Objective The aim of this study was to evaluate the expression of selected salivary oncomiRNAs among smokeless tobacco users and nonsmokers. Materials and Methods Twenty-five subjects with chronic smokeless tobacco habit (> 1 year) and 25 nonsmokers were selected for this study. MicroRNA was extracted from saliva samples using the miRNeasy Kit (Qiagen, Hilden, Germany). The forward primers used in the reactions include hsa-miR-21-5p, hsa-miR-146a-3p, hsa-miR-155-3p, and hsa-miR-199a-3p. Relative expression of miRNAs was calculated using the 2-ΔΔCt method. Fold change is calculated by raising 2 to the power of the negative ΔΔCT value. Statistical Analysis Statistical analysis was carried out using GraphPad Prism 5 software. A p-value less than 0.05 was considered statistically significant. Results The four tested miRNAs were found overexpressed in saliva of subjects with smokeless tobacco habit when compared with saliva from nontobacco users. miR-21 expression was 3.74 ± 2.26 folds higher among subjects with smokeless tobacco habit compared to nontobacco users (p < 0.01). The expression for miR-146a (5.56 ± 8.3 folds; p < 0.05), miR-155 (8.06 ± 23.4 folds; p < 0.0001) and miR-199a (14.39 ± 30.3 folds; p < 0.05) was significantly higher among subjects with smokeless tobacco habit. Conclusion Smokeless tobacco leads to salivary overexpression of the miRs 21, 146a, 155, and 199a. Monitoring the levels of these four oncomiRs may provide insight about the future development of oral squamous cell carcinoma, especially in patients with smokeless tobacco habits.

Increased oxidative stress in bone cells is known to negatively alter favorable bone regeneration. This study aimed to develop a porous polycaprolactone (PCL) membrane incorporated with 25 wt % Vitamin C (PCL-Vit C) and compared it to the PCL membrane to control oxidative stress and enhance biomineralization in vitro. Both membranes were characterized using SEM-EDS, FTIR spectroscopy, and surface hydrophilicity. Vitamin C release was quantified colorimetrically. Assessments of the viability and attachment of human fetal osteoblast (hFOB 1.19) cells were carried out using XTT assay, SEM, and confocal microscopy, respectively. ROS generation and wound healing percentage were measured using flow cytometry and ImageJ software, respectively. Mineralization study using Alizarin Red in the presence or absence of osteogenic media was carried out to measure the calcium content. Alkaline phosphatase assay and gene expression of osteogenic markers (alkaline phosphatase (ALP), collagen Type I (Col1), runt-related transcription factor 2 (RUNX2), osteocalcin (OCN), and osteopontin (OPN)) were analyzed by real-time PCR. SEM images revealed smooth, fine, bead-free fibers in both membranes. The FTIR spectrum of pure vitamin C was replaced with peaks at 3436.05 and 2322.83 cm^-1 in the PCL-Vit C membrane. Vitamin C release was detected at 15 min and 1 h. The PCL-Vit C membrane was hydrophilic, generated lower ROS, and showed significantly higher viability than the PCL membrane. Although both PCL and PCL-Vit C membranes showed similar cellular and cytoskeletal morphology, more cell clusters were evident in the PCL-Vit C membrane. Lower ROS level in the PCL-Vit C membrane displayed improved cell functionality as evidenced by enhanced cellular differentiation with more intense alizarin staining and higher calcium content, supported by upregulation of osteogenic markers ALP, Col1, and OPN even in the absence of osteogenic supplements. The presence of Vitamin C in the PCL-Vit C membrane may have mitigated oxidative stress in hFOB 1.19 cells, resulting in enhanced biomineralization facilitating bone regeneration.

Titanium Dental implant is the most successful treatment modality to replace missing teeth today. Although titanium is considered biologically biocompatible, strong, and corrosion-free, the risk of implant failure continues due to bone loss at the expense of optimum oral health. Current research points toward the presence of titanium dioxide (TiO₂) particles leached from dental implant surface, which occurred due to mechanical and chemical insults on the surface. This study aimed to investigate the influence of TiO₂ particles of different sizes leaching from implant surfaces on Human Osteoblast cells (HOB) in-vitro. Titanium dioxide particles in both nano (NPs) and micro (MPs) size and at different concentrations were introduced to human osteoblast cells with and without treatment with vitamin C. Production of ROS was measured using H2DCFDA cellular ROS Assay Kit and MCP-1 and IL-8 cytokines released were assayed at 24 h time point using ELISA technique. Results showed a dose dependent increase in ROS production following exposure of HOB to both nano and micro particles. MCP-1 and IL-8 were released and there was minimal difference between the amount generated by nano compared with micro size particles. Treatment of HOB with antioxidant vitamin C demonstrated a significant reduction in the generation of ROS. At the same time, MCP-1 release was reduced significantly for the 100 μg/mL TiO₂ NPs and MPs after Vitamin C treatment while IL-8 release increased significantly. This study suggests a positive role played by antioxidants in the control of ROS generation and chemokines production in the peri-implant tissue environment.

