Gingival Tissue MiRNA Expression Profiling and an Analysis of Periodontitis-Specific Circulating MiRNAs.

This study aimed to identify the microRNAs (miRNAs) associated with periodontitis (PD) in gingival tissues, and to evaluate the levels of these selected miRNAs in the saliva and blood plasma among participants with and without rheumatoid arthritis (RA). A genome-wide miRNA expression analysis in 16 gingival tissue samples revealed 177 deregulated miRNAs. The validation of the miRNA profiling results in 80 gingival tissue samples revealed that the PD-affected tissues had a higher expression of miR-140-3p and -145-5p, while the levels of miR-125a-3p were significantly lower in inflamed tissues. After a thorough validation, four miRNAs, namely miR-140-3p, -145-5p, -146a-5p, and -195-5p, were selected for further analysis in a larger sample of salivary (N = 173) and blood plasma (N = 221) specimens. Increased salivary levels of miR-145-5p were associated with higher mean values of pocket probing depth and bleeding on probing index. The plasma-derived levels of miR-140-3p were higher among the participants with PD. In conclusion, the gingival levels of miR-140-3p, -145-5p, and -125a-3p were independently associated with PD presence and severity. The salivary and blood plasma levels of the target miRNAs were diversely related to PD. Similar miRNA associations with PD were observed among the participants with and without RA.

SARS-CoV-2 RT-qPCR Ct values in saliva and nasopharyngeal swab samples for disease severity prediction.

Background: Comparison of clinical value of RT-qPCR-based SARS-CoV-2 tests performed on saliva samples (SSs) and nasopharyngeal swab samples (NPSs) for prediction of the COVID-19 disease severity. Methods: Three paired SSs and NPSs collected every 3 days from 100 hospitalised COVID-19 patients during 2020 Jul-2021 Jan were tested by RT-qPCR for the original SARS-CoV-2 virus and compared to 150 healthy controls. Cases were divided into mild+moderate (Cohort I, N = 47) and severe disease (Cohort II, N = 53) cohorts and compared. Results: SARS-CoV-2 was detected in 65% (91/140) vs. 53% (82/156) of NPSs and 49% (68/139) vs. 48% (75/157) of SSs collected from Cohort I and II, respectively, resulting in the total respective detection rates of 58% (173/296) vs. 48% (143/296) (P = 0.017). Ct values of SSs were lower than those of NPSs (mean Ct = 28.01 vs. 30.07, P = 0.002). Although Ct values of the first SSs were significantly lower in Cohort I than in Cohort II (P = 0.04), it became negative earlier (mean 11.7 vs. 14.8 days, P = 0.005). Multivariate Cox proportional hazards regression analysis showed that Ct value ≤30 from SSs was the independent predictor for severe COVID-19 (HR = 10.06, 95% CI: 1.84-55.14, P = 0.008). Conclusion: Salivary RT-qPCR testing is suitable for SARS-CoV-2 infection control, while simple measurement of Ct values can assist in prediction of COVID-19 severity.

Analysis of Intrinsic Breast Cancer Subtypes: The Clinical Utility of Epigenetic Biomarkers and TP53 Mutation Status in Triple-Negative Cases.

This study aimed at analyzing the DNA methylation pattern and TP53 mutation status of intrinsic breast cancer (BC) subtypes for improved characterization and survival prediction. DNA methylation of 17 genes was tested by methylation-specific PCR in 116 non-familial BRCA mutation-negative BC and 29 control noncancerous cases. At least one gene methylation was detected in all BC specimens and a 10-gene panel statistically significantly separated tumors from noncancerous breast tissues. Methylation of FILIP1L and MT1E was predominant in triple-negative (TN) BC, while other BC subtypes were characterized by RASSF1, PRKCB, MT1G, APC, and RUNX3 hypermethylation. TP53 mutation (TP53-mut) was found in 38% of sequenced samples and mainly affected TN BC cases (87%). Cox analysis revealed that TN status, age at diagnosis, and RUNX3 methylation are independent prognostic factors for overall survival (OS) in BC. The combinations of methylated biomarkers, RUNX3 with MT1E or FILIP1L, were also predictive for shorter OS, whereas methylated FILIP1L was predictive of a poor outcome in the TP53-mut subgroup. Therefore, DNA methylation patterns of specific genes significantly separate BC from noncancerous breast tissues and distinguishes TN cases from non-TN BC, whereas the combination of two-to-three epigenetic biomarkers can be an informative tool for BC outcome predictions.

