OBJECTIVE: This study aimed to determine the potential of ascorbic acid alone in inducing differentially expressed osteoblast-related proteins in dental stem cells via the liquid chromatography-mass spectrometry/ mass spectrometry (LC-MS/MS) approach.
METHODS: The cells were isolated from deciduous (SHED) and permanent teeth (DPSC) and induced with 10 μg/mL of ascorbic acid. Bone mineralisation and osteoblast gene expression were determined using von Kossa staining and reverse transcriptase-polymerase chain reaction. The label-free protein samples were harvested on days 7 and 21, followed by protein identification and quantification using LC-MS/MS. Based on the similar protein expressed throughout treatment and controls for SHED and DPSC, overall biological processes followed by osteoblast-related protein abundance were determined using the PANTHER database. STRING database was performed to determine differentially expressed proteins as candidates for SHED and DPSC during osteoblast development.
RESULTS: Both cells indicated brownish mineral stain and expression of osteoblast-related genes on day 21. Overall, a total of 700 proteins were similar among all treatments on days 7 and 21, with 482 proteins appearing in the PANTHER database. Osteoblast-related protein abundance indicated 31 and 14 proteins related to SHED and DPSC, respectively. Further analysis by the STRING database identified only 22 and 11 proteins from the respective group. Differential expressed analysis of similar proteins from these two groups revealed ACTN4 and ACTN1 as proteins involved in both SHED and DPSC. In addition, three (PSMD11/RPN11, PLS3, and CLIC1) and one (SYNCRIP) protein were differentially expressed specifically for SHED and DPSC, respectively.
CONCLUSION: Proteome differential expression showed that ascorbic acid alone could induce osteoblastrelated proteins in SHED and DPSC and generate specific differentially expressed protein markers.
METHOD: In vitro cell viability, morphology, and alkaline phosphatase (ALP) activity of MC3T3-E1 cells on HA and PCL scaffolds were determined in comparison to the accepted model outlined for two-dimensional systems. An in vivo study involving the transplantation of MC3T3-E1 cells with scaffolds into an artificial bone defect of 4 mm length and 1.5 mm depth in the rat's left maxilla was conducted. Three-dimensional analysis using micro-computed tomography (micro-CT), hematoxylin and eosin (H&E), and immunohistochemistry analyses evaluation were performed after six weeks of transplantation.
RESULTS: MC3T3-E1 cells on the HA scaffold showed the highest cell viability. The cell viability on both scaffolds decreased after 14 days of culture, which reflects the dominant occurrence of osteoblast differentiation. An early sign of osteoblast differentiation can be detected on the PCL scaffold. However, cells on the HA scaffold showed more prominent results with intense mineralized nodules and significantly (p
METHODS: This review was performed following the PRISMA guidelines. A systematic search of the study was conducted by retrieving articles from the electronic databases PubMed and Web of Science to identify articles focussed on gene expression and approaches for osteoblast and osteoclast differentiation.
RESULTS: Six articles were included in this review; there were original articles of in vitro human stem cell differentiation into osteoblasts and osteoclasts that involved gene expression profiling. Quantitative polymerase chain reaction (qPCR) was the most used technique for gene expression to detect differentiated human osteoblasts and osteoclasts. A total of 16 genes were found to be related to differentiating osteoblast and osteoclast differentiation.
CONCLUSION: Qualitative information of gene expression provided by qPCR could become a standard technique to analyse the differentiation of human stem cells into osteoblasts and osteoclasts rather than evaluating relative gene expression. RUNX2 and CTSK could be applied to detect osteoblasts and osteoclasts, respectively, while RANKL could be applied to detect both osteoblasts and osteoclasts. This review provides future researchers with a central source of relevant information on the vast variety of gene expression approaches in analysing the differentiation of human osteoblast and osteoclast cells. In addition, these findings should enable researchers to conduct accurately and efficiently studies involving isolated human stem cell differentiation into osteoblasts and osteoclasts.
BACKGROUND: High background signals resulting from nonspecific binding are a significant concern for microtiter plate-based enzyme-linked lectin sorbent assays (ELLSAs), as they can mask specific binding signals and cause false-positive results.
METHODS: In this study, we constructed an ELLSA based on different washing step parameters, including the number of washing cycles, NaCl and Tween-20 concentrations, and the type of blocking agent and evaluated the effects on both specific and nonspecific binding signals. Furthermore, we performed a PSA binding assay using the optimized ELLSA.
