Novel insights on oral squamous cell carcinoma management using long non-coding RNAs.

Oral squamous cell carcinoma (OSCC) is one of the most prevalent forms of head and neck squamous cell carcinomas (HNSCC) with a poor overall survival rate (about 50%), particularly in cases of metastasis. RNA-based cancer biomarkers are a relatively advanced concept, and non-coding RNAs currently have shown promising roles in the detection and treatment of various malignancies. This review underlines the function of long non-coding RNAs (lncRNAs) in the OSCC and its subsequent clinical implications. LncRNAs, a class of non-coding RNAs, are larger than 200 nucleotides and resemble mRNA in numerous ways. However, unlike mRNA, lncRNA regulates multiple druggable and non-druggable signaling molecules through simultaneous interaction with DNA, RNA, proteins, or microRNAs depending on concentration and localization in cells. Upregulation of oncogenic lncRNAs and down-regulation of tumor suppressor lncRNAs are evident in OSCC tissues and body fluids such as blood and saliva indicating their potential as valuable biomarkers. Targeted inhibition of candidate oncogenic lncRNAs or over-expression of tumor suppressor lncRNAs showed potential therapeutic roles in in-vivo animal models. The types of lncRNAs that are expressed differentially in OSCC tissue and bodily fluids have been systematically documented with specificity and sensitivity. This review thoroughly discusses the biological functions of such lncRNAs in OSCC cell survival, proliferation, invasion, migration, metastasis, angiogenesis, metabolism, epigenetic modification, tumor immune microenvironment, and drug resistance. Subsequently, we addressed the diagnostic and therapeutic importance of lncRNAs in OSCC pre-clinical and clinical systems, providing details on ongoing research and outlining potential future directions for advancements in this field. In essence, this review could be a valuable resource by offering comprehensive and current insights into lncRNAs in OSCC for researchers in fundamental and clinical domains.

Critical appraisal of the chorioallantoic membrane model for studying angiogenesis in preclinical research.

BACKGROUND: Angiogenesis, the biological mechanism by which new blood vessels are generated from existing ones, plays a vital role in growth and development. Effective preclinical screening is necessary for the development of medications that may enhance or inhibit angiogenesis in the setting of different disorders. Traditional in vitro and, in vivo models of angiogenesis are laborious and time-consuming, necessitating advanced infrastructure for embryo culture. MAIN BODY: A challenge encountered by researchers studying angiogenesis is the lack of appropriate techniques to evaluate the impact of regulators on the angiogenic response. An ideal test should possess reliability, technical simplicity, easy quantifiability, and, most importantly, physiological relevance. The CAM model, leveraging the extraembryonic membrane of the chicken embryo, offers a unique combination of accessibility, low cost, and rapid development, making it an attractive option for angiogenesis assays. This review evaluates the strengths and limitations of the CAM model in the context of its anatomical and physiological properties, and its relevance to human pathophysiological conditions. Its abundant capillary network makes it a common choice for studying angiogenesis. The CAM assay serves as a substitute for animal models and offers a natural setting for developing blood vessels and the many elements involved in the intricate interaction with the host. Despite its advantages, the CAM model's limitations are notable. These include species-specific responses that may not always extrapolate to humans and the ethical considerations of using avian embryos. We discuss methodological adaptations that can mitigate some of these limitations and propose future directions to enhance the translational relevance of this model. This review underscores the CAM model's valuable role in angiogenesis research and aims to guide researchers in optimizing its use for more predictive and robust preclinical studies. CONCLUSION: The highly vascularized chorioallantoic membrane (CAM) of fertilized chicken eggs is a cost-effective and easily available method for screening angiogenesis, in comparison to other animal models.

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.

Andrographolide demonstrates anti-proliferative activity in oral cancer by promoting apoptosis, the programmed cell death process.

