[Detection of molecular affecting sensitivity to local glucocorticoid therapy in oral lichen planus through transcriptome sequencing].

OBJECTIVE: To detect key genes of local glucocorticoid therapy in oral lichen planus (OLP) through transcriptome sequencing. METHODS: The study prospectively enrolled 28 symptomatic patients who visitied Department of Oral Mucosa, Peking University Hospital of Stomatology from November 2019 to March 2023. Topical inunction of 0.1 g/L of dexamethasone was applied for 1 min, 3 times daily for 4 weeks. The patients' signs and pain symptoms were recorded and they were classified as effective group and ineffective group according to the treatment outcome. Their mucosa samples were collected before treatment. After isolating total RNA, transcriptome sequencing was performed. The gene expression data obtained by sequencing were analyzed differently using the DESeq2 package in R software, and the Kyoto encyclopedia of genes and genomes (KEGG) pathway enrichment analysis was performed on the basis of the hypergeometric distribution algorithm to describe the biological function of differentially expressed genes (DEGs), accordingly detecting sensitivity related molecular affecting therapeutic effect of dexamethasone. RESULTS: After 4 weeks treatment by topical dexamethasone, 13 cases of the 28 OLP patients responding well with the sign score reducing from 7.0 (4.5, 9.0) to 5.0 (3.0, 6.3), pain score decreasing from 5.0 (2.0, 5.5) to 2.0 (0.0, 3.5), oral health impact profile lessening from 5.0 (3.5, 9.0) to 1.0 (0.0, 5.0) significantly (P<0.01) were classified as effective group and 15 cases with poor response to the drug were sorted as ineffective group. There were no significant differences of demographic and baseline levels of clinical features, especially disease severity between these two groups. A total of 499 DEGs including 274 upregulated and 225 downregulated genes were identified between effective group and ineffective group. KEGG enrichment analysis showed that upregulated genes in effective group compared with ineffective group including CLDN8, CTNNA3, MYL2 and MYLPF were associated with leukocyte transendothelial migration, while downregulated genes were significantly enriched in tumor necrosis factor (TNF), interleukin-17 (IL-17), nuclear factor kappa B (NF-κB) signaling pathways, and cortisol synthesis and secretory. CONCLUSION: High expressions of CLDN8, CTNNA3, MYL2 and MYLPF genes in patients with oral lichen planus have a good clinical response to topical dexamethasone, while patients with high expression genes of inflammation pathway such as TNF, IL-17, NF-κB and cortisol synthesis and secretion received poor effect.

The expression of p53, ki67 and COX-2 in erosive-type oral lichen planus.

Oral lichen planus is a chronic inflammatory disease that affects the oral mucosa and may undergo malignant changes, which can be reflected in the expression of specific proteins that are responsible for maintaining cellular mitosis and apoptosis. The study aimed to investigate the expression of p53, ki67, and COX-2 in erosive lichen planus using immunohistochemistry to correlate these findings with the histological aspects of the disease. Thirty-three biopsies of erosive lichen planus were collected and diagnosed based on histological and clinical criteria. The blocks were processed for immunohistochemistry to assess p53, ki67, and COX-2 expression in the basal layer, suprabasal, and inflammatory infiltrate respectively. The histological analysis of the samples showed no dysplasia or metastasis. P53 stained positively in 80% of the samples, while ki67 was positive in all the cases, ranging from 5% to 85% positivity. COX-2 expression ranged from 0-50% positivity. The highest expression of p53 was observed in 8 cases (24.2%), with a maximum of 5%, and ki67 exhibited the highest expression of 90% in 3 cases (9.1%). COX-2 was negative in 27 cases (81.8%) and positive in 6 cases (18.1%), with the highest expression at 50% in 1 case and 10 % positivity in 4 cases (12.1%). In our study, the markers p53, ki67, and COX-2 proved to be useful for detecting the proliferative, inflammatory, and physiologic states of the keratinocytes. However, they did not demonstrate utility in detecting any malignant transformation.

Patients with oral lichen planus display lower levels of salivary acidic glycoproteins than individuals without oral mucosal disease.

