From mice to men: An assessment of preclinical model systems for the study of vitiligo.

Vitiligo is an autoimmune skin disease of multiple etiology, for which there is no complete cure. This chronic depigmentation is characterized by epidermal melanocyte loss, and causes disfigurement and significant psychosocial distress. Mouse models have been extensively employed to further our understanding of complex disease mechanisms in vitiligo, as well as to provide a preclinical platform for clinical interventional research on potential treatment strategies in humans. The current mouse models can be categorized into three groups: spontaneous mouse models, induced mouse models, and transgenic mice. Despite their limitations, these models allow us to understand the pathology processes of vitiligo at molecule, cell, tissue, organ, and system levels, and have been used to test prospective drugs. In this review, we comprehensively evaluate existing murine systems of vitiligo and elucidate their respective characteristics, aiming to offer a panorama for researchers to select the appropriate mouse models for their study.

Serum cytokine profiles predict response to systemic glucocorticoid in active vitiligo.

Introduction: Vitiligo is an immune-related skin disease. Cytokines regulate immune response and inflammation and are involved in the pathogenesis of vitiligo. Aim: To assess the serum levels of pro-inflammatory cytokines pre- and post- systemic glucocorticoid treatment in patients with active vitiligo. Material and methods: We measured serum cytokine levels using the enzyme-linked immunosorbent assay in 31 patients with active vitiligo before and after treatment. All patients received systemic glucocorticoid (compound betamethasone injection) in combination with topical halometasone cream and tacrolimus ointment for 3 months. Twenty healthy controls were also examined. The cytokines measured included TNF-α, IL-1ß, IL-6, IFN-γ, IL-2, IL-17, IL-10, IL-8, and CXCL10. Results: The serum levels of TNF-α, IL-1ß, IL-6, IFN-γ, IL-2, IL-17, IL-8, and CXCL10 were significantly higher, and levels of IL-10 were lower in vitiligo patients compared to controls. Additionally, serum IFN-γ (r = 0.378; p = 0.036), IL-17 (r = 0.426; p = 0.017), and CXCL10 (r = 0.514; p = 0.003) showed a positive correlation with affected body surface area in vitiligo patients. After 3 months of systemic glucocorticoid treatment, the levels of IL-1ß, IFN-γ, IL-2, IL-17, and CXCL10 in responders were significantly decreased and nearly restored to normal levels. The IL-10 level was also increased in response to treatment. In contrast, the non-responder group had persistently high IL-6, IL-17, IL-8, and CXCL10 levels, and negligible changes in TNF-α, IL-1ß, IFN-γ, IL-2, and IL-10. Conclusions: Our study indicated that the levels of inflammatory cytokines were significantly ameliorated in the glucocorticoid responder group. Altered cell-mediated immunity may contribute to the resistance in vitiligo. The cytokines such as TNF-α, IL-1ß, IFN-γ and IL-2 could serve as therapeutic targets for managing glucocorticoid-resistant vitiligo.

Influences of vitiligo-associated characteristics on the occurrence of diabetes mellitus: Interactive analysis of a cross-sectional study.

The risk of diabetes mellitus (DM) in vitiligo patients is higher than that in non-vitiligo population. Our goal was to explore the influencing factors for DM in vitiligo patients. A matched-pair design of 107 cases with DM and 428 controls without DM was conducted among vitiligo patients in Xijing hospital from January 2010 to October 2021. The baseline characteristics of patients were analysed based on standard descriptive statistics. The vitiligo-associated characteristics were analysed by logistic regression to identify influencing factors of DM. Interaction analysis was performed to explore the additive interactions between vitiligo-associated characteristics and baseline characteristics. After adjustment for the baseline characteristics, the severity of vitiligo [odds ratio (OR) = 2.47, 95% confidence interval (CI): 1.47-4.14] and onset age of vitiligo (OR = 0.98, 95% CI: 0.97-0.99) had a significant correlation with occurrence of DM. The severity of vitiligo had additive interaction with family history of diabetes [relative excess risk due to interaction (RERI) = 132.51 (95% CI: 5.51-1100.20), attributable proportion (AP) = 0.91 (95% CI: 0.17-0.95), synergy index (S) = 11.53 (95% CI: 1.32-100.5)] and with smoking history [RERI = 6.54 (95% CI: 0.67-19.83), AP = 0.64 (95% CI: 0.04-0.80), S = 3.48 (95% CI: 1.17-10.36)]. Earlier onset age of vitiligo and greater BSA involvement might be two independent risk factors for DM in vitiligo patients. Interaction assessment identified the severity of vitiligo as additive interaction factors with diabetes family history and with smoking history for the DM occurrence.

