Long-term cryopreservation of whole gingival tissue.

Stem cells obtained from the body tissue, such as adipose tissue, dental pulp and gingival tissue. Fresh tissue is often used to isolate and culture for regenerative medicine. However, availability of tissue as and when required is one of the measure issue in regenerative medicine. Cryopreservation of tissue provides benefit over tissue availability, storage for significant amount of period and helps preserve the original cell structures. The effects of cryopreservation of gingival tissue for mesenchymal stem cell (MSC) are not well documented; however this process is of increasing importance for regenerative therapies. This study examined the effect of cryopreservation on the long term survival the whole gingival biopsy tissue. We studied cell outgrowth, cell morphology, MSC surface-markers and differentiation of mesenchymal stem cells derived from cryopreserved gingiva. In this study, gingival tissue was cryopreserved for 3, 6, 9 months. Cryopreserved tissue has been thawed and cells were isolated by using explant culture method. The fresh and cryopreserved gingival tissue cells were cultured and characterized for surface marker analysis, CFU-f, population doubling time, and osteogenic, chondrogenic and adipogenic differentiation. The fresh and cryopreserved tissue has similar stem cell properties. Results indicate that cryopreservation of the entire gingival tissue does not affect the properties of stem cells. This opens door for gingival tissue banking for future use in periodontology and regenerative medicine.

Stem Cell Secretome Modulated by Arsenicum album 30C Ameliorates Lipopolysaccharide-induced Cytokine Storm in Blood Mononuclear Cells in vitro.

BACKGROUND: The therapeutic effectiveness of mesenchymal stem cells (MSCs) and their secretome can be enhanced by means of physical, chemical and biological preconditioning. Arsenicum album 30C (AA30) has been one of the leading homeopathic medicines used in prophylaxis against SARS-CoV-2 infection. AIMS: This study aimed to investigate whether AA30 preconditioning could influence the growth factors and cytokine profile of the human dental pulp-derived MSC (DPD-MSC) secretome. Also, to test the efficacy of the AA30-preconditioned DPD-MSC secretome in ameliorating the lipopolysaccharide (LPS)-induced cytokine storm in human peripheral blood mononuclear cells (PBMCs) as an in-vitro cellular model. METHODS: The cytotoxicity of AA30 was assessed in DPD-MSCs by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Growth factors and cytokine levels in the AA30-preconditioned DPD-MSC secretome were analysed by fluorescence-activated cell sorting (FACS) analysis. The angiogenic potential of the AA30-preconditioned DPD-MSC secretome was assessed by chick yolk-sac membrane (YSM) assay. Culture medium with 0.001% ethanol was used as vehicle control. The efficacy of the AA30-preconditioned DPD-MSC secretome in ameliorating the cytokine storm was assessed in LPS pre-treated PBMCs. The mRNA and protein expression of inflammatory markers such as IL-1ß, IL-6 and IL-10 were analysed by using RT-PCR and FACS analysis respectively. RESULTS: AA30 did not exhibit cytotoxicity in the concentration range of 1% to 50%. Furthermore, the AA30-preconditioned DPD-MSC secretome exhibited a significant increase in the levels of angiogenic factors, such as human angiopoietin-2, EPO and PDGF-AA, and decreased levels of cytokines, such as TNF-α, CXCL-8 and IL-6. The AA30-preconditioned DPD-MSC secretome showed augmented angiogenesis compared to vehicle controls. The DPD-MSC secretome ameliorated LPS-induced mRNA and protein expression of IL-1ß, IL-6 and IL-10 in PBMCs. CONCLUSION: The AA30-preconditioned DPD-MSC secretome augmented angiogenesis and ameliorated the LPS-induced cytokine storm in human PBMCs in vitro. Our data demonstrate that AA30 preconditioning enhances the therapeutic potency of MSCs and their secretome.

Insulin-producing cell clusters derived from human gingival mesenchymal stem cells as a model for diabetes research.