OBJECTIVES: Salivary proteins, acidic glycoproteins, and free calcium might take part in oral mucosal defence against inflammation in oral lichen planus (OLP). The study aimed to investigate whether the levels of sulfated and sialylated glycoproteins, total protein, and free calcium in saliva from patients with OLP differ from those of individuals without oral mucosal diseases. MATERIAL AND METHODS: Patients diagnosed with OLP (n = 25) and two control groups without any oral mucosal disease; age- and gender-matched controls (n = 25, 65.6 ± 2.9 years), and younger controls (n = 25, 41.8 ± 2.5 years) were included. Subjective dry mouth (xerostomia) was assessed by asking a single-item question. Chew-stimulated whole saliva was collected to measure sulfated and sialylated glycoproteins by the Alcian Blue method. The total protein was determined spectrophotometrically, and the free calcium measured using an electrode. RESULTS: The output of salivary sulfated and sialylated glycoproteins in the OLP group (21.8 ± 2.4 µg/min) was lower than in the age- and gender-matched controls (43.0 ± 2.9 µg/min, p = 0.0002), whereas the total protein and calcium output did not differ between the three groups (p > 0.05). The prevalence of xerostomia was significantly higher in the OLP group compared to both control groups (p = 0.038). CONCLUSIONS: Patients with OLP showed a high prevalence of xerostomia and lower levels of salivary acidic type glycoproteins compared to the individuals without oral mucosa disease. CLINICAL RELEVANCE: It is relevant to investigate the role of acidic glycoproteins in the pathogenesis of OLP.

The expression of Th17/Treg in oral submucosal fibrosis carcinogenesis and the significance in the development of mucosal lesions.

It targets to explore the expression of Th17/Treg in oral submucous fibrosis (OSF) carcinogenesis and its significance in the development of mucosal lesions. In this research, 100 patients with OSF who visited our hospital for surgical treatment from March 2020 to April 2022 were selected. Based on pathological examination results, the patients were divided into 27 patients with oral leukoplakia (OK) group, 14 patients with oral lichen planus (OLP) group, 9 patients with oral squamous cell carcinoma (OSCC) group, and 50 patients with OSF group. It adopted flow cytometry (FC) to calculate the ratio of peripheral blood Th17 cells and Treg cells in four groups, and the Th17/Treg ratio was calculated; The area of oral mucosal lesions (OML) from patients was collected. It needs to compare the differences in Th17/Treg ratio and OML area among four groups and determine the correlation between indicators. ROC curve was used to analyze the diagnostic threshold of the Th17/Treg ratio for carcinogenesis. Except for the OK and OLP, it had statistical significance differences in Th17, Treg cells, and Th17/Treg ratio (P<0.001); The area of OML in the OK, OLP, and OSCC was higher than that in the simple OSF, with statistical significance (P<0.001); Th17 (%), Treg (%), and Th17/Treg all had direct ratio with the area of OML; The area of OML has directed ratio with the development of mucosal lesions (r>0, P<0.05); The areas under the ROC curve for patients with OSF combined with OK, OLP, or OSCC with Th17 (%), Treg (%), Th17/Treg, and OML area were 0.560, 0.986, 0.936, and 0.466, respectively. The expression of Th17/Treg is elevated in oral submucosal fibrosis and carcinogenesis. When mucosal lesions progress or become cancerous, the Th17/Treg ratio increases accordingly, and it has more clinical value than the increase in the OML area.

Immunohistochemical expression of T-cell subsets (CD4 and CD8) in oral lichen planus.

Introduction: oral lichen planus (OLP) is a common oral mucosal disease with various clinical manifestations. The most predominant types are reticular and erosive. Despite extensive research on the causes of OLP, the exact etiology remains unclear. However, it is believed that a T-cell-mediated response, which triggers the apoptosis of oral epithelial cells, may contribute to the development of this disorder. This study aims to investigate the different types of T-cells (specifically CD4 and CD8) present in OLP tissue samples. By using immunohistochemistry, the expressions of cluster of differentiation 4 (CD4) and cluster of differentiation 8 (CD8) will be evaluated in biopsy samples taken from OLP patients who exhibit various clinical presentations. Methods: this study was a retrospective analysis study. Oral lichen planus was established histologically in forty paraffin-embedded tissue samples. Blocks of OLP were diagnosed and characterized as reticular or erosive. Immunohistochemical staining was conducted using a monoclonal antibody for (CD4) and a polyclonal antibody for CD8. Semi-quantitative techniques were used to analyze the patterns of positively stained cells. Results: forty biopsies of OLP cases were obtained from 24 females and 16 males. The mean age was (49.15±11.39) years. Using an immunohistochemical method, the proportion of CD4 expression: CD8 expression among the epithelial-connective tissue interface was shown to be 24 (60%) cases with a predominance of CD8, 9 (22.5%) cases with no difference, and only 7 (17.5%) cases with a predominance of CD4. The proportion of CD4: CD8 among perivascular parts was shown to be 8 (20%) cases with a predominance of CD8, 20 (50%) cases with no difference, while only 12 (30%) cases had a predominance of CD4. The CD4 perivascular expression was significantly stronger in (71.4%) of erosive OLP than in reticular cases. Conclusion: T-cell subsets (CD4 and CD8) were found in the OLP infiltrates. The correlation may have contributed to the pathogenesis of OLP.