Th1-like Treg in vitiligo: An incompetent regulator in immune tolerance.

Vitiligo is an autoimmune skin disease resulting from epidermal melanocyte destruction mediated by CD8+T cells that breach the self-tolerance. Regulatory T cells (Tregs) are critical for keeping the CD8+T cells in check, but the deficiency of Tregs leading to the immune disequilibrium in vitiligo remains undefined. In the present study, we used RNA-sequencing (RNA-seq) to acquire the transcriptome data of Tregs from vitiligo patients and healthy controls, respectively. Further flow cytometry analysis and immunofluorescence assays substantiated the phenotype of Th1-like Tregs in vitiligo. CD8+T cell-/vitiligo serum-Treg co-culture assays and chemotaxis assays were used to functionally examine this subset of Tregs. As a result, RNA-seq, flow cytometry, and immunofluorescence all indicated the transition of bona fide Treg to the Th1-like T-bet+IFN-γ+Treg in vitiligo patients. Besides, these Th1-like Tregs exhibited significantly dampened suppression on the proliferation and activation of CD8+T cells and a markedly higher tendency to be chemoattracted by CXCL10 and CXCL16. More interestingly, vitiligo serum could even elicit bona fide Tregs of healthy controls to adopt the Th1-like phenotype and manifest impaired suppression. To conclude, Tregs from vitiligo patients are functionally disturbed and the Th1-skewed inflammatory microenvironment in the serum of vitiligo patients is responsible for the generation of Th1-like Tregs. We provide a clinical exploitable strategy that in addition to simply replenishing the bona fide Treg or promoting the homing of Treg to the skin, the normalization of the Th1-skewed inflammatory environment in vitiligo patients and targeting the incompetent Th1-like Tregs might be critical in the future treatment of vitiligo.

P53/PANK1/miR-107 signalling pathway spans the gap between metabolic reprogramming and insulin resistance induced by high-fat diet.

High-fat diet (HFD) leads to obesity, type II diabetes mellitus (T2DM) and increases the coincidence of cardiovascular diseases and cancer. Insulin resistance (IR) is considered as the 'common soil' of those diseases. Furthermore, people on HFD showed restrained glycolysis and enhanced fatty acid oxidation, which is the so-called metabolic reprogramming. However, the relationship between metabolic reprogramming and IR induced by HFD is still unclear. Here, we demonstrate that PANK1 and miR-107 were up-regulated in the liver tissue of mice on HFD for 16 weeks and involved in metabolic reprogramming induced by palmitate acid (PA) incubation. Importantly, miR-107 within an intron of PANK1 gene facilitated IR by targeting caveolin-1 in AML12 cells upon PA incubation. Moreover, we identify that HFD enhanced P53 expression, and activation of P53 with nutlin-3a induced PANK1 and miR-107 expression simultaneously in transcriptional level, leading to metabolic reprogramming and IR, respectively. Consistently, inhibition of P53 with pifithrin-α hydrobromide ameliorated PA-induced metabolic reprogramming and IR. Thus, our results revealing a new mechanism by which P53 regulate metabolism. In addition, the results distinguished the different roles of PANK1 and its intron miR-107 in metabolic regulation, which will provide more accurate intervention targets for the treatment of metabolic diseases.

miR-141-3p Promotes the Cisplatin Sensitivity of Osteosarcoma Cell through Targeting the Glutaminase [GLS]-Mediated Glutamine Metabolism.

