Discovery and Potential Functional Characterization of Long Noncoding RNAs Associated with Familial Acne Inversa with NCSTN Mutation.

BACKGROUND: Long noncoding RNAs (lncRNAs) are associated with many dermatologic diseases. However, little is known about the regulatory function of lncRNAs in familial acne inversa (AI) patients with nicastrin (NCSTN) mutation. OBJECTIVES: The aim of this study was to explore the regulatory function of lncRNAs in familial AI patients with NCSTN mutation. METHODS: The expression profiles of lncRNAs and mRNAs in skin tissues from familial AI patients with NCSTN mutation and healthy individuals were analysed in this study via RNA sequencing (RNA-seq). RESULTS: In total, 359 lncRNAs and 1,863 mRNAs were differentially expressed between the two groups. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that the dysregulated mRNAs targeted by lncRNAs were mainly associated with the immune regulation, Staphylococcus aureus infection and B cell receptor signalling pathways. The lncRNA-miRNA-mRNA coexpression network contained 265 network pairs comprising 55 dysregulated lncRNAs, 11 miRNAs, and 74 mRNAs. Conservation analysis of the differentially expressed lncRNAs between familial AI patients with NCSTN mutation and Ncstn keratinocyte-specific knockout (NcstnΔKC) mice identified 6 lncRNAs with sequence conservation; these lncRNAs may participate in apoptosis, proliferation, and skin barrier function. CONCLUSIONS: These findings provide a direction for exploring the regulatory mechanisms underlying the progression of familial AI patients with NCSTN mutation.

Hsa_circ_0105040 promotes Cutbacterium acnes biofilm induced inflammation via sponge miR-146a in human keratinocyte.

Acne is a chronic inflammatory skin disease, and the pathogenesis of acne induced by Cutibacterium acnes (C.acnes) is not well understood. Recently, circular RNAs (circRNAs) have attracted much attention because of its involvement in various diseases. However, the mechanisms by which circRNAs regulated acne have rarely been reported. We identified several differentially expressed circRNAs by sequencing patient-derived acne tissues. Among them, hsa_circ_0105040 was determined to be low expressed in acne tissues and localized in the cytoplasm of human primary keratinocytes. We established a C.acnes biofilms model of acne in vitro and showed that hsa_circ_0105040 promoted inflammation via MAPK and NF-κB pathway. Mechanistically, hsa_circ_0105040 could directly bind to miR-146a and inhibit the expression of miR-146a. Moreover, hsa_circ_0105040 promoted the expression of IRAK1 and TRAF6 by sponging miR-146a, thereby elevating the level of inflammation in acne. Collectively, our data suggested that hsa_circ_0105040- miR-146a -IRAK1/TRAF6 axis was involved in regulating the inflammatory response in acne, which provided a potential therapeutic target for acne and a novel insight into the pathogenesis of inflammatory acne.

Antimicrobial peptide-producing dermal preadipocytes defend against Candida albicans skin infection via the FGFR-MEK-ERK pathway.

Dermal fibroblasts (dFBs) defend against deep bacterial skin infections by differentiating into preadipocytes (pAds) that produce the antimicrobial peptide cathelicidin; this differentiation is known as the dermal reactive adipogenesis response. However, the role of dFBs in fungal infection remains unknown. Here, we found that cathelicidin-producing pAds were present in high numbers in skin lesions from patients with cutaneous Candida granulomas. Second, we showed that dermal Candida albicans (C. albicans) infection in mice robustly triggered the dermal reactive adipogenesis response and induced cathelicidin expression, and inhibition of adipogenesis with pharmacological inhibitors of peroxisome proliferator-activated receptor γ (PPARγ) impaired skin resistance to C. albicans. In vitro, C. albicans products induced cathelicidin expression in pAds, and differentiating pAds markedly suppressed the growth of C. albicans by producing cathelicidin. Finally, we showed that C. albicans induced an antimicrobial response in pAds through the FGFR-MEK-ERK pathway. Together, our data reveal a previously unknown role of dFBs in the defense against skin infection caused by C. albicans.

YTHDC1-Modified m6A Methylation of Hsa_circ_0102678 Promotes Keratinocyte Inflammation Induced by Cutibacterium acnes Biofilm through Regulating miR-146a/TRAF6 and IRAK1 Axis.