Implantation of biomaterials generates reactive oxygen species (ROS) in the peri-implant microenvironment. Bone loss occurs when ROS levels exceed the local antioxidant capacity. The aim of this study was to investigate the influence of vitamin C-incorporated polycaprolactone (PCL-Vit C) membrane in scavenging ROS and enhancing biomineralisation in osteoblast-osteoclast (OB-OC) co-culture system. OB-OCs were cultured on polycaprolactone (PCL) and PCL-Vit C membranes under osteogenic conditions to mimic the bone microenvironment. ROS generation was measured using flow cytometry. ALP and the RANKL/OPG ratio were determined by colorimetric and ELISA assays, respectively. Gene expression of ALP, Col1, Runx-2 and OCN and protein expression of Runx-2, BMP-7, Col1 and OCN were determined by real-time PCR and western blotting, respectively. Activation of P38, ERK and JNK and beta-catenin expression was also analysed. OB-OC grown on PCL membrane generated a higher amount of ROS compared to those on PCL-Vit C. Colorimetric assays revealed a significantly higher ALP activity in OB-OC co-cultures on PCL-Vit C membranes. Furthermore, OB-OC grown on PCL-Vit C membrane showed significant upregulation in mRNA levels of ALP, Col1 and OCN with lower RANKL/OPG ratio and higher amounts of mineralisation nodules. Runx-2 expression was comparable in both membranes. Western blotting showed a significant increase in phosphorylation of P38 MAPK, ERK, JNK and beta-catenin expression in OB-OC maintained on PCL membrane, plausibly due to increased ROS levels, compared to PCL-Vit C. OB-OC grown on PCL-Vit C membrane scavenged ROS and supported a higher osteogenic potential environment for OB-OC co-culture in vitro.

The current study elucidates potential alterations in inflammatory cytokines and dopamine (DA) levels in saliva following a 21-day fasting regimen during Ramadan and explores their associations with mental health parameters.Forty-four healthy young university students (22 females and 22 males; median age 22 (2) and body mass index 23.40 (6.23) were enrolled, and saliva samples were collected before and after fasting. Cytokine and DA levels were quantified by ELISA and LEGENDplex Human Inflammation Panel, respectively. Participants also completed the Patient Health Questionnaire-9 and Depression Anxiety Stress Scales questionnaires to assess mental health.Data analysis was performed using SPSS. Differences between pre- and postfasting were tested using Wilcoxon's signed-rank test. Mann–Whitney's U test determined disparities in DA and cytokine levels across sex. Simple and multiple linear regression analyses were performed to identify the factors influencing the change in DA. Correlation analysis was employed to explore the relationships between the changes in DA and cytokine levels before and after fasting.A significant increase in inflammatory cytokines such as interleukin (IL)-1β, interferon-α2, tumor necrosis factor-α, IL-23, IL-33, and IL-8 was observed after fasting. Anti-inflammatory cytokine, IL-10, levels remained unchanged. Females had significantly higher levels of proinflammatory cytokines before fasting compared with males, but no significant gender differences were observed after fasting. The current study also showed a significant decrease in DA levels after fasting; however, no significant difference in DA levels across genders was noted. Self-reported mental health status did not significantly change before and after fasting. Multiple linear regression analyses did not suggest potential associations between these variables and changes in DA levels before and after fasting. However, correlation analysis indicated that the change in inflammatory cytokine levels was inversely related to changes in DA levels.Fasting during Ramadan significantly increased salivary cytokine levels and decreased DA levels, indicating potential relationships between immune factors and mental well-being. The findings highlight the complex interplay between inflammation, immunity, and DA regulation during fasting. Further research is warranted to elucidate the potential long-term effects of these associations and their implications for mental health and well-being.