AIMS: This study aimed to evaluate the roles and molecular targets of miRNA-141-3p in the cisplatin sensitivity of osteosarcoma. BACKGROUND: Osteosarcoma is one of the most common-type bone tumors, occurring mainly in children and adolescents. Cancer cells display dysregulated cellular metabolism, such as the abnormally elevated glutamine metabolism. OBJECTIVE: Non-coding RNA miRNA-141-3p has been reported to act as a tumor suppressor in osteosarcoma. Currently, the precise molecular mechanisms for the miR- 141-3p-mediated chemosensitivity through regulating glutamine metabolism remain unclear. METHODS: We collected thirty paired OS tumors and their adjacent normal tissues. The osteosarcoma cell lines [Saos-2] and normal osteoblast cells, hFOB1.19, were used for in vitro experiments. RT-qPCR and Western blot were applied for gene expression detections. Targets of miR-141-3p were predicted from starBase. The MTT and flow cytometric assays were performed to determine cell growth and apoptosis rates. The cellular glutamine metabolism was monitored by glutamine uptake assay and the glutaminase [GLS] activity assay. RESULTS: We reported that miR-141-3p were significantly downregulated in osteosarcoma tissues and cells. Overexpression of miR-141-3p suppressed OS cell growth and sensitized OS cells to cisplatin. In addition, glutamine metabolism was significantly increased in osteosarcoma. We characterized that GLS played oncogenic roles in osteosarcoma and validated GLS was a direct target of miR-141-3p in OS cells. Rescue experiments consistently demonstrated that miR-141-3p-promoted cisplatin sensitivity was achieved by targeting GLS directly. CONCLUSION: Overall, our findings revealed new molecular mechanisms of the miR-141- 3p-modulated cisplatin sensitization through targeting the GLS-glutamine metabolism pathway. This study will contribute to developing new therapeutic approaches for the treatments of chemoresistant osteosarcoma.

Management of the refractory vitiligo patient: current therapeutic strategies and future options.

Vitiligo is an autoimmune disease that leads to disfiguring depigmented lesions of skin and mucosa. Although effective treatments are available for vitiligo, there are still some patients with poor responses to conventional treatment. Refractory vitiligo lesions are mostly located on exposed sites such as acral sites and lips, leading to significant life stress. Understanding the causes of refractory vitiligo and developing targeted treatments are essential to enhance vitiligo outcomes. In this review, we summarized recent treatment approaches and some potential methods for refractory vitiligo. Janus kinase inhibitors have shown efficacy in refractory vitiligo. A variety of surgical interventions and fractional carbon dioxide laser have been widely applied to combination therapies. Furthermore, melanocyte regeneration and activation therapies are potentially effective strategies. Patients with refractory vitiligo should be referred to psychological monitoring and interventions to reduce the potential pathogenic effects of chronic stress. Finally, methods for depigmentation and camouflage may be beneficial in achieving uniform skin color and improved quality of life. Our ultimate focus is to provide alternative options for refractory vitiligo and to bring inspiration to future research.

Using Ex Situ Seedling Baiting to Capture Seedling-Associated Mycorrhizal Fungi in Medicinal Orchid Dendrobium officinale.

Using orchid mycorrhizal fungi (OMFs) to facilitate orchid proliferation is considered an effective method of orchid conservation. Based on the success of using in situ seedling baiting to obtain plant growth-promoting fungi in our previous study, in this study, we developed the method of using ex situ seedling baiting to capture seedling-associated fungi from Dendrobium officinale. We collected substrates (e.g., litters, barks and mosses) from six original habitats of D. officinale in different geographical locations in China, and then, transplanted in vitro-produced seedlings of D. officinale into the substrates. After cultivation for 75 days, it was obvious that fungi colonized the seedling roots and formed large numbers of pelotons in all six groups. From these seedling roots, a total of 251 fungal strains, which were divided into 16 OMF and 11 non-OMF species, were successfully isolated. The 16 OMFs included 13 Tulasnella and 3 Serendipitaceae species. The fungal species isolated from the different groups (original habitat sources) were not identical, but the dominant OMFs with high isolation frequencies (more than 10 times) were commonly isolated from more than four original sources. Among the 11 non-OMFs, Fusarium oxysporum TP-18 and Muscodor sp. TP-26 were the dominant endophytes. Fusarium oxysporum is a common endophyte associated with many orchid species, including D. officinale. The results suggest that ex situ seedling baiting is an easy and efficient approach to obtaining seedling-associated fungi for this species and could be performed for other over-collected species, especially orchids for which wild plants have disappeared in the field but their original habitats are known. This approach has great potential for application in OMF studies in the future.

