Topical Delivery of Mesenchymal Stem Cells "Secretomes" in Wound Repair.

Wound healing requires a coordinated interplay among cells, growth factors, and extracellular matrix proteins. Central to this process is the endogenous mesenchymal stem cell (MSC), which coordinates the repair response by recruiting other host cells and secreting growth factors and matrix proteins. MSCs are self-renewing multipotent stem cells that can differentiate into various lineages of mesenchymal origin such as bone, cartilage, tendon, and fat. In addition to multilineage differentiation capacity, MSCs regulate immune response and inflammation and possess powerful tissue protective and reparative mechanisms, making these cells attractive for treatment of different diseases. The beneficial effect of exogenous MSCs on wound healing was observed in a variety of animal models and in reported clinical cases. Specifically, they have been successfully used to treat chronic wounds and stimulate stalled healing processes. Recent studies revealed that human placental membranes are a rich source of MSCs for tissue regeneration and repair. This review provides a concise summary of current knowledge of biological properties of MSCs and describes the use of MSCs for wound healing. In particular, the scope of this review focuses on the role MSCs have in each phase of the wound-healing process, and clinical reports transplatation MSCs - secretomes in chronical ulcer.

Potential Use of L-Arginine Amino Acids towards Proliferation and Migratory Speed Rate of Human Dental Pulp Stem Cells (hDPSCs).

OBJECTIVE: Inflamed pulp has a decreased amount of protein and amino acids. L-Arginine is a semi-essential amino acid, the production of which is insufficient under oxidative stress and inflammation. L-Arginine is the sole substrate for nitric oxide synthase (NOS) to produce nitric oxide (NO), which plays an important role in cell proliferation and migration. This study aims to evaluate the proliferation and migration rate of hDPSCs after L-Arginine supplementation. METHODS: Serum-starved hDPSCs were divided into four groups: control: hDPSCs in Dulbecco's modified Eagle medium; hDPSCs in 300 µmol/L of the L-Arginine based culture media group; hDPSCs in 400 µmol/L of the L-Arginine based culture media group; and hDPSCs in 500 µmol/L of the L-Arginine based culture media group, which were planted in two separate 24-well-plates (5x104 cell/well) for proliferation and migration evaluation. The proliferation of all groups was measured by using a cell count test (haemacytometer and manual checker) after 24 h. The migratory speed rate of all groups was measured by using cell migration assay (scratch wound assay) after 24 h. Cell characteristics were evaluated under microscope (Inverted microscope, Zeiss®, Observer Z1, UK) that can be read through image-J® interpretation. This image J represented the measurement of migratory speed rate (nm/h) data. Statistical analysis was conducted using one-way ANOVA and post hoc Bonferroni (p<0.05) for proliferation and post hoc LSD (p<0.05) for migration (IBM SPSS Statistics Software, version 22.0). RESULTS: There was a statistically significant difference in hDPSC proliferation among various concentration groups of the L-Arginine based solution (300, 400 and 500 µmol/L) compared to the control group. There was a statistically significant difference in the migratory speed rate of hDPSCs in 500 µmol/L of the L-Arginine based solution group compared to lower concentrations and control group. CONCLUSION: The highest potency of hDPSC proliferation and migration was observed in the 500 µmol/L group. (EEJ-2023-01-08).

Resolving trapdoor phenomenon without secondary procedure after forehead flap on medial canthal region.

Herein, we present a case of basal cell carcinoma in a 59-yearold woman. It presented with painless itchy, black, gradually enlarged patches which were easily bled under her left eye since three years ago. A dermatological examination of the left medial canthus region obtained hyperpigmented plaques (2x0.8x0.1 cm) with uneven skin texture, irregular borders, and erosion on the center of the lesion. We performed forehead flap technique surgery followed by eight-month monitoring, resulting in a satisfying outcome in both function and appearance. The thinning technique and adjusting the flap size from the forehead area to the medial canthus should be as thin as possible to avoid differences in skin thickness and post-reconstruction hypertrophic scars. A bulging appears on the surgical site a month after the procedure, known as the trapdoor phenomenon, on the 8th month of followup, the trapdoor phenomenon disappeared.