INTRODUCTION: CircRNAs are closely related to many human diseases; however, their role in acne remains unclear. This study aimed to determine the role of hsa_circ_0102678 in regulating inflammation of acne. METHODS: First, microarray analysis was performed to study the expression of circRNAs in acne. Subsequently, RNase R digestion assay and fluorescence in situ hybridization assay were utilized to confirm the characteristics of hsa_circ_0102678. Finally, qRT-PCR, Western blotting analysis, immunoprecipitation, luciferase reporter assay, circRNA probe pull-down assay, biotin-labeled miRNA pull-down assay, RNA immunoprecipitation assay, and m6A dot blot assay were utilized to reveal the functional roles of hsa_circ_0102678 on inflammation induced by C. acnes biofilm in human primary keratinocytes. RESULTS: Our investigations showed that the expression of hsa_circ_0102678 was significantly decreased in acne tissues, and hsa_circ_0102678 was a type of circRNAs, which was mainly localized in the cytoplasm of primary human keratinocytes. Moreover, hsa_circ_0102678 remarkably affected the expression of IL-8, IL-6, and TNF-α, which induced by C. acnes biofilm. Importantly, mechanistic studies indicated that the YTHDC1 could bind directly to hsa_circ_0102678 and promote the export of N6-methyladenosine-modified hsa_circ_0102678 to the cytoplasm. Besides, hsa_circ_0102678 could bind to miR-146a and sponge miR-146a to promote the expression of IRAK1 and TRAF6. CONCLUSION: Our findings revealed a previously unknown process by which hsa_circ_0102678 promoted keratinocyte inflammation induced by C. acnes biofilm via regulating miR-146a/TRAF6 and IRAK1 axis.

Effect of 5-Aminolevulinic Acid Photodynamic Therapy on Aspergillus fumigatus Biofilms in Vitro.

Aspergillus fumigatus biofilm development results in enhanced pathogenicity and treatment resistance. Most contemporary antibiotics, however, are unable to eliminate biofilms. In recent years, with the application of new photosensitizers and the development of treatment, ALA-PDT (5-aminolevulinic acid photodynamic treatment) has achieved remarkable curative effect in the treatment of fungal infectious diseases; however, no research has been conducted on ALA-PDT against A. fumigatus. This study investigated the inhibitory effect of ALA-PDT at various 5-aminolevulinic acid concentrations and light doses on A. fumigatus planktonic and biofilms in vitro. We found that ALA-PDT may successfully inhibit the development of A. fumigatus biofilm and disintegrate mature biofilm. After ALA-PDT treatment, the adherence rate and vitality dramatically decreased, and the biofilm's structure was severely compromised. Our findings show for the first time that ALA-PDT may be used to prevent the formation of A. fumigatus biofilm and disturb the structure of mature biofilm, and that it could be employed as a therapeutic therapy for A. fumigatus superficial infection.

The immune function of dermal fibroblasts in skin defence against pathogens.

Dermal fibroblasts are the main resident cells of the dermis. They have several significant functions related to wound healing, extracellular matrix production and hair cycling. Dermal fibroblasts can also act as sentinels in defence against infection. They express pattern recognition receptors such as toll-like receptors to sense pathogen components, followed by the synthesis of pro-inflammatory cytokines (including IL-6, IFN-ß and TNF-α), chemokines (such as IL-8 and CXCL1) and antimicrobial peptides. Dermal fibroblasts also secrete other molecules-like growth factors and matrix metalloproteinases to benefit tissue repair from infection. Crosstalk between dermal fibroblasts and immune cells may amplify the immune response against infection. Moreover, the transition of a certain adipogenic fibroblasts to adipocytes protects skin from bacterial infection. Together, we discuss the role of dermal fibroblasts in the war against pathogens in this review. Dermal fibroblasts have important immune functions in anti-infection immunity, which should not be overlooked.

The impacts of non-farming income on rural household energy choices: Empirical evidence from China.