Circulatory levels of alarmins in patients with non-segmental vitiligo: Potential biomarkers for disease diagnosis and activity/severity assessment.

Background: Non-segmental vitiligo (NSV) is an autoimmune skin disorder that is difficult to determine disease activity/severity and thus to treat. Alarmins have emerged as promising biomarkers in various diseases, so further confirmation of their potential roles in NSV would be of considerable value. With the present work, we aimed to determine the serum levels of alarmins in patients with NSV, correlate these alarmins with disease activity and severity, and analyze the predictive value of the combination of these markers. Methods: 104 NSV patients and 56 healthy controls were enrolled at the Xijing Hospital of Fourth Military Medical University between September 1, 2018, and June 30, 2019. The serum levels of alarmins (including IL-33, IL-1α, S100A9, S100A12, S100B, and HMGB1) were measured with enzyme-linked immunosorbent assays. The predictive performance of these biomarkers was evaluated with the area under the receiver operating characteristic curve (AUC), sensitivity, specificity, and other representative statistics. Results: A total of 104 patients with NSV (mean [SD] age, 34.2 [13.0] years; 62 [59.6%] male) and 56 healthy controls (mean [SD] age, 34.8 [13.5] years; 34 [60.7%] male) were enrolled. For vitiligo diagnosis, S100B had the highest sensitivity (92.31%), whereas HMGB1 had the highest specificity (85.71%); the combination of IL-1α, S100B, S100A9, and HMGB1 increased the AUC value to 0.925, with a sensitivity of 87.50% and a specificity of 85.71%. Multivariate logistic regression analysis showed S100B (OR, 1.019; 95% CI, 1.002-1.038; P =0.03), S100A9 (OR, 1.002; 95% CI, 1.001-1.003; P<0.001), and HMGB1 (OR, 1.915; 95% CI, 1.186-3.091; P =0.008) were significantly associated with vitiligo activity. S100A9 had the highest accuracy in discriminating patients at the active stage from the stable stage, with an AUC value of 0.827. The combination of these alarmins had an AUC value of 0.860 to assess disease activity, with a sensitivity of 90.00% and a specificity of 72.97%. Furthermore, S100B (r=0.61, P <0.001), S100A9 (r=0.33, P <0.001), and HMGB1 (r = 0.51, P <0.001) levels were positively correlated with the affected body surface area (BSA) in NSV patients. Conclusions: Serum S100B, S100A9, and HMGB1 might be biomarkers for diagnosing and assessing the activity/severity of NSV, either used alone or in combination.

The Regulatory Network and Potential Role of LINC00973-miRNA-mRNA ceRNA in the Progression of Non-Small-Cell Lung Cancer.

Background: The occurrence and development of cancer could be promoted by abnormally competing endogenous RNAs (ceRNA) network. This article aims to determine the prognostic biomarker of ceRNA for non-small-cell lung cancer (NSCLC) prognosis. Methods: The expression and clinical significance of LINC00973 in NSCLC tissues were analyzed via the The Cancer Genome Atlas (TCGA), Gene Expression Profiling Interactive Analysis (GEPIA), lnCAR, and clinical samples in Taihe Hospital. The biological functions and signaling pathways involved in target genes of ceRNA network were analyzed via Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG). Survival analysis, univariate and multivariate Cox regression analysis were used for prognostic-related mRNA. Results: Expression of LINC00973 was increased in NSCLC tissues. High expression of LINC00973 was associated with poor prognosis of NSCLC patients. There were 15 miRNA and 238 differential mRNA in the INC00973-miRNA-mRNA ceRNA network, involving cell migration, endothelial cell proliferation, tumor growth factor (TGF)-ß, cellular senescence, phosphatidylinositol 3-hydroxy kinase (PI3K)-Akt, Hippo, Rap1, mitogen-activated protein kinase (MAPK), cell cycle signaling pathway, etc. The expression levels of RTKN2, NFIX, PTX3, BMP2 and LOXL2 were independent risk factors for the poor prognosis of NSCLC patients. Conclusions: LINC00973-miRNA-mRNA ceRNA network might be the basis for determining pivotal post-translational regulatory mechanisms in the progression of NSCLC. BMP2, LOXL2, NFIX, PTX3 and RTKN2 might be valuable prognostic markers and potential therapeutic targets.

