Early activation of the interleukin-23-17 axis in a murine model of oropharyngeal candidiasis

P>Candida albicans is an oral commensal yeast that causes oropharyngeal candidiasis (OPC) in immunocompromised individuals. The immunological pathways involved in OPC have been revisited after the interleukin-17 (IL-17) pathway was implicated in fungal immunity. We studied immediate (< 24 h) and adaptive (3-6 day) IL-12 and IL-23-17 pathway activation in naive p40-/- mice, which lack IL-12 and IL-23 and develop severe, chronic OPC upon oral inoculation with C. albicans. Macrophages from p40-/- mice were less efficient than C57BL/6J controls at killing C. albicans in vitro but very low numbers in the oral mucosae of infected C57BL/6J mice suggest that they are not critical in vivo, at least in this strain. Migration of macrophages to regional lymph nodes of infected p40-/- mice was impaired; however, dendritic cell migration was not affected. Recombinant IL-12 therapy provided only temporary relief from OPC, suggesting that IL-23 is required for full protection. In C57BL/6J mice, but not p40-/- mice, messenger RNAs encoding IL-23p19 and IL-17 were induced in the oral mucosa within 24 h of infection (6 +/- 0.6 and 12 +/- 2.7-fold). By day 6 of infection in C57BL/6J mice, IL-17A messenger RNA level had increased 5.1 +/- 1.8 and 83 +/- 21-fold in regional lymph nodes and oral tissues respectively. Ablation of p40 was associated with delayed or abrogated induction of IL-17A pathway targets (monocyte chemoattractant protein-1, IL-6 and macrophage inflammatory protein-2), and a lack of organized recruitment of neutrophils to the infected oral mucosa. Overall our data show that the IL-23-17A axis is activated early in the oral mucosae of immunologically naive mice with OPC

Medicinska gljiva Lignosus rhinocerus svojim imunomodulacijskim učinkom i reguliranjem signalnog puta posredovanog faktorom nekroze tumora uzrokuje apoptozu i zaustavlja stanični ciklus stanica karcinoma usne šupljine ORL-204

Research background. Tiger milk mushroom (Lignosus rhinocerus) is a medicinal mushroom that is geographically distributed in the region of South China, Thailand, Malaysia, Indonesia, Philippines and Papua New Guinea. Consumption of its sclerotium has been reported to treat various ailments. However, its anticancer potential towards oral cancer cell lines is yet to be determined considering the traditional method of its consumption by biting/chewing of the sclerotium. Experimental approach. Mushroom sclerotial powder of cultivar TM02® was extracted and fractionated in a chromatographic column prior to cytotoxicity testing against a panel of human oral cancer cell lines. The capability of the identified bioactive fraction in regulating several molecules associated with its tumour necrosis factor (TNF) pathway was investigated. Results and conclusions. 2,5-Diphenyl-2H-tetrazolium bromide (MTT) proliferation assay indicated that cell lines ORL-48 (derived from gingiva), ORL-188 (derived from the tongue) and ORL-204 (derived from buccal mucosa) were inhibited by cold water extract of L. rhinocerus sclerotia and its high-molecular-mass fraction (HMM) in varying degrees with ORL-204 being most affected. Hence, the treatment of ORL-204 with HMM mushroom extract was further investigated. HMM mushroom extract induced apoptosis and G0/G1 phase cell cycle arrest through caspase-3/7 cleavage. Activities of MIP2 and COX-2 were downregulated by 0.2- and 4.6-fold respectively in the HMM mushroom extract-treated ORL-204 cells. Novelty and scientific contribution. Using ORL-204, we showed that HMM mushroom extract may act via the TNF pathway at various network sites as a potential dietary compound for cancer prevention and natural adjunct therapeutic to conventional cancer treatment.Pozadina istraživanja. Medicinska gljiva Lignosus rhinocerus rasprostranjena je na području južne Kine, Tajlanda, Malezije, Indonezije, Filipina i Papua Nove Gvineje. Sklerocij gljive koristi se za liječenje različitih oboljenja. Međutim, dosad još nije ispitan antikacerogeni učinak sklerocija ove medicinske gljive, koja se tradicionalno konzumira tako da se grize odnosno žvače, na stanice karcinoma usne šupljine. Eksperimentalni pristup. Ekstrakt praha sklerocija kultivara gljive TM02® frakcioniran je pomoću kromatografske kolone, te je zatim ispitan njegov citokosični učinak na različite stanične linije humanih karcinoma usne šupljine. Ispitana je sposobnost bioaktivne frakcije da regulira molekule koje sudjeluju u sintezi faktora nekroze tumora (TNF). Rezultati i zaključci. Ispitivanjem proliferacije stanica pomoću testa redukcije 2,5-difenil-2H-tetrazolijeva bromida (MTT) utvrđeno je da ekstrakt sklerocija L. rhinocerus dobiven hladnom vodom, i to frakcija velike molekulske mase, u različitoj mjeri inhibira rast staničnih linija ORL-48 (izoliranih iz desni), ORL-188 (izoliranih ih jezika), a ponajviše onih linije ORL-204 (izoliranih iz sluznice obraza). Stoga smo dodatno istražili učinak ekstrakta gljive velike molekulske mase na staničnu liniju ORL-204. Ektrakt je potaknuo apoptozu i zaustavio stanični ciklus u fazi G0/G1 cijepanjem kaspaze 3/7. U stanicama ORL-204 tretiranim ekstraktom gljive velike molekulske mase smanjila se aktivnost enzima MIP2 za 0,2 puta, a enzima COX-2 za 4,6 puta. Novina i znanstveni doprinos. Pomoću stanične linije ORL-204 pokazali smo da ekstrakt medicinske gljive velike molekulske mase može djelovati na sintezu faktora nekroze tumora, te se upotrijebiti kao prirodni dodatak prehrani za prevenciju razvoja karcinoma ili kao dodatak konvencionalnom liječenju karcinoma