BACKGROUND: The human gingiva-derived mesenchymal stem cells (hGMSCs) possess a great potential to develop the cell-based therapy for diabetes due to its unscarred healing capacity and reparative potential. In this current study, we isolated, cultured and characterised the GMSCs and explored their potential to differentiate into Insulin Producing Cell Clusters (IPCCs). METHODS: The cells derived from gingival tissues exhibited fibroblast-like morphology. The flow cytometric analysis revealed positive expression of CD73(97.43%), CD90(95.05%), and CD105(93.17%) and negative expression of CD34(0.05%), CD45(0.09%), and HLA-DR (0.025) surface markers. We then converted this adherent fibroblast-like GMSCs into floating IPCCs using a sequential three-step protocol containing a different combination of differentiating agents. Initially, the presence of insulin in IPCCs was confirmed by dithizone staining. Glucose-stimulated insulin secretion (GSIS) assay confirmed that IPCCs secrete insulin in response to glucose. RESULTS: Generated IPCCs express pancreatic markers such as insulin, pdx1, glucagon, GLUT4 and GLUT2 as evidenced by RT-PCR analysis. Our results unequivocally showed that IPCCs can be generated from gingiva which is a potential source of postnatal MSCs. Our results offer the IPCCs generated from hGMSCs a platform for screening anti-diabetic drugs and a new autologous source of tissue for islet transplantation for the treatment of diabetes. CONCLUSIONS: Our results unequivocally demonstrate for the first time that hGMSCs can be used as an attractive non-invasive tissue source for generating IPCCs, which can be employed in diabetes research for screening antidiabetic agents and also for transplantation in type 1 diabetic patients as autologous source without the need of immunosuppression.

Long term explant culture for harvesting homogeneous population of human dental pulp stem cells.

Dental pulp stem cells have emerged as a preferred source of mesenchymal stem cells, because of its easy availability and high stem cell content. Dental pulp is a specific fibrous tissue that contains heterogeneous populations of odontoblasts, fibroblasts, pericytes, progenitors, stem cells, leukocytes and neuronal cells. In this study, we propose sustained explant culture as a simple, economical and efficient process to isolate dental pulp stem cells from human Dental pulp Tissue. Historically explant cultures were used to get fibroblast cells from embryonic chick heart using plasma clot cultures. The subculture was performed by lifting mother explant (original explant) and grafting it in a new plasma clot. We modified this age old technique to suit the modern times. Here we demonstrate for the first time that the mother explant (E0) of human dental pulp tissue could be sub-cultured consecutively seven times (E7) without displacement. This technique is highly reproducible and permits growth and proliferation of dental pulp stem cells yielding an enriched homogeneous mesenchymal stem cells population in the first passage itself as revealed by surface marker expression. These dental pulp stem cells exhibit differentiation into adipogenic, chondrogenic and osteogenic lineage revealing their mesenchymal stem cell nature. We propose that dental pulp stem cells isolated by sustained explant culture are phenotypically and functionally comparable to those obtained by enzymatic method. It is a simple, inexpensive and gentle method, which may be preferred over the conventional techniques for obtaining stem cells from other tissue sources as well especially in cases of limited starting material.

Role of the Stem Cells from Human Deciduous Teeth in Dentistry: A Review.

Stem cells isolated from human exfoliated deciduous teeth or SHED possess high rate of proliferation along with high osteogenic as well as neurogenic differentiation when compared with dental pulpal stem cells. This stem cell population is an important source for therapeutic regeneration of tissues. Hence, the aim of present review article was to analyze the potential applications of SHED.

Decellularized leaf-based biomaterial supports osteogenic differentiation of dental pulp mesenchymal stem cells.