Analysis of Thrombin-Activated Platelet-Derived Exosome (T-aPDE) Potential for Dental Pulp Regeneration: In-Vitro Study.

OBJECTIVE: This study analyzed the potential of various concentrations of the thrombin-activated platelet-derived exosome (T-aPDE) to regenerate the dental pulp by performing an in-vitro analysis of the cell viability, migration activity, and vascular endothelial growth factor A (VEGF-A) expression of human dental pulp stem cells (hDPSCs). MATERIAL AND METHODS: The hDPSCs were collected from nine third molar teeth of nine healthy donors and were isolated and cultured using the explant method. They were harvested between the third and fourth passages and starved, after which they were seeded in the following treatments: Dulbecco's Modified Eagle Medium and 10% platelet-rich plasma-thrombin as the control groups, and 0.5, 1, and 5% T-aPDE as the experimental groups. All groups had three biological triplicates (Triplo) and two number of experiments. The T-aPDE was analyzed using transmission electron microscopy testing, particle size analyzer, and CD63 + and CD81 + specific immune phenotyping flow cytometry tests for plasma exosomes. The cell viability was evaluated using the colorimetric assay of activity cellular enzymes (MTT assay); the migration activity, using scratch assay; and the VEGF-A expression, using enzyme-linked immunosorbent assay. RESULTS: The highest viability absorbance value of hDPSCs after 24, 48, 72 hours of observation was in the 5% T-aPDE group (p<0.05). Whereas, the closest distance result of migratory activation hDPSCs was also in the same group (p<0.05). However the highest VEGF-A expression of hDSPCs was noted in the same group at 72 hours observation (p<0.05). STATISTICAL ANALYSIS: The data were analyzed using one-way analysis of variance and the Kruskal-Wallis test. The statistical power was set at p <0.05 CONCLUSION: The 5% T-aPDE had a higher potential to induce dental pulp regeneration than the other groups.

Viability of human dental pulp stem cells: The potential of L-arginine-based culture media.

Dental pulp is built by proteins that have various roles in the biological process of pulp, such as structural protein, regulation protein, and catalytic protein. L-arginine, an amino acid and one of the building blocks of proteins, regulates pro-inflammatory and anti-inflammatory activity. Therefore, L-arginine-based culture has potential to promote dental pulp regeneration. This study aimed to investigate the potential of L-arginine-based culture in improving the viability of human dental pulp stem cells (hDPSCs). We evaluated the viability of hDPSCs in culture media supplemented with different concentrations of L-arginine amino acid (250, 300, 350, and 400 µmol/L) and Dulbecco's Modified Eagle Medium plus fetal bovine serum 10% (control) using a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay after 24-h incubation time. Statistical analysis was conducted using a one-way analysis of variance and post hoc least significant difference test. In qualitative analysis, the 4´, 6-diamidino-2-phenylindole staining method was used. The evaluation has shown a significant result when 250, 300, and 350 µmol/L concentration of L-arginine amino acid culture media compared with control, and 400 µmol/L has the best result and was not significantly different with control toward viability of hDPSCs.

The Effect of Hydrogel Hyaluronic Acid on Dentine Sialophosphoprotein Expression of Human Dental Pulp Stem Cells.

OBJECTIVE: Hyaluronic acid (HA) is glycosaminoglycan and one of important factors in extracellular matrix. In an inflamed pulp, when niche biology is conducive, the recruitment of human dental pulp stem cells (hDPSCs) will take place and differentiate into odontoblast like cell, creating reparative dentine and expressing dentine sialophosphoprotein (DSPP). Therefore, the purpose of this study was to analyze the potential of hydrogel HA in various concentration towards hDPSCs differentiation via DSPP expression at day 7 and 14. METHODS: After hDPSCs incubation reaching 80% confluence, cells were then starved for 24 hours. Then, culture media were supplemented with osteogenic media. hDPSCs planted into 96 well plate and HA 10 µg/mL, 20 µg/mL, and 30 µg/mL were added. DSPP expression was analysed using elisa reader at day 7 and 14, qualitative result was analysed using alizarin red at day 21. Data was analysed using one-way ANOVA. RESULTS: At day 7, there was a statistically significant different potential of HA conditioned media in various concentration (p<0.05) towards hDPSCs differentiation via expression of DSPP with HA 30 µg/mL being the most potential concentration to increase DSPP expression. CONCLUSION: HA have the potential to increase odontoblast differentiation process via expression of DSPP, with HA 30 µg/mL being the optimum concentration for hDPSCs. (EEJ-2022-12-169).