Analysis of Epigenetic Changes in Vitamin D Pathway Genes in Rheumatoid Arthritis Patients.

Background: Rheumatoid arthritis (RA) is an autoimmune inflammatory disease with complex etiopathogenesis launched by multiple risk factors, including epigenetic alterations. RA is possibly linked to vitamin D that is epigenetically active and may alter DNA methylation of certain genes. Therefore, the study aimed to evaluate the relationship between DNA methylation status of vitamin D signaling pathway genes (VDR, CYP24A1, CYP2R1), vitamin D level and associations with RA. Materials and Methods: Totally 76 participants (35 RA patients and 41 healthy controls) were enrolled from a case-control vitamin D and VDR gene polymorphisms study regarding age and vitamin D concentration. CpG islands in promoter regions of the VDR, CYP24A1, CYP2R1 genes were chosen for DNA methylation analysis by means of pyrosequencing. Chemiluminescent microplate immunoassay was used to assess 25(OH)D serum levels. RA clinical data, i.e. the disease activity score C-reactive protein 28 (DAS28 - CRP) as well as patient-reported outcome questionnaires were recorded. Results: The study showed similar methylation pattern in the promoter regions of vitamin D pathway genes in RA and control group with p>0.05 (VDR gene 2.39% vs. 2.48%, CYP24A1 gene 16.02% vs. 15.17% and CYP2R1 2.53% vs. 2.41%). CYP24A1 methylation intensity was significantly higher in compare to methylation intensity of VDR and CYP2R1 genes in both groups (p<0.0001). A tendency of higher vitamin D concentration in cases having methylated VDR (57.57±28.93 vs. 47.40±29.88 nmol/l), CYP24A1 (53.23±26.22 vs. 48.23±34.41 nmol/l) and CYP2R1 (60.41±30.73 vs. 44.54±27.63 nmol/l) genes and a positive correlation between VDR, CYP2R1 methylation intensity and vitamin D level in RA affected participants was revealed (p>0.05). A significantly higher CYP24A1 methylation intensity (p=0.0104) was detected in blood cells of vitamin D deficient (<50 nmol/l) RA patients vs. vitamin D deficient controls. Conclusions: Our data suggests some indirect associations between DNA methylation status of vitamin D pathway genes and vitamin D level in RA.

Gingival crevicular fluid microRNA associations with periodontitis.

PURPOSE: The present study was performed to assess the associations of gingival crevicular fluid (GCF) microRNAs miR-140-3p, miR-145-5p, miR-146a-5p, and miR-195-5p with periodontitis (PD) and to evaluate the possible influence of rheumatoid arthritis (RA) in this context. METHOD: GCF samples were collected from 134 individuals with PD and 76 periodontally healthy individuals, with or without RA. After miRNA extraction from GCF, the levels of miR-140-3p, miR-145-5p, miR-146a-5p, and miR-195-5p were assessed using RT-qPCR. RESULTS: MiR-146a-5p levels were significantly lower among the patients with PD than among the healthy individuals (P < 0.001) and negatively correlated with PD severity based on PD stage and periodontal outcome parameters (P < 0.05). Patients with severe PD had higher GCF levels of miR-140-3p and miR-145-5p than did periodontally healthy individuals (P < 0.05). Significant AUC values for diagnosis of severe PD were revealed for miR-140-3p (AUC = 0.614, P = 0.022), miR-145-5p (AUC = 0.621, P = 0.016) and miR-146a-5p (AUC = 0.702, P < 0.001). Combination of the aforementioned miRNAs increased the diagnostic performance (AUC = 0.709, P < 0.001). CONCLUSION: It was demonstrated that miR-140-3p, miR-145-5p and miR-146a-5p were associated with PD and would be potentially effective for GCF-based non-invasive periodontitis diagnostics in patients with and without RA.