Upregulation of IL-37 in epithelial cells: A potential new mechanism of T cell inhibition induced by tacrolimus.

Oral lichen planus (OLP) is a chronic T cell-mediated mucocutaneous disease characterized by T cell infiltration at the connective tissue-epithelium interface. Traditionally, topical corticosteroids are used as the first-line drugs to treat OLP. However, long-term use of corticosteroids may lead to drug tolerance, secondary candidiasis, and autoimmune adrenal insufficiency. Although topical tacrolimus has often been recommended for short-term use in corticosteroid-refractory OLP, the precise role of tacrolimus in epithelial cells remains elusive. This study showed that tacrolimus could directly upregulate the expression of IL-37 in human gingival epithelial cells by promoting the TGF-ßRI/Smad3 pathway independently of calcineurin inhibition and MAPKs. In contrast, dexamethasone, one of the corticosteroids, did not have the same effect. Moreover, IL-37 could inhibit the proliferation of activated T cells and the secretion of effector cytokines and alleviate epithelial cell apoptosis and death caused by activated T cells ina co-culturesystem. Furthermore, compared with healthy controls, IL-37 and p-Smad3 levels significantly increased in the oral mucosa affected by OLP, especially in the epithelium. IL-37 might have mediated a negative feedback mechanism to curb excessive inflammation in OLP. However, the expression of IL-37 was not associated with the infiltration of CD8+ T cells and Tregs in OLP, implying that IL-37 might mostly affect T cell activation rather than T cell differentiation and migration. Overall, this study discovered a potential novel mechanism by which tacrolimus might indirectly inhibit T cell-mediated immune damage by upregulating IL-37 in human gingival epithelial cells.

METTL14-upregulated miR-6858 triggers cell apoptosis in keratinocytes of oral lichen planus through decreasing GSDMC.

Oral lichen planus (OLP), a chronic inflammatory disorder, is characterized by the massive cell apoptosis in the keratinocytes of oral mucosa. However, the mechanism responsible for triggering oral keratinocyte apoptosis is not fully explained. Here, we identify that Gasdermin C (GSDMC) downregulation contributes to apoptosis in human oral keratinocytes. Mechanistically, we describe that activated nuclear factor kappa B (NF-κB) pathway induces overexpression of methyltransferase-like 14 (METTL14), which increases N6-adenosine methylation (m6A) levels in the epithelial layer of OLP. m6A modification is capable of regulating primary miR-6858 processing and alternative splicing, leading to miR-6858 increases. miR-6858 can bind and promote GSDMC mRNA degradation. Forced expression of GSDMC is able to rescue cell apoptosis in human oral keratinocyte models resembling OLP. Collectively, our data unveil that m6A modification regulates miR-6858 production to decrease GSDMC expression and to trigger keratinocyte apoptosis in the context of OLP.

CD8 + T-cell marker genes reveal different immune subtypes of oral lichen planus by integrating single-cell RNA-seq and bulk RNA-sequencing.

BACKGROUND: Oral lichen planus (OLP) is a local autoimmune disease induced by T-cell dysfunction that frequently affects middle-aged or elderly people, with a higher prevalence in women. CD8 + T cells, also known as killer T cells, play an important role in the progression and persistence of OLP. In order to identify different OLP subtypes associated with CD8 + T cell pathogenesis, consensus clustering was used. METHODS: In this study, we preprocessed and downscaled the OLP single-cell dataset GSE211630 cohort downloaded from Gene Expression Omnibus (GEO) to finally obtain the marker genes of CD8 + T cells. Based on the expression of marker genes, we classified OLP patients into CMGs subtypes using unsupervised clustering analysis. The gene expression profiles were analyzed by WGCNA using the "WGCNA" R package based on the clinical disease traits and typing results, and 108 CD8 + T-cell related OLP pathogenicity-related genes were obtained from the intersection. Patients were once again classified into gene subtypes based on intersection gene expression using unsupervised clustering analysis. RESULTS: After obtaining the intersecting genes of CD8 + T cells related to pathogenesis, OLP patients can be precisely classified into two different subtypes based on unsupervised clustering analysis, and subtype B has better immune infiltration results, providing clinicians with a reference for personalized treatment. CONCLUSIONS: Classification of OLP into different subtypes improve our current understanding of the underlying pathogenesis of OLP and provides new insights for future studies.