The effect of hyaluronic acid conditioned media on hDPSCs differentiation through CD44 and transforming growth factor-ß1 expressions.

Hyaluronic acid (HA) has the capability to influence dentin niche which is important in regenerative process. The CD44 as a specific receptor of HA was found to be related to dentin mineralization process. Meanwhile, transforming growth factor ß1 (TGF-ß1) has a vital role in the transition from proliferation into the differentiation of human dental pulp stem cell human dental pulp stem cells (hDPSCs) to become odontoblast cells and dentin mineralization. This study aims to analyzed HA's effect on dentin mineralization through CD44 and TGF-ß1 expressions. Stem cells were cultured in four different supplemented conditioned media (control, +10 µg/mL, +20 µg/mL, and + 30 µg/mL of HA). Evaluation of CD44 expression was analyzed using flow cytometry and TGF-ß1 was analyzed using enzyme-linked immunosorbent assay reader. Qualitative result using Alizarin red test after 21 days was done to confirm the formation of mineralization nodules. It was shown that HA expression of CD44 and TGF-ß1 on hDPSCs were higher in AH groups compared to the control group and 30 µg/mL HA induced the highest TGF-ß1 expression on hDPSCs. Alizarin red test also showed the highest mineralization nodules in the same group. Therefore, from this study, we found that supplemented 30 µg/mL of HA was proved in initiating hDPSCs differentiation process and promote dentin mineralization.

The Effect of Calcium Gluconate on Platelet Rich Plasma Activation for VEGF-A Expression of Human Dental Pulp Stem Cells.

OBJECTIVE: Platelet-rich plasma (PRP) activation is an important factor in triggering the initial release of blood-derived growth factors from platelets. Vascular endothelial growth factor-A (VEGF-A) can be expressed by human dental pulp stem cells (hDPSCs) and plays an important role in dental pulp angiogenesis. The aim of this study is to analyze the effects of calcium gluconate on PRP activation in hDPSC VEGF-A expression. MATERIALS AND METHODS: Two types of PRP and their corresponding activators were analyzed in this study: PRP (activated using calcium chloride/CaCl2) and PRP-T (activated using CaCl2 with the addition of 10% calcium gluconate). hDPSCs were obtained by using an out-growth method (DPSCs-OG), and harvest between the fifth and sixth passages, then cultured in three different media groups: control, PRP, and PRP-T, which were planted in 96 wells (5 × 103 each well). The VEGF-A expression of hDPSCs was analyzed by using an ELISA test and observed at 24, 48, and 72 hours. STATISTICAL ANALYSIS: This study was performed by using one-way ANOVA (p < 0.05) test. RESULTS: There were significant differences between all groups (p < 0.05) at 48 and 72 hours of observations, and no significant differences in the PRP and PRP-T groups at 48 and 72 hours of observations (p > 0.05). CONCLUSION: PRP and PRP-T were equally effective in inducing VEGF-A expression of hDPSCs.

The Difference in Interleukin-19 Serum on Degrees of Acne Vulgaris Severity.