This study uses data from the China Family Panel Studies to analyze the possible impact of non-farming income on household energy choices. We use ordinary least squares and instrumental variable estimation methods to investigate the causal effect of non-farming income on household energy choices. We find that an increase in non-farming income assisted farmers in reducing their use of solid fuels in favor of clean energy. Our heterogeneity analysis, based on the average rural household income and geographical location of the village, shows that the energy upgrade effect of non-farming income is more obvious in high-income areas and suburbs closer to the county seat center. Further, we find that non-farming income has an impact on rural household energy choice mainly through the optimization of household energy-saving appliances and the enhancement of environmental awareness.

Low curcumin concentrations combined with blue light inhibits cutibacterium acnes biofilm-induced inflammatory response through suppressing MAPK and NF-κB in keratinocytes.

BACKGROUND: Curcumin has been employed as a photosensitizer agent during photodynamic therapy (PDT). Cutibacterium acnes (C. acnes) can cause an inflammatory response in human keratinocytes; however, no research has been conducted to determine whether curcumin and its photodynamic properties can prevent this inflammatory reaction. OBJECTIVE: We hypothesized that curcumin may control the C. acnes biofilm-induced inflammatory response in keratinocytes, either alone or in combination with blue light photodynamic therapy. METHODS: Following C. acnes biofilm stimulation, human primary keratinocytes were treated with 20 µM curcumin solution alone or 5 µM curcumin with combined blue light irradiation. The amount of secreted protein was measured using an ELISA kit. The expression levels of Toll-like receptor 2 (TLR2) and its downstream proteins were determined using western blot. RESULTS: Treatment with 20 µM curcumin, but not 5 µM curcumin, reduced the inflammatory response to C. acnes biofilms in keratinocytes by blocking the TLR2/MAPK/NF-κB pathway. Interestingly, 5 µM curcumin combined with blue light also reduced the C. acnes biofilm-induced inflammation indicated above by blocking the TLR2/MAPK/NF-κB pathway. CONCLUSION: Curcumin alone, in sufficient concentrations, or low-concentration curcumin with blue light had anti-inflammatory activity on keratinocytes stimulated by C. acnes biofilms through inhibition of MAPK and NF-κB signaling pathways by downregulating TLR2 expression.

Interaction of Amiodarone with Azoles Against Aspergillus Planktonic Cells and Biofilms in vitro.

Aspergillus spp. is the most common clinical pathogen of invasive fungal infection with high mortality. Existing treatments for Aspergillus spp. infection are still inefficient and accompanied by drug resistance, so it is still urgent to find new treatment approaches. The antiarrhythmic drug amiodarone (AMD) has demonstrated antifungal activity against a range of fungi. This study evaluated the efficacy of AMD in combination with triazoles for Aspergillus spp. infection. We tested the combined effect of AMD and three triazole drugs, namely, itraconazole (ITR), voriconazole (VRC), and posaconazole (POS), on the planktonic cells and biofilms of 20 strains of Aspergillus spp. via a checkerboard microdilution assay derived from 96-well plate-based method. Our results reveal that the combination of AMD with ITR or POS against Aspergillus biofilms has synergistic fungicidal effects. By contrast, the combination of AMD with VRC exhibits no antagonistic and synergistic effects. In this way, the use of AMD in combination with ITR or POS could be an effective adjunctive treatment for Aspergillus spp. infection.

GP IIb/IIIa-Mediated Platelet Activation and Its Modulation of the Immune Response of Monocytes Against Candida albicans.

Candida albicans is the most common fungal pathogen in humans, causing invasive disease and even potentially life-threatening systemic infections when tissue homeostasis is disrupted. Previous studies have identified an essential role of platelets in infection and immunity, especially when they are activated. However, it is still unclear whether platelets can be activated by C. albicans, and even less is known about the role of platelets in C. albicans infection. Herein, we showed that C. albicans induced platelet activation in vitro. C. albicans elevated the levels of AKT Ser473 phosphorylation, and inhibition of the PI3K-AKT signaling pathway reversed C. albicans-induced platelet activation. Surprisingly, C. albicans-induced platelet activation occurred in an integrin glycoprotein (GP) IIb/IIIa-dependent manner but was independent of the pattern recognition receptors toll-like receptor (TLR) 2 and TLR4. Interestingly, platelets enhanced the phagocytosis of human monocytes challenged with C. albicans and upregulated the expression of inflammatory cytokines, which were dependent on platelet activation mediated by GP IIb/IIIa. The present work provides new insights into the role of activated platelets in the defense against C. albicans, highlighting the importance of GP IIb/IIIa in the recognition of C. albicans.