Análise da imunoexpressão de BAX e MMP-9 no líquen plano oral / Analysis of BAX and MMP-9 immunoexpression in oral lichen planus

Introdução: O líquen plano oral é uma doença crônica imunologicamente mediada relativamente comum, que acomete a mucosa oral. Clinicamente, o LPO é classificado em seis padrões bem identificados: placa, reticular, bolhoso, atrófico, papular e erosivo.Sendo os mais comuns oos tipos reticulares e erosivos. A ativação dos linfócitos TCD4+ no LPO, pode induzir os ceratinócitos ao processo de apoptose através da respostaimunológica citotóxica. A proteína Bax desempenha uma função relevante para o processo apoptótico. Deste modo, a presente pesquisa consistiu em um estudo transversal retrospectivo, descritivo, quantitativo e comparativo. Objetivo: Avaliar a expressão imuno-histoquímica das proteínas MMP9 e Bax no LPO. Método: Foram utilizados 43 casos de LPO para análise da imunoexpressão de Bax e MMP-9. Os resultados foram analisados através dos testes estatísticos apropriados e serão considerados significativos, valores onde p<0,05. Resultado: A imunoexpressão de MMP9 foi significativamente maior nos ceratinócitos e quando analisados os subtipos de líquen plano oral, não foram observados diferenças estatísticas entre os tipos reticulares e erosivos para as proteínas analisadas. Conclusões: Com essas observações, infere-se que a alteração na expressão das proteínas estudadas sugere um distúrbio nos mecanismos apoptóticos, os quais estão associados às lesões de LPO, e podemos concluir também que as imunoexpressões dessas proteínas não apresentaram diferença, quando relacionada ao tipo clínico reticular ou erosivo. Com esse resultado pode-se contribuir para um maior entendimento sobre os possíveis mecanismos celulares envolvidos na etiopatogenia dessa lesão (AU).

Background: Oral lichen planus is a relatively common immune-mediated chronic disease that affects the oral mucosa. Clinically, OLP is classified into six well-identified patterns: plaque, reticular, bullous, atrophic, papular, and erosive. The most common being the reticular and erosive types. The activation of TCD4+ lymphocytes in the OLP can induce keratinocytes to the process of apoptosis through the cytotoxic immune response. Thus, the present research consisted of a retrospective, descriptive, quantitative and comparative crosssectional study. Objective: to evaluate the immunohistochemical expression of MMP-9 and Bax proteins in OLP. Methods: We used 20 cases of Inflammatory Fibrous Hyperplasia as control. The results were analyzed through the appropriate statistical tests and will be considered significant, values where p<0.05. Results: The immunoexpression of MMP-9 was significantly higher in keratinocytes and when the subtypes of oral lichen planus were analyzed, no statistical differences were observed between the reticular and erosive types for the proteins analyzed. Conclusions: With these observations, it is inferred that the alteration in the expression of the studied proteins suggests a disturbance in the proliferative and apoptotic mechanisms, which are associated with a pathological behavior of the oral mucosa, and consequently with a repercussion on the lesions of OLP, and we can also conclude that the immunoexpression of these proteins had no difference, when related to the reticular or erosive clinical type. This research aims to contribute to a greater understanding of the possible cellular mechanisms involved in the etiopathogenesis of this lesion, thus enabling the understanding of the clinical aspects of the pathology (AU).

Effects of interleukin-23 on the activation of mucosal-associated invariant T cells from oral lichen planus.

OBJECTIVE: Oral lichen planus (OLP) is a T cell-mediated inflammatory condition in the oral cavity. Mucosal-associated invariant T (MAIT) cells are gaining more relevance in immune diseases because they can be activated by cytokines without T cell receptor stimulation. Herein, we tested the effect of interleukin-23 (IL-23) on the activation status of OLP MAIT cells. METHODS: Peripheral blood mononuclear cells (PBMCs) isolated from OLP patients were stimulated by IL-23 in the absence or presence of phorbol myristate acetate (PMA) and ionomycin. The activation status of MAIT cells was analyzed by flow cytometry after staining for CD3, CD4, CD8, CD161, TCR Vα7.2, and CD69. RESULTS: The fraction of MAIT cells in OLP peripheral blood was approximately 0.38% to 3.97%, and CD8+ subpopulations overwhelmed CD4+ cells. The mean percentages of OLP MAIT cells in PBMCs and CD8+MAIT cells in MAIT cells were approximately 40%. PMA and ionomycin significantly increased CD69 expression on OLP T cells, MAIT cells, and CD8+MAIT cells. Cells with enhanced activation had different responsiveness to exogenous IL-23, showing increased CD69 expression on OLP T cells, decreased CD69 on OLP CD8+MAIT cells, and no significant change on OLP MAIT cells. CONCLUSIONS: IL-23 showed different effects on the activation status of OLP MAIT cells and CD8+MAIT cells.