Decellularized tissues are an attractive scaffolds for 3D tissue engineering. Decellularized animal tissues have certain limitations such as the availability of tissue, high costs and ethical concerns related to the use of animal sources. Plant-based tissue decellularized scaffolds could be a better option to overcome the problem. The leaves of different plants offer a unique opportunity for the development of tissue-specific scaffolds, depending on the reticulate or parallel veination. Herein, we decellularized spinach leaves and employed these for the propagation and osteogenic differentiation of dental pulp stem cells (DPSCs). DPSCs were characterized by using mesenchymal stem cell surface markers CD90, CD105 and CD73 and CD34, CD45 and HLA-DR using flow cytometry. Spinach leaves were decellularized using ethanol, NaOH and HCL. Cytotoxicity of spinach leaf scaffolds were analysed by MTT assay. Decellularized spinach leaves supported dental pulp stem cell adhesion, proliferation and osteogenic differentiation. Our data demonstrate that the decellularized spinach cellulose scaffolds can stimulate the growth, proliferation and osteogenic differentiation of DPSCs. In this study, we showed the versatile nature of decellularized plant leaves as a biological scaffold and their potential for bone regeneration in vitro.

Evaluating the ability of cultivated odontoblasts to form dentin-like tissue in vitro using fibroblast growth factor and insulin-like growth factor.

Aim: The aim of the study was to evaluate the ability of cultivated odontoblast to form dentin-like tissue using fibroblast growth factor (FGF) and insulin-like growth factor (IGF). Materials and Methods: Dental pulp stem cells (DPSCs) were extracted from 10 human teeth. They were isolated and cultivated in vitro with the use of stem cell markers. The human DPSCs were characterized for trilineage differentiation. They were then differentiated into odontoblasts. The ability of cultivated odontoblasts to form dentin-like tissue was evaluated using FGF and IGF. Results: IGF showed superior ability to form dentin-like tissue as compared to FGF. The addition of FGF showed no significant difference in the formation of dentin-like tissue. A combination of FGF and IGF in odontoblast showed an enhanced ability to form dentin-like tissue. Conclusion: The use of growth factors IGF and FGF with dental stem cells showed a greater potential to form dentin-like tissue. This can profoundly alter the paradigms of conservative vital pulp therapy, which may eventually make it possible to treat dental diseases by regeneration of lost dentine.

Chemical preconditioning escalates chondrogenic activity in explant cultured human dental pulp stem cell study model for future temporomandibular joint regeneration.

Context: Human dental pulp stem cells (hDPSC) derived from dental pulp in conducive environment activated by chemicals can enhance chondrogenic cells for future animal model temporomandibular joint model. Aim: The study aims at evaluating the chemicals preconditioning (curcumin and rapamycin) efficacy toward chondrogenic proliferation of human dental pulp stem cells. Settings and Design: The in vitro study model with 10 premolar teeth extirpated pulp was processed under sterile chemical conditions. The cells viability was checked with calorimetric assay for adipogenic and chondrogenic, osteogenic lineages. The viability of the cells and the concentration of curcumin (CU) and rapamycin (RP) required for cell differentiation toward chondrogenic lineage were assessed. Material and Methods: The hDPSC was evaluated after explant long-term cultivation with characterization and chemical conditioning with dimethyl sulfoxide (DMSO) as control. MTT assay was used for cytotoxicity evaluation, cell viability, and proliferation. The dose optimization was observed with RP and CU. Chondrogenic proliferation was assessed with standard staining method of 0.1% Safranin O and 0.1% Alcian blue. Statistical Design: The flow cytometry analysis revealed good results for CD 90 compared to others. The intergroup analysis was done by ANOVA, and intragroup analysis was done by Post hoc Tukey's test. The intragroup analysis showed P value < 0.05 for RP in comparison between the various preconditioning agents CU and RP. The dosage of 10 µg/ml RP was considered statistically significant. Results: The flow cytometer analysis revealed good results for CD 90 compared to other surface markers. The dosage of 10 µg/ml RP was having good chondrogenic cell proliferation. The intragroup analysis showed P value < 0.05 for RP in comparison between the various preconditioning agents CU and RP. The calorimetric assay (MTT) quantitative analysis of the chondrogenic cells with Safranin O stain the standard deviation (SD = 0.017 for rapamycin), Alcian blue (SD = 0.49 for RP) in comparison to DMSO (control) and CU. Conclusion: RP activates mTOR pathway and hence stabilizes the stem cell maintenance of human dental pulp stem cell and the dose quantified can be used for future animal temporomandibular joint animal model.