MiR-146a Negatively Regulates Aspergillus fumigatus-Induced TNF-α and IL-6 Secretion in THP-1 Macrophages.

Aspergillus fumigatu (A. fumigatus) is one of the most common important fungal pathogens that cause life-threatening infectious disease in immunocompromised individuals. However, the host immune response against this pathogenic mold is not fully understood. MicroRNAs (miRNAs) play essential roles in regulating innate immunity. Thus, we investigated the function of miR-146a in inflammatory responses in macrophages after A. fumigatus stimulation in this study. We found that TNF-α and IL-6 were increased in THP-1 macrophage-like cells treated with A. fumigatus at both the mRNA and protein levels. The interaction between THP-1 macrophage-like cells and A. fumigatus resulted in a long-lasting increase in miR-146a expression dependent on p38 MAPK and NF-κB signaling. In A. fumigatus-challenged THP-1 macrophage-like cells, overexpression of miR-146a by miR-146a mimics decreased TNF-α and IL-6 production, whereas downregulation of miR-146a by anti-miR-146a significantly enhanced the level of TNF-α and IL-6. Our study demonstrates that the crosstalk between miR-146a and the inflammation-regulating p38 MAPK and NF-κB pathways might be a fine-tuning mechanism in the modulation of the inflammatory response in macrophages infected with A. fumigatus. Our findings illuminate the crucial role of miR-146a in the pathogenesis of human diseases associated with A. fumigatus infection.

Gene expression profile and long noncoding RNA analysis in Candida albicans insoluble ß-glucan-stimulated CD14+ monocytes and THP-1 cells.

Emerging evidence suggests that long noncoding RNAs (lncRNAs) play important roles in disease development. However, the roles of lncRNAs in the pathogenesis of Candida albicans (C. albicans) remain unclear. Our study aimed to investigate and characterize the mRNA and lncRNA transcriptomes of CD14+ monocytes and THP-1 cells stimulated with insoluble ß-glucan by RNA-seq. We identified a total of 10788 differentially expressed (DE) mRNAs and 2021 DE lncRNAs in CD14+ monocytes, while 3349 DE mRNAs and 291 DE lncRNAs were observed in THP-1 cells. A total of 808 DE mRNAs and 51 DE lncRNAs overlapped between the two groups. We examined five collectively DE mRNAs and lncRNAs in both cells using quantitative real-time PCR, validating the reliability of the RNA-seq results. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that the 808 DE mRNAs were mostly enriched in the inflammatory response and NF-kappa B signaling pathway, respectively. Next, lncRNA-mRNA coexpression analysis was performed for the 51 DE lncRNAs and the 808 DE mRNAs in the two groups. We chose the common network pairs of the two groups to construct the coexpression network and revealed 97 network pairs comprising 8 dysregulated lncRNAs and 60 dysregulated mRNAs. We found that lncRNA lnc-CCL3L3-1:1 might be involved in the NF-kappa B signaling pathway in C. albicans infection. In conclusion, the aberrantly expressed lncRNAs might play a role in the pathogenesis of C. albicans infection and could be used as therapeutic targets in the future.

[Effect of 5-aminolevulinic Acid Photodynamics Therapy on Biofilm of Propionibacterium Acnes].