Transcriptional analysis highlights three distinct immune profiles of high-risk oral epithelial dysplasia

Oral potentially malignant disorders (OPMD) are precursors of oral squamous cell carcinoma (OSCC), and the presence of oral epithelial dysplasia (OED) in OPMD confers an increased risk of malignant transformation. Emerging evidence has indicated a role for the immune system in OPMD disease progression; however, the underlying immune mechanisms remain elusive. In this study, we used immune signatures established from cancer to delineate the immune profiles of moderate and severe OED, which are considered high-risk OPMD. We demonstrated that moderate and severe OEDs exhibit high lymphocyte infiltration and upregulation of genes involved in both immune surveillance (major histocompatibility complex-I, T cells, B cells and cytolytic activity) and immune suppression (immune checkpoints, T regulatory cells, and tumor-associated macrophages). Notably, we identified three distinct subtypes of moderate and severe OED: immune cytotoxic, non-cytotoxic and non-immune reactive. Active immune surveillance is present in the immune cytotoxic subtype, whereas the non-cytotoxic subtype lacks CD8 immune cytotoxic response. The non-immune reactive subtype showed upregulation of genes involved in the stromal microenvironment and cell cycle. The lack of T cell infiltration and activation in the non-immune reactive subtype is due to the dysregulation of CTNNB1, PTEN and JAK2. This work suggests that moderate and severe OED that harbor the non-cytotoxic or non-immune reactive subtype are likely to progress to cancer. Overall, we showed that distinct immune responses are present in high-risk OPMD, and revealed targetable pathways that could lead to potential new approaches for non-surgical management of OED

Apoptosis-Inducing Effect of Three Medicinal Plants on Oral Cancer Cells KB and ORL-48

Brucea javanica, Azadirachta indica, and Typhonium flagelliforme are medicinal plants commonly used to treat conditions associated with tumour formation. This study aimed to determine the antiproliferative activity of these plants extracts on KB and ORL-48 oral cancer cell lines and to suggest their mode of cell death. The concentration producing 50% cell inhibition (IC50) was determined and the activity was examined under an inverted microscope. Immunohistochemistry fluorescent staining method (TUNEL) was performed to indicate the mechanism of cell death and the fragmented DNA band pattern produced was obtained for verification. Compared to Azadirachta sp. and Typhonium sp., the antiproliferative activity of Brucea sp. extract was the most potent on both KB and ORL-48 cells with IC50 of 24.37 ± 1.75 and 6.67 ± 1.15 µg/mL, respectively. Signs of cell attrition were observed 24 hr after treatment. Green fluorescent spots indicating cell death by apoptosis were observed in images of both cells following treatment with all the three extracts. DNA fragments harvested from Brucea-treated cells produced bands in a ladder pattern suggesting the apoptotic effect of the extract. It is thus concluded that Brucea sp. extract exhibited cytotoxic activity on ORL-48 cells and their action mechanism is via apoptosis