Ascorbic acid and IFNγ preconditioning enhance the potency of human mesenchymal stem cells to ameliorate LPS induced cytokine storm.

The mesenchymal Stem Cells (MSCs) is one of the leading contender in therapeutic management of cytokine storm implicated in the COVID-19 and other inflammatory conditions. This study was aimed to investigate the effect of Interferon gamma (IFN-γ) and Ascorbic Acid (AA) preconditioning on the secretome of the human Umbilical Cord Derived MSCs (UCMSCs) and their potential to ameliorate the lipopolysaccharide (LPS) induced cytokine storm in the human peripheral blood mononuclear cells (PBMCs). UCMSCs were preconditioned with IFN-γ, AA and secretome (UCMSCs-S, IFNγ-UCMSCs-S and AA-UCSMCs-S) was analysed for the levels of growth factors and cytokines by flow cytometry. The potential of secretome to ameliorate cytokine storm and augment angiogenesis was assessed in the LPS induced PBMCs and yolk sac membrane (YSM) assay respectively. The mRNA transcript and protein levels of IL-6, IL-1ß and TNF-α was analysed by RT-PCR and flow cytometry respectively. IFNγ-UCMSCs-S and AA-UCSMCs-S ameliorated the LPS induced cytokine storm as revealed by the decreased mRNA and protein expression of IL-6, IL-1ß and TNF-α as compared to the UCMSCs-S. IFNγ-UCMSCs-S and AA-UCSMCs-S augmented angiogenesis in YSM assay. Furthermore, IFNγ and AA preconditioning of UCMSCs exhibited distinct growth factors and cytokine profile in the secretome. Our results unequivocally show that IFNγ and AA preconditioning of MSCs could give better therapeutic outcomes in the cell mediated therapies for COVID-19 and other inflammatory conditions.

Therapeutic Potential of Metformin-preconditioned Mesenchymal Stem Cells for Pancreatic Regeneration.

BACKGROUND: Diabetes occurs due to insulin deficiency or less insulin. To manage this condition, insulin administration as well as increased insulin sensitivity is required, but exogeneous insulin cannot replace the sensitive and gentle regulation of blood glucose levels same as ß cells of healthy individuals. By considering the ability of regeneration and differentiation of stem cells, the current study planned to evaluate the effect of metformin preconditioned buccal fat pad (BFP) derived mesenchymal stem cells (MSCs) on streptozotocin (STZ) induced diabetes mellitus in Wistar rats. MATERIALS & METHODS: The disease condition was established by using a diabetes-inducing agent STZ in Wistar rats. Then, the animals were grouped into disease control, blank, and test groups. Only the test group received the metformin-preconditioned cells. The total study period for this experiment was 33 days. During this period, the animals were monitored for blood glucose level, body weight, and food-water intake twice a week. At the end of 33 days, the biochemical estimations for serum insulin level and pancreatic insulin level were performed. Also, histopathology of the pancreas, liver and skeletal muscle was performed. RESULTS: The test groups showed a decline in the blood glucose level and an increase in the serum pancreatic insulin level as compared to the disease group. No significant change in food and water intake was observed within the three groups, while body weight was significantly reduced in the test group when compared with the blank group, but the life span was increased when compared with the disease group. CONCLUSION: In the present study, we concluded that metformin preconditioned buccal fat pad-derived mesenchymal stem cells have the ability to regenerate damaged pancreatic ß cells and have antidiabetic activity, and this therapy is a better choice for future research.

Xanthium strumarium seed extract boosts osteogenesis in human dental pulp stem cell model.