Predicting the Activity of Oral Lichen Planus with Glycolysis-related Molecules: A Scikit-learn-based Function.

OBJECTIVE: Oral lichen planus (OLP) is one of the most common oral mucosa diseases, and is mainly mediated by T lymphocytes. The metabolic reprogramming of activated T cells has been shown to transform from oxidative phosphorylation to aerobic glycolysis. The present study investigated the serum levels of glycolysis-related molecules (lactate dehydrogenase, LDH; pyruvic acid, PA; lactic acid, LAC) in OLP, and the correlation with OLP activity was assessed using the reticular, atrophic and erosive lesion (RAE) scoring system. METHODS: Univariate and multivariate linear regression functions based on scikit-learn were designed to predict the RAE scores in OLP patients, and the performance of these two machine learning functions was compared. RESULTS: The results revealed that the serum levels of PA and LAC were upregulated in erosive OLP (EOLP) patients, when compared to healthy volunteers. Furthermore, the LDH and LAC levels were significantly higher in the EOLP group than in the nonerosive OLP (NEOLP) group. All glycolysis-related molecules were positively correlated to the RAE scores. Among these, LAC had a strong correlation. The univariate function that involved the LAC level and the multivariate function that involved all glycolysis-related molecules presented comparable prediction accuracy and stability, but the latter was more time-consuming. CONCLUSION: It can be concluded that the serum LAC level can be a user-friendly biomarker to monitor the OLP activity, based on the univariate function developed in the present study. The intervention of the glycolytic pathway may provide a potential therapeutic strategy.

Altered Expression of Vascular Cell Adhesion Molecule-1 in Oral Lichen Planus.

Oral lichen planus (OLP) is a T cell-mediated chronic inflammatory mucocutaneous disease affected by the interaction between keratinocytes and T cells. Recent evidence indicates that vascular cell adhesion molecule-1 (VCAM1) plays a vital role in mediating immune and inflammatory responses. In this study, the expression of VCAM1 in OLP was detected by immunohistochemical staining and its correlations with clinical features were analyzed. The disease severity of OLP was assessed by the reticular, atrophic, and erosive scoring system. We found that VCAM1 was generally localized in the cytoplasm of epithelial cells, and in nucleus, cytoplasm, and extracellular matrix of subepithelial infiltrated cells in superficial layer of lamina propria. Moreover, VCAM1 levels in epithelium and lamina propria of OLP were significantly higher than that in controls, respectively. In addition, VCAM1 level in epithelium was increased compared with that of lamina propria. There were no significant differences for VCAM1 expression between nonerosive and erosive forms of OLP. The expression of VCAM1 in OLP was not associated with the severity of disease, gender, and age. Thus, we speculated that spatial expression differences of VCAM1 in local lesions of OLP may involve the pathogenesis of OLP.

Differential proteomic expression in indolent versus transforming oral lichen planus.

Oral lichen planus (OLP) confers an approximately 1% risk of transformation to oral squamous cell carcinoma (OSCC). Early identification of high-risk OLP would be very helpful for optimal patient management. We aimed to discover specific tissue-based protein biomarkers in patients with OLP who developed OSCC compared to those who did not. We used laser capture microdissection- and nanoLC-tandem mass spectrometry to assess protein expression in fixed lesional mucosal specimens in patients with indolent OLP (no OSCC after at least 5-year follow-up, n = 6), transforming OLP (non-dysplastic epithelium with lichenoid inflammation marginal to OSCC, n = 6) or normal oral mucosa (NOM, n = 5). Transforming OLP protein profile was enriched for actin cytoskeleton, mitochondrial dysfunction and oxidative phosphorylation pathways. CA1, TNNT3, SYNM and MB were overexpressed, and FBLN1 was underexpressed in transforming OLP compared with indolent OLP. Integrin signalling and antigen presentation pathways were enriched in both indolent and transforming OLP compared with NOM. This proteomic study provides potential biomarkers, such as CA1 overexpression, for higher-risk OLP. While further validation studies are needed, we propose that epithelial-mesenchymal transition may be involved in OLP carcinogenesis.

Characterization of Mucosal-Associated Invariant T Cells in Oral Lichen Planus.