INTRODUCTION: Acne vulgaris is a multifactorial disease. Recent study showed that inflammation does have a central role in the formation of both inflammatory and noninflammatory lesions in acne vulgaris. There are various findings of proinflammatory cytokines related to acne vulgaris, but no previous study correlate interleukin- (IL-) 19 to acne vulgaris. This pilot study aims to look at difference in IL-19 serum concentration on degrees of severity of acne vulgaris. METHODS: This is an analytical observational cross-sectional study. Sample subjects were patients with acne vulgaris who met the inclusion criteria. Enzyme-linked immunosorbent assay (ELISA) study was applied to measure IL-19 serum. RESULT: Analysis test found statistically significant difference between IL-19 serum concentration of group of patients with mild acne vulgaris and that of group of patients with severe acne vulgaris. Moreover, analysis revealed significant difference between IL-19 serum concentration of group of patients with moderate acne vulgaris and that of group of patients with severe acne vulgaris. CONCLUSIONS: There are differences in serum levels of IL-19 on the severity of acne vulgaris. The significant difference might show that inflammation has a core role in severity of acne vulgaris, and IL-19 might potentially be related to acne vulgaris.

Comparison of Transforming Growth Factor 1 (TGF-β 1) Expression in Various Lysate Platelet-Rich Fibrin (L-PRF) Concentrations on Human Dental Pulp Stem Cell Differentiation

ABSTRACT Objective: To compare Transforming growth factor-β1 (TGF-β1) expression in various L-PRF concentrations on the hDPSC differentiation process. Material and Methods: hDPSC cell cultures were subjected to serum starvation by reducing FBS levels in the hDPSC culture media. Lysate PRF was obtained from the PRF gel, which was then incubated at 4°C for 24 h. The supernatant was dried, transferred to a 2-ml Eppendorf tube, and stored at −20°C. The evaluation of TGF-β1 expression in 1%, 5%, 10%, and 25% L-PRF samples and 10% FBS (control) during the process of hDPSC differentiation was quantified using an ELISA reader on day 7. The expression of TGF-β1 was subjected to a one-way ANOVA test, followed by Bonferroni's post hoc test with significant values (p<0.05). Results: Significant differences were noted in TGF-β1 expression between 1%, 5%, 10%, and 25% L-PRF and the control group (10% FBS). The highest TGF-β1 expression occurred with 25% L-PRF (0.645 ± 0.048), followed by 10% L-PRF (0.461 ± 0.035), 10% FBS (0.374 ± 0.013), 5% L-PRF (0.275 ± 0.045), and the lowest expression was with 1% L-PRF (0.160 ± 0.045). Conclusion: The best result of TGF-B1 expression in hDPSC differentiation was in the 25% L-PRF group.

Ideal Concentration of Advanced-Platelet Rich Fibrin (A-PRF) Conditioned Media for Human Dental Pulp Stem Cells Differentiation

Abstract Objective: To discover the ideal concentration of Advanced Platelet Rich Fibrin (A-PRF) as modification of PRF, for human Dental Pulp Stem Cells (hDPSCs) differentiation. Material and Methods: hDPSCs were devided into five experimental groups: Group I (control group) consist of hDPSCs cultured in 10% FBS, Group II consist of hDPSCs cultured in 1% A-PRF, Group III consist of hDPSCs cultured in 5% A-PRF, Group IV consist of hDPSCs cultured in 10% A-PRF and Group V consist of hDPSCs cultured in 25% A-APRF. All group have been observed for 7 and 14 days and each group had three biological replicates (triplo). Formation of the mineralized nodules was detected after 7 days by Alizarin red-based assay and Dentin Sialophosphoprotein (DSPP) expression after 7 and 14 days quantified by ELISA reader. Statistical analysis was proven with Kruskal-Wallis and post hoc Mann-Whitney test Results: The differentiation of hDPSCs in all A-PRF groups was significantly different on day-7 (p<0.05) compare to control group (Group I). There were no significant differences between all groups on day-14 (p>0.05). Significantly differences between Group II (1% A-PRF) and Group I (control), Group II (1% A-PRF) and Group III (5% A-PRF), also Group II (1% A-PRF) and Group V (25% A-PRF) was found from post hoc test analysis Conclusion: The ideal conditioned media concentration for differentiation of human dental pulp stem cells was on 1% up to 5% A-PRF group.