Urinary microRNAs can predict response to abiraterone acetate in castration resistant prostate cancer: A pilot study.

BACKGROUND: Despite novel agents have been introduced to treat castration resistant prostate cancer (CRPC) during the last decade, up to one-third of CRPC patients face primary resistance to new generation compounds. Therefore, sensitive molecular tools are urgently needed for reliable treatment selection and response prediction. This study aimed to evaluate urinary miRNAs and blood circulating androgen receptor (AR) transcript level as a tool for noninvasive outcome prediction for CRPC patients undergoing abiraterone acetate (AA) therapy. METHODS: Prostate cancer-specific miR-148a, -365, -375, and -429 were analyzed in 129 urine samples collected from 100 CRPC patients before and during AA therapy via quantitative reverse transcription PCR. To test the prognostic value, urinary miRNA levels alone, as well as combined with AR level were associated with progression-free survival (PFS) and overall survival (OS). RESULTS: Level of urinary miR-375 was the highest in CRPC in comparison to noncancerous controls, as well as in combination with miR-429 was predictive for short PFS in AA-treated patients (HR = 2.2, 95% CI: 1.1-4.2, p = 0.023). Especially high prognostic power of all analyzed miRNAs was observed in CRPC cases with high blood AR levels. For PFS prediction a tandem of miR-429 and high AR reached HR of 5.0 (95% CI: 2.2-11.8, p < 0.001), while for prediction of OS the best combination was demonstrated by miR-148a and AR with HR of 3.1 (95% CI: 1.4-7.1, p = 0.006). CONCLUSIONS: Urinary miRNAs could be used as prognostic biomarkers for CRPC patients to predict response to AA therapy, especially for the cases with high blood AR levels.

Analysis of periodontitis-associated miRNAs in gingival tissue, gingival crevicular fluid, saliva and blood plasma.

OBJECTIVE: Periodontitis (PD) is a chronic inflammatory disease which is associated with multiple systemic comorbidities, including rheumatoid arthritis (RA), meanwhile the etiopathology of PD may be modulated by various factors including microRNA (miRNA). The present study aimed to reveal miRNAs associated with PD in gingival tissue, gingival crevicular fluid (GCF), saliva, plasma and to assess the possible influence of RA. DESIGN: The cross-sectional study included 30 patients with PD and 31 periodontally healthy participants. A total of 25 participants were additionally diagnosed with RA. Microarray analysis of eight gingival tissue samples was performed and four PD-associated miRNAs were selected: miR-199a-5p, miR-483-5p, miR-3198 and miR-4299. Target miRNAs were further assessed by means of RT-qPCR in 61 gingival tissue samples and corresponding bodily fluids - GCF, saliva and plasma. RESULTS: The upregulation of miR-199a-5p and downregulation of miR-4299 in gingival tissue was associated with the presence of PD and RA (P < 0.05). GCF level of miR-3198 was higher amongst participants with PD (P = 0.019) and showed a good diagnostic ability (AUC = 0.72, P = 0.008). Increased miR-199a-5p salivary level and decreased miR-199a-5p plasma level were observed amongst patients with worse clinical status of PD (P < 0.05). MiR-3198 and miR-4299 combination in GCF demonstrated AUC value of 0.86 and reached sensitivity of 68 % and specificity of 96 %. CONCLUSIONS: Aberrant expression of miR-199a-5p, miR-483-5p, miR-3198, miR-4299 in gingival tissues is associated with the presence and/or severity of PD. MiR-3198, miR-4299 level in GCF and miR-199a-5p level in plasma strongly correlated with PD, demonstrating significant diagnostic performance.