Oral lichen planus (OLP) is an inflammatory condition of unknown cause that has been associated with concurrent candidal infection. Mucosal-associated invariant T (MAIT) cells express the T cell receptor TCRVα7.2 and are activated by riboflavin intermediates produced by microbes. The interaction between MAIT cells, Candida, and OLP is unknown. This study aimed to determine mucosal-associated T cell presence in OLP and whether the abundance of these cells changed due to the presence of either Candida or symptoms, using multiplex immunohistochemistry (mIHC). Ninety formalin fixed-paraffin-embedded (FFPE) tissue samples were assessed using mIHC for the cellular markers CD3, interleukin 18 receptor one (IL18R1), TCRVα7.2, CD161, CD8, and major histocompatibility complex class I-related (MR-1) protein. The samples were stratified into five groups on the basis of clinical (presence/absence of symptoms) and microbiological (presence/absence of Candida) criteria. Results demonstrated the presence of MAIT cell phenotypes in OLP inflammatory infiltrate within the connective tissue. Significant differences existed between different OLP groups with the percentage of log(CD3+ CD161+) and log(CD3+ TCRVα7.2+) positive cells (p < 0.001 and p = 0.005 respectively). Significant differences also existed with the relative abundance of triple-stained log(CD3+ CD161+ IL18R1+) cells (p = 0.004). A reduction in log(CD3+ CD161+ IL18R1+) cells was observed in lesional tissue of patients with symptomatic OLP with and without Candida when compared to controls. When present in OLP, MAIT cells were identified within the connective tissue. This study demonstrates that mIHC can be used to identify MAIT cell phenotypes in OLP. Reduced percentage of log(CD3+ CD161+ IL18R1+) cells seen in symptomatic OLP with and without Candida suggests a role for these cells in OLP pathogenesis.

Autophagy-related 9 homolog B regulates T-cell-mediated immune responses in oral lichen planus.

OBJECTIVES: This study aimed to explore the impacts of autophagy-related 9 homolog B (ATG9B)-mediated autophagy on T-cell immune responses in oral lichen planus. DESIGN: ATG9B expression was detected in lesions and local T cells by immunohistochemical analysis and immunofluorescence assay. The effects of ATG9B-mediated autophagy on T-cell immune responses were explored after ATG9B-overexpression or ATG9B-knockdown lentivirus transfection. A coculture system of activated T cells and lipopolysaccharide-induced keratinocytes was used to simulate the main cell crosstalk in oral lichen planus. RESULTS: The expression of ATG9B upregulated in lesions and local T cells of oral lichen planus, especially in non-erosive oral lichen planus, suggesting that ATG9B may be a diagnostic factor for oral lichen planus. Notably, ATG9B-knockdown T cells of oral lichen planus demonstrated autophagy suppression, enhanced proliferation, and attenuated apoptosis, whereas overexpression of ATG9B showed opposite effects on T cells. In the coculture system of T cells and keratinocytes, ATG9B-knockdown T cells of oral lichen planus, but not ATG9B-overexpression T cells, promoted the proliferation and apoptosis of their cocultured keratinocytes. Additionally, exogenous insulin-like growth factor 1 (IGF1) significantly reversed the apoptosis rates of keratinocytes cocultured with T cells expressing abnormal ATG9B. Furthermore, ATG9B-overexpression T cells showed decreased secretion of interferon-γ and tumor necrosis factor-α in the coculture system. CONCLUSIONS: This study revealed the regulatory roles of ATG9B-mediated T-cell autophagy on T-cell immune responses and crosstalk between T cells and keratinocytes in of oral lichen planus.

The critical role of IFNγ in the epidermotropic migration of lymphocytes in oral lichen planus.

INTRODUCTION: The chemokines play a crucial role in the recruitment of lymphocytes in oral lichen planus, and the activated epithelial cells are the main producers of the chemokines. However, the signals provoking chemokine secretion still remain to be elucidated. METHODS: The global expression profile of chemokines in oral epithelial cell line induced by IFNγ was determined by microarray analysis. The gene and protein expression was validated in primary culture of oral epithelial cells, and the effects of IFNγ on regulating chemokine production were compared with that of TNFα and IL2. Moreover, the capability of primary culture of oral epithelial cells to attract peripheral lymphocytes in response to IFNγ was investigated in oral lichen planus patients, and the cell phenotype of the recruited lymphocytes was analyzed using flow cytometry. RESULTS: IFNγ triggered the expression of multiple chemokines in the oral epithelial cells. The expression pattern of the chemokines closely resembled that in the epithelial cell layer of oral lichen planus lesions. Compared with IL2 and TNFα, IFNγ demonstrated a distinct maximal effect on the chemokines secretion in primary culture of oral epithelial cells. The migration of peripheral lymphocytes toward the culture supernatant of IFNγ-treated primary culture of oral epithelial cells was significantly enhanced in the oral lichen planus group compared to that in the healthy control group. CONCLUSION: IFNγ plays an important role in the chemokine secretion and epidermotropic migration of lymphocytes in oral lichen planus.