Objective To investigate the effect of 5-aminolevulinic acid photodynamic therapy (ALA-PDT) on Propionibacterium acnes (P.acnes) biofilm. Methods P.acnes biofilms were constructed on a cell slide and treated with ALA-PDT.According to different light doses,the biofilms were divided into six groups:ALA-PDT group [ALA-PDT1 (50 J/cm2),ALA-PDT2 group (100 J/cm2),ALA-PDT3 group (200 J/cm2)],ALA-only group (ALA group),light-only group (LED),and a negative control group (ALA-PDT-group).The biofilm structure and the ratio of the dead bacteria/live bacteria were observed using a laser confocal microscope (CLSM).Biofilm viability was measured using the XTT assay. Results CLSM showed that the biofilm structures of ALA group and LED group were not significantly different from that of ALA-PDT-group,whereas the biofilm structure was more seriously damaged in ALA-PDT1 group,ALA-PDT2 group,and ALA-PDT3 group than in the ALA-PDT-group.The ratios of the dead/live bacteria in ALA-PDT-group,ALA group,LED group,ALA-PDT1 group,ALA-PDT2 group,and ALA-PDT3 group were 0.350±0.033, 0.305±0.046, 0.330±0.032, 1.525±0.439, 2.293±0.148 and 3.092±0.189,respectively.ALA group(md=0.003, P=1.000)and LED group(md=-0.025, P=1.000)did not significantly differ from the ALA-PDT-group.However,the ratio of dead/live bacteria in ALA-PDT-group was significantly lower than those in ALA-PDT1 group (md=-0.162, P<0.001),ALA-PDT2 group (md=-0.254, P<0.001),and ALA-PDT3 group (md=-0.352, P<0.001).The values of the XTT assay were were 0.462±0.028,0.465±0.044,0.437±0.047,0.301±0.040,0.207±0.001,and 0.110±0.007,respectively,in ALA-PDT-group,ALA group,LED group,ALA-PDT1 group,ALA-PDT2 group,and ALA-PDT3 group.Although the values of XTT assay in ALA(md=-0.044, P=1.000)and LED groups (md=-0.020, P=1.000)did not significantly differ from that in ALA-PDT-group,it was significantly higher in ALA-PDT-group than in ALA-PDT1 group (md=1.175, P<0.001),ALA-PDT2 group (md=1.942, P<0.001),and ALA-PDT3 group (md=-0.352, md=2.742, P<0.001). Conclusions ALA-PDT has an inhibitory effect on P.acnes biofilm.ALA-PDT destroys biofilm structure and inhibits biofilm viability.

miR-146a Inhibits Biofilm-Derived Cutibacterium acnes-Induced Inflammatory Reactions in Human Keratinocytes.

Acne is a chronic inflammatory skin disorder that often involves the formation of Cutibacterium acnes (C. acnes) biofilms. Several microRNAs (miRNAs) are known to be involved in inflammatory responses. However, it is unknown whether miRNAs play a role in the inflammatory reaction triggered by C. acnes biofilm. In this study, we investigated the role of miR-146a in biofilm-derived C. acnes-induced inflammatory responses. Increased expressions of miR-146a and toll-like receptor (TLR) 2 were detected in acne lesions. In the presence of biofilm-derived C. acnes, TLR2 and its downstream NF-kB and MAPK pathways were activated in keratinocytes. Subsequently, miR-146a was upregulated in these cells along with the induction of IL-6, IL-8, and tumor necrosis factor (TNF)-α. Furthermore, our data indicates that miR-146a could directly bind the 3'-untranslated region of IRAK1 and TNF receptor-associated factor 6 (TRAF6) and suppress their expression, leading to an inhibition of biofilm-derived C. acnes-induced activation of NF-kB, p38, and ERK1/2 pathways. Overall, our results indicate that biofilm-derived C. acnes induces miR-146a, which can downregulate the production of IL-6, -8, and TNF-α in acne inflammation by inhibiting the TLR2/IRAK1/TRAF6/NF-κB and MAPK pathways.

Candida tropicalis induces pro-inflammatory cytokine production, NF-κB and MAPKs pathways regulation, and dectin-1 activation.

The prevalence of Candida infection induced by non-albicans Candida (NAC) species is increasing. However, as a common NAC species, C. tropicalis has received much less study in terms of host immunity than C. albicans has. In this study, we evaluated the pro-inflammatory cytokine responses evoked by C. tropicalis and determined whether dectin-1 and downstream NF-κB and mitogen-activated protein kinases (MAPKs) signaling pathways played roles in inflammation in human peripheral blood mononuclear cells (PBMCs) and THP-1 macrophage-like cells. Exposure of PBMCs and THP-1 macrophage-like cells to C. tropicalis led to the enhanced gene expression and secretion of TNF-α and IL-6 in a time- and dose-dependent manner. THP-1 macrophage-like cells being challenged by C. tropicalis resulted in the activation of the NF-κB, p38, and ERK1/2 MAPK signaling pathways. We also found that the expression of dectin-1 was increased with C. tropicalis treatment. These data reveal that dectin-1 may play a role in sensing the inflammation response induced by C. tropicalis and that NF-κB and MAPK are involved in the downstream signaling pathways in macrophages.