The effect of activated κ-opioid receptor (κ-OR) on the role of calcium sensing receptor (CaSR) in preventing hypoxic pulmonary hypertension development.

κ-opioid receptor (κ-OR) plays a key role in preventing hypoxic pulmonary hypertension (HPH) development after activated by exogenous agonist U50,488H. Calcium sensing receptor (CaSR) activation induces HPH by promoting vasoconstriction and vascular remodeling. The activated κ-OR is reported to inhibit the expression of CaSR in pulmonary artery smooth muscle cells (PASMCs). Thus, in this study, we aimed to explore the effect of activated κ-OR on the role of CaSR in preventing HPH development. An HPH rat model was constructed using Sprague-Dawley rats. Changes in mean pulmonary arterial pressure (mPAP) and right ventricular pressure (RVP) mediated by κ-OR agonist U50,488H and CaSR inhibitor NPS2143 were observed. The effects of CaSR agonist spermine and inhibitor NPS2143 on pulmonary artery tension were tested. The expression and localization of κ-OR and CaSR were measured in isolated PASMCs. A cell-counting kit-8 assay was performed to evaluate the effect of spermine in PASMC proliferation. Expression of proliferating cell nuclear antigen (PCNA), Erk, and p-Erk was evaluated by western blot analysis. Results showed that κ-OR and CaSR were co-expressed and colocalized in PASMCs under normoxic and hypoxic conditions. Interactions between κ-OR and CaSR were also observed. Spermine improved vasoconstriction in the pulmonary artery in HPH rats, which was abolished by U50,488H. RVP and mPAP were significantly increased in HPH rats under CaSR stimulation, but were significantly reduced when the rats were pretreated with U50,488H and NPS2143 (P < 0.01). Spermine treatment significantly promoted PASMC proliferation, which was significantly inhibited by U50,488H, p38 inhibitor SB203580, JNK inhibitor SP600125, Erk inhibitor SCH772984, and MEK inhibitor U0126, especially Erk inhibitor (P < 0.01). Spermine significantly increased PCNA and P-Erk expression in hypoxic conditions, which was inhibited by U50,488H and NPS2143. κ-OR stimulation prevented HPH development via the CaSR/MAPK signaling pathway.

The relationship between the variants of the bovine MBL2 gene and milk production traits, mastitis, serum MBL-C levels and complement activity.

Mannose-binding lectin (MBL), a calcium-dependent collagenous lectin, plays an important role in the host immune defence against a wide range of pathogens. There are MBL1 and MBL2 genes which encode the MBL-A and MBL-C proteins, respectively. This study was carried out to investigate the relationship between the variants of the bovine MBL2 gene and milk production traits, mastitis, serum MBL-C levels and hemolytic complement activity in both classical pathway (CH50) and alternative pathway (ACH50) in Chinese Holstein cattle. Four single-nucleotide polymorphisms (SNPs) in the exon 1 of the MBL2 gene in Chinese Holstein cattle and Luxi yellow cattle were identified by the direct sequencing method. The SNP g.201 G>A was identified as a non-synonymous mutation (codon 31, Arg>Gln) at the N-terminus cysteine-rich domain and the SNPs g.234 C>A and g.235 G>A (codon 42) made Pro to Gln at the 1st Gly-X-Y repeat of the collagen-like domain, while the SNP g.244 T>C (codon 45) was identified as a synonymous mutation (Asn>Asn) at the 2 th Gly-X-Y repeat of the collagen-like domain. The SNP markers (g.201 G>A, and g.234 C>A) were significantly correlated with somatic cell score (SCS) (P<0.05). The concentration of MBL-C protein in serum ranges from 0.8 to 7.4 µg/mL by enzyme-linked immunosorbent assay. Six combinations of different haplotypes from the four SNPs were identified in Chinese Holstein cattle. Statistical analysis revealed that cows with the haplotype combination H4H5 exhibited the lowest SCS. The CH50 value of H4H5 and H5H5 cow are significantly higher than H2H5 haplotype combination (P<0.05). The association analysis results showed that the haplotype combination H4H5 may be used as a tolerance haplotype combination for the bovine mastitis.