BACKGROUND: In traditional medicine, Xanthium strumarium is used as an anti-inflammatory and anti-arthritic plant-based medicine. Human Dental Pulp Stem Cells (hDPSCs) are an ideal in vitro model for drug and bioactive compound screening. This study assessed the potential of X. strumarium aqueous extract on hDPSCs differentiation towards the osteogenic lineage. MATERIALS AND METHODS: HDPSCs were isolated and cultured by explant method and characterized by surface marker expression, Colony Forming units fibroblasts (CFU-F), Population Doubling time (PDT), and tri-lineage differentiation. X. strumarium aqueous seed extract (XSE) was prepared and its cytotoxic effect on hDPSCs was examined by MTT assay. The effect of XSE on hDPSC differentiation into osteocytes was investigated by biochemical staining and gene expression. RESULTS: The hDPSCs were positive for CD73, CD90, and CD105 and negative for CD34, CD45, and HLA-DR surface markers. The cells had a colony-forming ability with a PDT of 44.91 h. The hDPSCs differentiated into osteocytes, chondrocytes, and adipocytes. The XSE concentration of 15 µg/ml had a significant increase in hDPSC viability. Alizarin Red S staining revealed that XSE treatment enhanced calcium accumulation and matrix mineralization in hDPSCs. XSE treatment also increased osteonectin and IL-6 transcript expression in osteogenesis-induced hDPSCs. CONCLUSION: X. strumarium aqueous extract is a suitable candidate for bone repair because it promotes osteogenic differentiation in hDPSCs. Therefore this could be explored further in the treatment of bone disorders.

CC group of chemokines and associated gene expression of transcription factors: Deciphering immuno-pathogenetic aspect of oral submucous fibrosis.

BACKGROUND: Oral submucous fibrosis (OSMF) is a chronic disease with significantly increasing malignant transformation rate. To date the pathogenesis of OSMF has been considered to be associated with areca nut constituents and their action on fibroblasts. However, fibrosis is also associated with immunological factors such as chemokines. In-depth analysis of such factors is the need of the hour in OSMF to better understand the pathogenesis so that effective therapeutic strategies can be developed in the future. MATERIALS AND METHOD: Clinically diagnosed cases of OSMF (n=21) and healthy individuals (n=10) were enrolled in the present study. Chemokines such as CCL2, CCL3, CCL4, CCL5, CCL11, CCL17, CCL28, CXCL1, CXCL5, CXCL8, CXCL9, CXCL10, and CXCL11 were assessed using the chemokine bead array in conjunction with the flow cytometry, along with real-time PCR (RT-PCR). The transcription factors CREB, NF-κB and NFAT5 were also studied for their expressions. The analysis of pg/ml (picogram/milliliter) values was done by using LEGENDplex™ Data Analysis Software. RESULTS: The results obtained demonstrated early phase transient increase in CXCL-11, CCL20, CXCL9, CCL3, CCL2, CXCL10 and CXCL8. However, the expression of CCL3, CXCL10 and CXCL8 was higher in the late stage as compared to the early stage. The relative gene expression of CREB, NF-κB, NFAT5 were upregulated in the late stage of OSMF when compared to normal. CONCLUSION: Distinctive sets of chemokine expression during the early and late stages of OSMF suggest a unique pattern of disease progression playing an important role in the pathogenesis.

Angiogenic Potential of Various Oral Cavity-Derived Mesenchymal Stem Cells and Cell-Derived Secretome: A Systematic Review and Meta-Analysis.

Recent evidence suggests the immense potential of human mesenchymal stem cell (hMSC) secretome conditioned medium-mediated augmentation of angiogenesis. However, angiogenesis potential varies from source and origin. The hMSCs derived from the oral cavity share an exceptional quality due to their origin from a hypoxic environment. Our systematic review aimed to compare the mesenchymal stem cells (MSCs) derived from various oral cavity sources and cell-derived secretomes, and evaluate their angiogenic potential. A literature search was conducted using PubMed and Scopus from January 2000 to September 2020. Source-wise outcomes were systematically analyzed using in vitro, in vivo, and in ovo studies, emphasizing endothelial cell migration, tube formation, and blood vessel formation. Ninety-four studies were included in the systematic review, out of which 4 studies were subsequently included in the meta-analysis. Prominent growth factors and other bioactive components implicated in improving angiogenesis were included in the respective studies. The findings suggest that oral tissues are a rich source of hMSCs. The meta-analysis revealed a positive correlation between dental pulp-derived MSCs (DPMSCs) and stem cells derived from apical papilla (SCAP) compared to human umbilical cord-derived endothelial cell lines as a control. It shows a statistically significant positive correlation between the co-culture of human umbilical vein endothelial cells (HUVECs) and DPMSCs with tubule length formation and total branching points. Our meta-analysis revealed that oral-derived MSCs (dental pulp stem cells and SCAP) carry a better angiogenic potential in vitro than endothelial cell lines alone. The reviewed literature illustrates that oral cavity-derived MSCs (OC-MSCs) increased angiogenesis. The present literature reveals a dearth of investigations involving sources other than dental pulp. Even though OC-MSCs have revealed more significant potential than other MSCs, more comprehensive, target-oriented interinstitutional prospective studies are warranted to determine whether oral cavity-derived stem cells are the most excellent sources of significant angiogenic potential.