RESULTS: The optimal washing parameters based on the highest specific binding signal proposed four cycles of washing steps using a washing buffer containing a high salt concentration (0.5 M NaCl) and mild detergent (0.05% Tween-20). The utilization of the optimized washing parameters in this assay was shown to be sufficient to obtain the optimal binding signals without the use of any blocking agent. Binding assays performed using the optimized ELLSA revealed that the glycan of the PSA sample used in this study mainly consists of terminal α2,6-linked sialic acid, as strongly recognized by Sambucus nigra agglutinin (SNA) with a KD value of 12.38 nM.
CONCLUSION: The ELLSA reported in this study provides a simple yet sensitive assay for sialic acid linkage recognition.
AIM: This study described characteristics of adulterated THM products in Malaysia and aimed to quantify THM products' safety signals of adverse reactions (ARs).
METHODS: THM products that were seized by Pharmacy Enforcement Division between 2008 and 2014 were extracted and analysed for 59,440 THM products. Of these, only 6452 THM products with complete information were included in the final analyses. Safety signalling tools were used to measure AR signals from AR reports obtained from the National Pharmaceutical Regulatory Agency Adverse Drug Reaction Database.
RESULTS: More than half of adulterated THM products originated from countries outside of Malaysia, with the majority were from Indonesia. The most common claimed indication of adulterated THM products was for pain and fever relief, while steroids were the most common adulterant. AR signals were generated for cough and cold products for respiratory and thoracic disorders, weight-loss products for cardiac disorders, and women's health products for reproductive and breast disorders.
CONCLUSION: Health authorities from various fields can work collaboratively by implementing strategic actions that include the use of safety signalling tools to curb the increasing number of adulterated THM products in the Malaysian market.
METHODS: Ten subjects (n = 10) who had upper and lower fixed appliances (MBT, 3 M Unitek, 0.022″ × 0.028″) were recruited for this study. Human gingival crevicular fluid (GCF) was obtained using periopaper strips at pre-treatment (T0), 1 month (T1), 3 months (T3), and 6 months (T6) of orthodontic treatment. Periapical radiographs of the upper permanent central incisors were taken at T0 and T6 to measure the amount of root resorption. Identification of changes in PA was performed using liquid chromatography-tandem mass spectrometry. Student's t-test was then performed to determine the significance of the differences in protein abundance before and after orthodontic treatment.
RESULTS: Our findings showed that all ten subjects had mild root resorption, with an average resorption length of 0.56 ± 0.30 mm. A total of 186 proteins were found to be commonly present at T0, T1, T3, and T6. There were significant changes in the abundance of 16 proteins (student's t-test, p ≤ 0.05). The increased PA of S100A9, immunoglobulin J chain, heat shock protein 1A, immunoglobulin heavy variable 4-34 and vitronectin at T1 suggested a response to stress that involved inflammation during the early phase of orthodontic treatment. On the other hand, the increased PA of thymidine phosphorylase at T3 suggested growth promotion and, angiogenic and chemotactic activities.
CONCLUSIONS: The identified proteins can be potential early markers for root resorption based on the increase in their respective PA and predicted roles during the early phase of orthodontic treatment. Non-invasive detection of root resorption using protein markers as early as possible is extremely important as it can aid orthodontists in successful orthodontic treatment.
METHOD: DPSC from murine incisors were isolated through either the outgrowth (DPSC-OG) or the enzymatic digestion (DPSC-ED) method. Cells at passage 4 were used in this study. The cells were characterized through morphology and expression of cell surface markers. The cells' doubling time when cultured using different seeding densities was calculated and analyzed using one-way ANOVA and Tukey's multiple comparison post-test. The ability of cells to differentiate to chondrocyte and osteoblast was evaluated through staining and analysis on the matrices secreted.
RESULTS: Gene expression analysis showed that DPSC-OG and DPSC-ED expressed dental pulp mesenchymal stem cell markers, but not hematopoietic stem cell markers. The least number of cells that could have been used to culture DPSC-OG and DPSC-ED with the shortest doubling time was 5 × 10(2) cells/cm(2) (11.49 ± 2.16 h) and 1 × 10(2) cells/cm(2) (10.55 h ± 0.50), respectively. Chondrocytes differentiated from DPSC-ED produced 2 times more proteoglycan and at a faster rate than DPSC-OG. FTIR revealed that DPSC-ED differentiated into osteoblast also secreted matrix, which more resembled a calvaria.
DISCUSSION: Isolation approaches might have influenced the cell populations obtained. This, in turn, resulted in cells with different proliferation and differentiation capability. While both DPSC-OG and DPSC-ED expressed mesenchymal stem cell markers, the percentage of cells carrying each marker might have differed between the two methods. Regardless, enzymatic digestion clearly yielded cells with better characteristics than outgrowth.