Microbiome Profile in Patients with Adult Balanoposthitis: Relationship with Redundant Prepuce, Genital Mucosa Physical Barrier Status and Inflammation.

Balanoposthitis is a common inflammatory condition of male genitalia, while the overall microbiota spectrum and its relevance to contributing factors have yet to be determined. This case-control study included patients with balanoposthitis (n = 26) and matched healthy controls (n = 29), both uncircumcised. Overt fungal infection in balanoposthitis was excluded, swab samples were collected, 16S rRNA gene sequenced and analysed. The profile of the microbiome was further analysed in relation to the clinical severity of the disease and the physical barrier status of the glans penis, including mucosa pH, transepidermal water loss, and mucosa hydration. In general, the microbiota composition was similar between patients with balanoposthitis and healthy controls, while it was different between patients with balanoposthitis and healthy controls with redundant prepuce. Decreased hydration of the mucosa and increased pH were found in patients with balanoposthitis. Staphylococcus warneri and Prevotella bivia are the 2 most abundant balanoposthitis-associated species and are positively correlated with disease severity.

Vibrio cholerae Virulence Activator ToxR Regulates Manganese Transport and Resistance to Reactive Oxygen Species.

Like many other pathogens, Vibrio cholerae, the causative agent of cholera, can modulate its gene expression to combat stresses encountered in both aquatic and host environments, including stress posed by reactive oxygen species (ROS). We previously reported that the virulence activator AphB in V. cholerae is involved in ROS resistance. In this study, we found that another key virulence regulator, ToxR, was important for V. cholerae resistance to hydrogen peroxide. Through a genome-wide transposon screen, we discovered that a deletion in mneA, which encodes a manganese exporter, restored ROS resistance of the toxR mutant. We then showed that ToxR did not affect mneA transcription but that the ToxR-regulated major porin OmpU was critical for ROS resistance. The addition of manganese in culture medium restored ROS resistance in both the toxR and ompU mutants. Furthermore, elemental analysis indicated that the intracellular concentration of manganese in both the toxR and ompU mutants was reduced. This may result in intracellular ROS accumulation in these mutants. Our data suggest that ToxR plays an important role in the resistance to reactive oxygen species through the regulation of manganese transport.

Cell wall mannoprotein of Candida albicans polarizes macrophages and affects proliferation and apoptosis through activation of the Akt signal pathway.

Candida albicans is a commensal fungus that associates with human hosts. Under normal circumstances this interaction does not produce any severe life-threatening disease, as macrophages of the innate immune system will result in its clearance. However, disorders may arise in immunosuppressed individuals. To understand the bioactivity of Candida albicans cell wall polysaccharides, which represent an important component of its function, mannoprotein from this fungus was extracted, purified and analyzed. Mannoprotein with α-(1,2) and α-(1,6) linkages was investigated with use of HPLC and NMR. Co-incubation of mannoprotein with macrophages resulted in a mannoprotein with the potential to polarize macrophages to M1 and promote phagocytosis/microbial killing ability thus increasing the clearance of pathogens through Akt2. Moreover, mannoprotein within the cell wall promoted cell proliferation and inhibited apoptosis by activation of the Akt signaling pathway. Collectively, α-(1,6)(1,2)-mannoprotein, one of the five polysaccharides extracted from the cell wall of Candida albicans, demonstrates immune-enhancing effects by activation of the Akt signaling pathway. These findings provide important new insights into the biological effects of polysaccharides on macrophages. Such information can then serve as the foundation for the development of novel anti-fungal medications.

Extracellular vesicles derived from Malassezia furfur stimulate IL-6 production in keratinocytes as demonstrated in in vitro and in vivo models.

BACKGROUND: Malassezia is one of the commensal microorganisms colonized on human skin and has been shown to be related to several inflammatory cutaneous disorders. Previous studies indicated that Malassezia. sympodialis (M. sympodialis) can produce extracellular vesicles, however, the immunoregulatory function of Malassezia extracellular vesicles on keratinocytes has not been studied. OBJECTIVE: To investigate the extracellular vesicular production capability of Malassezia. furfur (M. furfur) and examine their immunoregulatory effects both in vitro and in vivo. METHODS: Extracellular vesicles derived from M. furfur were isolated by sequential ultracentrifugation procedure. Their structure and diameter were determined by negative stain TEM and NTA, respectively. Confocal microscopy was used to visualize the internalization of these nanoparticles into HaCaT cells and mice epidermal keratinocytes. The expressions of inflammatory cytokines were screened using PCR Array assay and validated in vitro by qPCR and ELISA assays. In vivo cytokine production was measured by the IHC method. The role of NF-κB in such process was evaluated in HaCaT cells by western blot assay. RESULTS: Our results showed that M. furfur produced ovoid-shaped nanoparticles, which could be then internalized into HaCaT cells, as well as mice epidermal keratinocytes. IL-6 expression was significantly enhanced in response to extracellular vesicular stimulation both in vitro and in vivo, in which process the activation of NF-κB was involved. CONCLUSION: M. furfur has the ability to release extracellular vesicles, which can be internalized into keratinocytes and promote the production of IL-6 with the involvement of NF-κB dependent pathway. Such findings reveal some important new insights into Malassezia pathogenesis and therapy.