Predicting malignant progression in clinically high-risk lesions by DNA ploidy analysis and dysplasia grading

The value of image cytometry DNA ploidy analysis and dysplasia grading to predict malignant transformation has been determined in oral lesions considered to be at 'high' risk on the basis of clinical information and biopsy result. 10-year follow up data for 259 sequential patients with oral lesions clinically at 'high' risk of malignant transformation were matched to cancer registry and local pathology database records of malignant outcomes, ploidy result and histological dysplasia grade. In multivariate analysis (n = 228 patients), 24 developed carcinoma and of these, 14 prior biopsy samples were aneuploid. Aneuploidy was a significant predictor (hazard ratio 7.92; 95%CI 3.45, 18.17) compared with diploidy (p < 0.001). The positive predictive value (PPV) for severe dysplasia was 50% (95%CI 31.5, 68.5) and for aneuploid lesions, 33.3% (95%CI 19.0, 47.6). Combined DNA aneuploidy and severe dysplasia increased PPV to 56.3% (95% CI 31.9, 80.6). Diploid-tetraploid and non-dysplastic status had high negative predictive values (NPV) of 94.6% (95% CI 91.4, 97.8) and 99.17% (95% CI 97.4, 100.8) respectively. DNA ploidy predicts malignant transformation well and combining it with dysplasia grading gave the highest predictive value. The predictive values reported here exceed those from other investigations to date

DNA aneuploidy and tissue architecture in oral potentially malignant disorders with epithelial dysplasia assessed by a 10 locus FISH panel

Subjectivity in oral dysplasia grading has prompted evaluation of molecular-based tests to predict malignant transformation. Aneuploidy detected by DNA image-based cytometry (ICM) is currently the best predictor but fails to detect certain high risk lesions. A novel multiplex fluorescence in situ hybridization (FISH) panel was used to explore possible explanations by detecting aneuploidy at the single cell level. FISH was compared to reference standard DNA ICM in 19 oral lesions with epithelial dysplasia and used to characterize the cellular architecture. Copy number variation at 3q28, 7p11.2, 8q24.3, 11q13.3 and 20q13.12 and matched chromosome specific loci were assessed by dual-color FISH to assess numerical and spatial patterns of copy number increase and gene amplification. FISH revealed wide variation in copy number at different loci. Only low level copy number gain was present and often in only a small proportion of cells, although usually with all or all but one locus (9/12). Four cases showed gene amplification, one at two loci. Some probes revealed an internal presumed clonal structure within lesions not apparent in routine histological examination. Both methods produced similar diagnostic results with concordance in detection of aneuploidy by both methods in 17 out of 19 samples (89%). We have shown that oral dysplastic lesions may contain very few aneuploid cells at a cellular level, high copy number gain is rare and changes appear to arise from large chromosomal fragment duplications. Single stem lines are relatively homogeneous for loci with copy number gain but there is a subclonal structure revealed by gene amplification in some lesions. © 2020 Spandidos Publications. All rights reserved

Effect of reconstruction parameters on cone beam CT trabecular bone microstructure quantification in sheep

Background: Cone Beam Computed Tomography (CBCT) is a reliable radiographic modality to assess trabecular bone microarchitecture. The aim of this study was to determine the effect of CBCT image reconstruction parameters, namely, the threshold value and reconstruction voxel size, on trabecular bone microstructure assessment. Methods: Five sectioned maxilla of adult Dorper male sheep were scanned using a CBCT system with a resolution of 76 μm3 (Kodak 9000). The CBCT images were reconstructed using different reconstruction parameters and analysed. The effect of reconstruction voxel size (76, 100 and 200 μm3) and threshold values (±15% from the global threshold value) on trabecular bone microstructure measurement was assessed using image analysis software (CT analyser version 1.15). Results: There was no significant difference in trabecular bone microstructure measurement between the reconstruction voxel sizes, but a significant difference (Tb.N = 0.03, Tb.Sp = 0.04, Tb.Th = 0.01, BV/TV = 0.00) was apparent when the global threshold value was decreased by 15%. Conclusions: Trabecular bone microstructure measurements are not compromised by changing the CBCT reconstruction voxel size. However, measurements can be affected when applying a threshold value of less than 15% of the recommended global value