The Marriage between Nanotechnology and Stem Cells Secretome for COPD.

Chronic Obstructive Pulmonary Disease (COPD) is a disorder characterized by narrow alveoli as a result of emphysema. As such, there is no treatment to cure this disorder completely, and existing drugs only delay the progression of the disease. In recent years, the stem cell secretome as a drug is remarkably used as a regenerative therapy. In particular, cell-free therapy approaches offer great opportunities for the treatment of COPD. However, a few issues, such as the delivery of stem cell secretome as a drug to the alveolar region, have obstructed their application in clinical scales. To address these challenges, a combination of stem-cells secretome as a drug with nanotechnology could be a smart solution. We suggest that the combinational approach of delivering nanoparticles loaded with stem cell secretome could be a translational medicine approach for the successful outcome of COPD.

A novel approach to develop an animal model for oral submucous fibrosis.

Epidemiological data have proved the association of consumption of areca nut with the causation of oral submucous fibrosis (OSF). OSF is a chronic inflammatory disease with the potential for malignant transformation from 7 to 13%. The establishment of animal models makes it easier for researchers to focus on the therapeutic options to combat this disease further. We developed and compared two areca nut extract (ANE) administration methods in Swiss albino mice to establish OSF. This study compared an invasive intrabuccal injection technique with a non-invasive intraoral droplet administration. The duration of induction was around 12 weeks. Histopathology (H&E, Masson's trichrome staining) and gene expression analysis (COL-I, COL-II, and α-SMA) were performed using RT-PCR to confirm the OSF in animals. Our study showed that ANE administration through the intraoral droplet method exhibited significantly higher fibrosis than the intrabuccal injections, as evidenced by the H&E and Masson's trichrome staining. Furthermore, intraoral administration of ANE significantly upregulated the mRNA expression of COL-I, COL-II, and α-SMA, as revealed by the RT-PCR analysis. The non-invasive droplet method could simulate the absorption of areca nut seen in humans through daily dosing. This study establishes the intraoral droplet method as an efficient and non-invasive method to administer the ANE to develop OSF. These findings will aid in the efficient development of OSF animal models for interventional studies, including screening novel drugs in the reversal of the OSF.

Synovial Fluid in Knee Osteoarthritis Extends Proinflammatory Niche for Macrophage Polarization.