Proteomic and bioinformatic analysis of condyloma acuminata: mild hyperthermia treatment reveals compromised HPV infectivity of keratinocytes via regulation of metabolism, differentiation and anti-viral responses.

BACKGROUND: Hyperthermia has proved successful in treating cutaneous human papillomavirus infectious diseases such as plantar wart and condyloma acuminata (CA). Moreover, this treatment provides improved therapeutic efficacy in these conditions as compared with conventional therapies. OBJECTIVES: To investigate the global proteome changes in CA in response to hyperthermia and achieve a better understanding of the mechanisms of hyperthermia therapy against HPV-infectious diseases. METHODS: CA tissue was obtained from patients undergoing pathological examinations. Diagnosis was verified as based on results of both HE staining and HPV-DNA PCR assay. Hyperthermia was achieved with a 44 °C water bath. Differentially expressed proteins (DEPs) were identified by iTRAQ labeling, SCX chromatography and LC-MS/MS assay. Validation of proteomic results was performed using real-time qPCR and western blot, while bioinformatic analysis of DEPs was accomplished by R 3.4.1, STRING and Cytoscape softwares. RESULTS: In response to hyperthermia, a total of 102 DEPs were identified with 37 being upregulated and 65 downregulated. Among these DEPs, hyperthermia induced proteins involved with anti-viral processes such as OAS1, MX1, BANF1, CANX and AP1S1, whereas it inhibited proteins that participated in cellular metabolism, such as GALT, H6PD, EXOSC4 and EXOSC6; protein translation, such as RPS4Y1; as well as keratinocyte differentiation, such as KRT5, KRT27, KRT75, KRT76 and H2AFY2. CONCLUSIONS: Hyperthermia inhibited enzymes and molecules responsible for metabolism modulation and keratinocyte differentiation in CA tissue, whereas it promoted factors involved in anti-viral responses. Such effects may, in part, contribute to the efficacy of local hyperthermia therapy against HPV infection.

Screening and identification of differentially expressed serum proteins in patients with vitiligo using two­dimensional gel electrophoresis coupled with mass spectrometry.

In the clinic, vitiligo is characterized by two stages: Stable and progressive. The pathogenesis of vitiligo is still not clear. Here, we identified serum markers of vitiligo by screening for differentially expressed proteins in patients with vitiligo compared to healthy individuals. Serum samples were collected from patients with vitiligo (n=10 for both the stable and progressive stages) and healthy individuals (n=10). Two­dimensional gel electrophoresis followed by matrix­assisted laser desorption/ionization time­of­flight mass spectrometry and western blotting were used to validate the differential expression of the proteins in the serum (n=20 each, at both stages for patients and healthy individuals). A total of 48 differentially expressed proteins were identified by gel image analysis. There were 28 differentially expressed proteins in patients with progressive vitiligo (PV) and 13 differentially expressed proteins in patients with stable vitiligo (SV) compared with that in healthy individuals. Additionally, 7 differentially expressed proteins were identified in patients with PV compared with those in patients with SV. The western blotting results showed that Peroxiredoxin­6, apolipoprotein L1, apolipoprotein E and mannose­binding protein were differentially expressed in patients with different stages of vitiligo. Our results showed that change serum levels of several proteins might be useful as biomarkers or in understanding the pathogenesis of vitiligo.

Cell wall mannoprotein of Candida albicans induces cell cycle alternation and inhibits apoptosis of HaCaT cells via NF-κB signal pathway.

Candida albicans (C. albicans) is a commensal organism in human and a well-known dimorphic opportunistic pathogenic fungus. Though plenty of researches on the pathogenesis of C. albicans have been performed, the mechanism is not fully understood. The cell wall components of C. albicans have been documented to play important roles in its pathogenic processes. To further study the infectious mechanism of C. albicans, we investigated the potential functional role of its cell wall mannoprotein in cell cycle and apoptosis of HaCaT cells. We found that mannoprotein could promote the transition of cell cycle from G1/G0 to S phase, in which Cyclin D1, CDK4 and p-Rb, the major regulators of the cell cycle progression, showed significant upregulation, and CDKN1A (cyclin dependent kinase inhibitor 1A (p21)) showed significant downregulation. Mannoprotein also could inhibit apoptosis of HaCaT cells, which was well associated with increased expression of BCL2 (Bcl-2). Moreover, mannoprotein could increase the phosphorylation levels of RELA (p65) and NFKBIA (IκBα), as the key factors of NF-κB signal pathway in HaCaT cells, suggesting the activation of NF-κB signal pathway. Additionally, a NF-κB specific inhibitor, PDTC, could rescue the effect of mannoprotein on cell cycle and apoptosis of HaCaT cells, which suggested that mannoprotein could activate NF-κB signal pathway to mediate cell cycle alternation and inhibit apoptosis.