[Expression and significance of microtubule associated protein 1 light chain 3B, p62 and Beclin1 in lesion tissues of oral lichen planus patients].

Objective: To explore the expression of autophagy related factors microtubule associated protein 1 light chain 3B (LC3B), p62, autophagy key factor Beclin1 in oral lichen planus (OLP) tissues and their relationships with the clinicopathological characteristics of OLP, investigating the function and significance of autophagy in pathogenesis of OLP. Methods: Forty-one lesion tissues (OLP group, twenty-one cases of erosive OLP and twenty cases of non-erosive OLP) were selected from OLP patients visiting the Department of Periodontal and Oral Medicine, School and Hospital of Stomatology, Guizhou Medical University from October 2017 to December 2019. Fifteen cases of normal oral mucosal tissues (control group) were collected from oral and maxillofacial surgery at The Affiliated Stomatology Hospital of Guizhou Medical University during the same period. Protein and mRNA expression levels of LC3B, p62 and Beclin1 were detected by immunohistochemistry (IHC) and real-time quantitative PCR (RT-qPCR) in OLP lesions respectively. The protein expression levels of LC3B, p62, Beclin1 and ratio of LC3B-Ⅱ/LC3B-Ⅰ in sixteen cases (eight cases of erosive OLP and eight cases of non-erosive OLP) from the OLP group were detected by Western blotting (WB). The potential relationship between LC3B, p62, Beclin1, LC3B-Ⅱ/LC3B-Ⅰ ratio and clinical features of OLP were analyzed. Results: IHC results showed that the positive expression rates of LC3B and p62 proteins in OLP lesion tissues [LC3B: 68% (28/41); p62: 59% (24/41)] were higher than those in the control group [LC3B: 5/15; p62: 3/15] (LC3B: χ2=5.55, P=0.019; p62: χ2=5.55, P=0.015). The positive expression rates of LC3B and p62 proteins in the erosive OLP group [LC3B: 86% (18/21); p62: 76% (16/21)] were higher than those in the non-erosive OLP group [LC3B: 50% (10/20); p62: 40% (8/20)] (LC3B: χ2=4.50, P=0.034; p62:χ2=5.53, P=0.019). The positive expression rate of Beclin1 protein in the OLP lesions[20% (8/41)] was lower than that in the control group (7/15) (χ2=4.13, P=0.042), but was not statistically different between the two types of OLP (P>0.05). The RT-qPCR results showed that the mRNA expression levels of LC3B and p62 in OLP lesions [LC3B: 2.78 (1.59, 6.15); p62: 4.30 (2.34, 6.29)] were higher than those in the control group [LC3B: 1.05 (0.88, 1.21); p62: 1.12 (0.89, 1.36)] (LC3B: Z=-4.56, P<0.001; p62: Z=-4.78, P<0.001), and the mRNA expression levels of LC3B and p62 in the erosive OLP group were higher than those in the non-erosive OLP group (LC3B: Z=-2.87, P=0.004; p62: Z=-2.95, P=0.003). The mRNA expression level of Beclin1 in OLP tissues was lower than that in the control group (Z=-2.43, P=0.015), but the difference was not statistically significant between the two types of OLP (P>0.05). WB results showed that the LC3B-Ⅱ/LC3B-Ⅰ ratio was higher in the OLP lesions than that in the control group (t=-2.45, P=0.021), and the LC3B-Ⅱ/LC3B-Ⅰ ratio was higher in the non-erosive OLP group than in the erosive OLP group (t=-2.38, P=0.032). Spearman's correlation analysis showed that the ratio was negatively correlated with the clinical staging and the degree of basal cell liquefaction in OLP (clinical staging: r=-0.57, P=0.021; basal cell liquefaction: r=-0.54, P=0.032), but not with the disease duration and the degree of lymphocytic infiltration (P>0.05). Conclusions: Autophagy related factors LC3B, p62 and Beclin1 may play a role in the formation and progression of OLP lesions. The autophagy level was relatively lack in erosive OLP compared to non-erosive OLP, contributing to the increased local lesion destruction in erosive OLP. Abnormal cellular autophagy may play an important role in the formation of OLP lesions.