Macrophage polarization is a steering factor of osteoarthritis (OA) progression. Synovial fluid (SF) obtained from OA patients with different Kellgren-Lawrence grades (KL grades) holds several proinflammatory factors and was hypothesized to induce macrophage differentiation and polarization by providing the needed microenvironment. U937 cells and peripheral-blood-mononuclear-cell-derived monocytes (PBMC-derived CD14+ cells) were induced with SFs of progressive KL grades for 48 h, and the status of the differentiated cells was evaluated by cell surface markers representing M1 and M2 macrophage phenotypes. Functional viability assessment of the differentiated cells was performed by cytokine estimation. The fraction of macrophages and their phenotypes were estimated by immunophenotyping of SF-isolated cells of different KL grades. A grade-wise proteome analysis of SFs was performed in search of the factors which are influential in macrophage differentiation and polarization. In the assay on U937 cells, induction with SF of KL grade III and IV showed a significant increase in M1 type (CD86+). The percentage of M2 phenotype (CD163+) was significantly higher after the induction with SF of KL grade II. A Significantly higher M1/M2 ratio was estimated in the cells induced with KL grade III and IV. The cell differentiation pattern in the assay on PBMC-derived CD14+ cells showed a grade-wise decline in both M1 (CD11C+, CD86+) and M2 phenotype (CD163+). Cytokine estimation specific to M1 (TNF-α, IL-6, IL-1ß, IFN-γ) and M2 (IL-4 and IL-10) macrophages corelated with the differentiation pattern in the U937 cell assay, while it did not reveal any significant changes in the PBMC-derived CD14+ cells assay. SF cells' immunophenotyping showed the highest percentage of CD14+ macrophages in KL grade II; CD86+ and CD163+ cells were minimal in all KL grades' SFs. The proteome analysis revealed significantly expressed MIF, CAPG/MCP, osteopontin, and RAS-related RAB proteins in KL grade III and IV samples, which are linked with macrophages' movement, polarization, and migration-behavior. In conclusion, this study demonstrated that SF in OA joints acts as a niche and facilitates M1 phenotype polarization by providing a proinflammatory microenvironment.

Assessment of angiogenic potential of mesenchymal stem cells derived conditioned medium from various oral sources.

Background: Abnormal angiogenesis hamper blood vessel proliferation implicated in various biological processes. The current method available to clinically treat patients to enhance angiogenesis is administering the angiogenic growth factors. However, due to a lack of spatiotemporal control over the substantial release of these factors, numerous drawbacks are faced such as leaky vasculature. Hence, stem-cell-based therapeutic applications are running their race to evolve as potential targets for deranged angiogenesis. In clinical dentistry, adequate tissue vascularization is essential for successful endodontic therapies such as apexogenesis and apexification. Furthermore, wound healing of the extraction socket and tissue regeneration post-surgical phase of treatment including implant placement require angiogenesis as a foundation for the ultimate success of treatment. Mesenchymal stem cells (MSCs) secrete certain growth factors and cytokines in the culture medium during the proliferation. These factors and cytokines are responsible for various biological activities inside human body. Oral cavity-derived stem cells can secrete growth factors that enhance angiogenesis. Aim: The aim of the study was to investigate the angiogenic potential of conditioned medium (CM) of MSCs derived from different oral sources. Methods: Oral tissues such as dental pulp of adult and deciduous teeth, gingiva, and buccal fat were used to isolate dental pulp MSCs (DPSCs), exfoliated deciduous teeth, gingival MSCs, and buccal fat derived MSCs. MSCs conditioned medium (CM) from passage four cells from all the sources were obtained at 48 h interval and growth factor analysis was performed using flow cytometry. To assess the functionality of the CM, Chick Yolk Sac Membrane (YSM) assay was performed. Results: CM obtained from DPSCs showed higher levels of vascular endothelial growth factor, fibroblast growth factor, and hepatocyte growth factor as evidenced by flow cytometry. Furthermore, DPSC-CM exhibited significantly higher pro-angiogenic potential when assessed in in-ovo YSM assay. Conclusion: DPSCs so far seems to be the best source as compare to the rest of oral sources in promoting angiogenesis. A novel source of CM derived from buccal fat stem cells was used to assess angiogenic potential. Thus, the present study shows that CM derived from oral cavity-derived-MSCs has a dynamic and influential role in angiogenesis. Relevance for Patients: CM derived from various oral sources of MSCs could be used along with existing therapies in medical practice where patients have compromised blood supply like in diabetes and in patients with debilitating disorders. In clinical dentistry, adequate tissue vascularization is essential for successful wound healing, grafting procedures, and endodontic therapies. DPSCs-CM shows better angiogenic potential in comparison with other oral sources of MSCs-CM. Our findings could be a turning point in the management of all surgical and regenerative procedures requiring increased angiogenesis.