Phagocytosis of Candida albicans Inhibits Autophagic Flux in Macrophages.

Autophagy machinery has roles in the defense against microorganisms such as Candida albicans. Lipidated LC3, the marker protein of autophagy, participates in the elimination of C. albicans by forming a single-membrane phagosome; this process is called LC3-associated phagocytosis (LAP). However, the influence of C. albicans on autophagic flux is not clear. In this study, we found that C. albicans inhibited LC3 turnover in macrophages. After the phagocytosis of C. albicans in macrophages, we observed fewer acridine orange-positive vacuoles and RFP-GFP-LC3 puncta without colocalization with phagocytized C. albicans. However, phagocytosis of C. albicans led to LC3 recruitment, but p62 and ATG9A did not colocalize with LC3 or C. albicans. These effects are due to an MTOR-independent pathway. Nevertheless, we found that the C. albicans pattern-associated molecular pattern ß-glucan increased LC3 turnover. In addition, phagocytosis of C. albicans caused a decrease in BrdU incorporation. Blocking autophagic flux aggravated this effect. Our findings suggest that phagocytosis of C. albicans decreases autophagic flux but induces LAP in an MTOR-independent manner in macrophages. Occupation of LC3 by recruiting engulfed C. albicans might contribute to the inhibition of autophagic flux. Our study highlights the coordinated machinery between canonical autophagy and LAP that defends against C. albicans challenge.

Inflammation Induced by Candida parapsilosis in THP-1 Cells and Human Peripheral Blood Mononuclear Cells (PBMCs).

Candida parapsilosis is one of the most prevalent Candida species; however, the inflammation response induced by C. parapsilosis and related mechanism received few studies. In this study, we analyzed the pro-inflammatory cytokine responses evoked by C. parapsilosis in human peripheral blood mononuclear cells (PBMCs) and THP-1 cells, determined the signal pathways related to the inflammation response and investigated the expression of dectin-1 modified with C. parapsilosis. Exposure of PBMCs and THP-1 cells to C. parapsilosis led to the increased gene expression and production of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). C. parapsilosis induced TNF-α and IL-6 release in a time- and dose-dependent manner. Western blotting was used to analyze p38, ERK1/2 mitogen-activated protein kinases (MAPKs) and IκB-α phosphorylation and degradation. Nuclear translocation of NF-κB was detected by confocal microscopy. THP-1 cells challenged by C. parapsilosis resulted in the activation of NF-κB and phosphorylation of p38 and ERK1/2 MAPKs. The expression of dectin-1 was up-regulated after the stimulation of C. parapsilosis. Our results suggest that C. parapsilosis could stimulate the inflammatory response, increase the expression of dectin-1 and activate NF-κB and MAPKs signaling pathways in macrophages.

MiR-146a negatively regulates dectin-1-induced inflammatory responses.

Dectin-1 is the critical sensor for ß-glucan from Candida which is the most common human fungal pathogen and cause superficial and system infection. MicroRNAs (miRNAs) play crucial roles in regulating innate immunity. However, the functional role of miRNAs in inflammatory response dependent on the activation of dectin-1 pathway has not been defined. In the present study, we found insoluble ß-glucan from the cell wall of Candida albicans (CaIG) was able to increase the production of of IL-6 and TNFα through Dectin-1-Syk-NF-κB and p38MAPK pathway. MiRNAs profiles combined with real-time PCR validation revealed that miR-146a, miR-30-5p, miR-210-3p expression level were increased in THP-1 cells treated with CaIG. The interaction between Dectin-1 and CaIG resulted in an long lasting increase of miR-146a expression dependent on Dectin-1-Syk-NF-κB, p38MAPK, contrasting with a rapid and transient increase of IL-6 and TNFα. Overexpression of miR-146a significantly suppressed the production of IL-6 and TNFα. MiR-146a mimics inhibited CaIG-induced activity of p-IκBα and translocation of NF-κB p65. Luciferase reporter assays showed miR-146a inhibited NF-κB promoter-binding activity. Together, our data suggest miR-146a may play the potent negative feedback regulator in inflammatory response following Dectin-1 stimulation.