Immunohistochemical correlation of mast cells and angiogenesis in oral lichen planus.

OBJECTIVE: The study was conceived with an aim to correlate mast cell expression and angiogenesis in oral lichen planus (OLP) and inflamed oral mucosa. DESIGN: An observational study was performed on fifty tissue blocks of OLP and inflamed oral mucosa which were stained immunohistochemically using mast cell tryptase, CD34 and CD105. The data was analysed using Mann Whitney U-test and Spearman's rank correlation coefficient test (p < 0.05). RESULTS: OLP showed significantly higher mast cell density for total and degranulated mast cells and higher microvessel density (CD105) compared to inflamed oral mucosa (p = 0.006, 0.000). Microvessel density, total and mean vascular area (CD34) were higher in inflamed oral mucosa with a non significant difference (p = 0.985, 0.977, 0.720). Mild negative association was observed between mast cells and angiogenesis in OLP (p = 0.879). A significant positive correlation was seen between degranulated mast cells and microvessel density (CD34) in inflamed oral mucosa (p = 0.009). CONCLUSION: Mast cells execute different roles in OLP depending upon stage of the disease. We found that although degranulation of mast cells is not the predominant pathway for angiogenesis in OLP, yet is one of the important precursor phenomena in inflamed tissues.

Serum vitamin D level in healthy individuals versus patients with symptomatic and asymptomatic oral lichen planus.

The aetiology of oral lichen planus (OLP) is multifactorial, having variable triggers. A role for vitamin D related to the immune system has been established. Vitamin D modulating effect is on the adaptive and innate immune responses. Our study aimed to compare serum levels of vitamin D in patients having different clinical symptoms of OLP (symptomatic or asymptomatic) with healthy individuals. Also, in this study, for further evaluation, the expression level of interleukin-17A and interleukin-6 (IL-17A and IL-6) was evaluated because the presence of active vitamin D reduces the expression of these pro-inflammatory factors. This study was included three groups with 30 volunteers in each. The first group included asymptomatic oral lichen planus patients (reticular or plaque-like lesions). The second group consisted of symptomatic oral lichen planus patients (atrophic or bullous-erosive lesions). In contrast, the third group consisted of healthy control subjects. The serum 25-hydroxyvitamin D was measured between the three groups and then correlated with clinical manifestation of oral lichen planus, either symptomatic or non-symptomatic. The Real-Time PCR technique was used to evaluate the expression of IL-17A and IL-6. Patients with symptomatic OLP (second group) had statistically significantly lower Vitamin D levels than asymptomatic OLP patients (first group). Healthy Controls (third group) exhibited statistically significantly higher vitamin D levels than OLP groups. The results of IL-17A and IL-6 genes expression showed that the presence of vitamin D had a statistically significant effect on reducing the expression of these two pro-inflammatory cytokines among symptomatic and asymptomatic OLP patients. Also, the results showed that there was a statistically significant difference between OLP patients (group I and II) and the control group (group III). In general, the current study results showed that lack of vitamin D had an important role in initiating or increasing the OLP's severity.

Automatic Detection of Image-Based Features for Immunosuppressive Therapy Response Prediction in Oral Lichen Planus.

Oral lichen planus (OLP) is a chronic inflammatory disease, and the common management focuses on controlling inflammation with immunosuppressive therapy. While the response to the immunosuppressive therapy is heterogeneous, exploring the mechanism and prediction of the response gain greater importance. Here, we developed a workflow for prediction of immunosuppressive therapy response prediction in OLP, which could automatically acquire image-based features. First, 38 features were acquired from 208 OLP pathological images, and 6 features were subsequently obtained which had a significant impact on the effect of OLP immunosuppressive therapy. By observing microscopic structure and integrated with the corresponding transcriptome, the biological implications of the 6 features were uncovered. Though the pathway enrichment analysis, three image-based features which advantageous to therapy indicated the different lymphocytes infiltration, and the other three image-based features which bad for therapy respectively indicated the nicotinamide adenine dinucleotide (NADH) metabolic pathway, response to potassium ion pathway and adenosine monophosphate (AMP) activated protein kinase pathway. In addition, prediction models for the response to immunosuppressive therapy, were constructed with above image-based features. The best performance prediction model built by logistic regression showed an accuracy of 90% and the area under the receiver operating characteristic curve (AUROC) reached 0.947. This study provided a novel approach to automatically obtain biological meaningful image-based features from unannotated pathological images, which could indicate the immunosuppressive therapy in OLP. Besides, the novel and accurate prediction model may be useful for the OLP clinical management.