Objectives: Andrographolide has been studied on different types of human cancer cells, but very few studies have been conducted on oral cancer. The study aimed to evaluate the anticancer potential of Andrographolide on an oral cancer cell line (KB) through in-silico network analysis and in vitro assays. Materials and Methods: The in-silico analysis involved the determination of drug-likeness prediction, prediction of common targets between oral cancer and andrographolide, Protein-Protein Interactions (PPI), hub genes, top 10 associated pathways by Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway, gene ontology (GO), and molecular docking experiments. In vitro assays comprised MTT assay, apoptosis assay, cell cycle analysis, intracellular reactive oxygen species (ROS) measurement, mitochondrial membrane potential (MMP), anti-migration activity, and gene expressions using polymerase chain reaction (PCR). Results: Fifteen common genes were obtained and were seen to be involved in cellular proliferation, regulation of apoptosis, migration of cells, regulation of MAPK cascade, and regulation of cell cycle. The most common genes involved in the top 10 pathways were MAPK1, MAPK8, MAPK14, and IL6 which were seen to be associated with the MAPK signaling pathway which may be the key pathway through which andrographolide may aid in treating oral cancer. In vitro assays showed anti-proliferative properties, late apoptosis, and anti-migratory properties. Conclusion: According to the results obtained, andrographolide has shown anticancer properties and has the potential to be used as a chemotherapeutic drug. The in-silico approach used in the present study can aid as a model for future research in developing efficient cancer treatments.

Compliance to the Tobacco-Free Educational Institution(ToFEI) guidelines at Madhyamik Vidhyalayas of Pimpri-Chinchwad, Pune, Maharashtra-A cross sectional study.

INTRODUCTION: Revised guidelines for Tobacco-Free Educational Institutes (ToEFI) were laid down in 2019 and they provide for tobacco free environment leading to a healthy life, implementation of legal provisions, and recognition about various approaches available for tobacco cessation. OBJECTIVE: To assess Madhyamik Vidyalays (MVs) for their compliance to the guidelines for ToFEI at the baseline using self-evaluation score card as part of operational research. MATERIAL AND METHOD: A cross sectional study was carried out during March 2021 among 19 MVs of Pimpri-Chinchwad block in Pune District, Maharashtra using census sampling. Trained data collectors scored for all 9 ToFEI criteria including the mandatory one's and their weightage points were calculated. RESULTS AND DISCUSSION: Eight {42%(0.21-0.64)} MVs had displays on tobacco-free area and awareness on the harms of tobacco displayed inside the premises and another three (16%(0.04-0.37)} had only the display of ToFEI signage at their boundary wall. No MV met with 4 or more criteria out of the total 9 criteria. The highest weightage of 29-30 out of 100 was achieved by only 2 {11%(0.01-0.30)} MVs and 5 {26%(0.10-0.49)} MVs achieved 0 points. No significance was given to tobacco free school probably because of untrained teachers and unawareness of the guidelines. CONCLUSION: This study demonstrates that minimal importance has been given to the revised ToEFI guidelines in making MVs tobacco-free. Hence, none of the them could attain the tobacco-free status.

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.

Characterization of stromal calcifications in odontogenic keratocyst: a multicentric study.

Odontogenic keratocysts (OKCs) are locally aggressive cysts that exhibit typical histopathological features and have a propensity for recurrence. Though histological variations are observed in OKCs, hard tissue formation and metaplastic changes are rare, and the underlying pathogenesis is not well understood. This study aimed to characterize stromal calcifications and analyze their association with odontogenic components in non-syndromic and syndrome-associated cases of OKCs. We analyzed 153 cases of OKCs from healthcare institutes in India and Japan. The epithelial and stromal features were evaluated, and the relationship of calcifications with odontogenic rests was determined. Immunohistochemistry for cytokeratin-19 and special stains including Masson Trichrome and Van Gieson, were used for identification of odontogenic rests and calcifications respectively. Stromal calcifications were observed in 29.41% OKCs. The calcification patterns included irregular dystrophic, dentinoid with linear or calcospherite-type mineralization, and psammoma calcifications. Psammoma and dentinoid calcifications were found in the proximity of cytokeratin-19-positive odontogenic rests or satellite cysts, whereas majority cases with dystrophic calcifications did not exhibit co-localization with stromal odontogenic components. Distinct patterns of calcifications were observed in OKCs. Calcifications found in proximity of the odontogenic rests were possibly indicative of an inductive or host-mediated response.

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.

Effectiveness of leukocyte- and platelet-rich fibrin in the management of medication-related osteonecrosis of the jaws ­ a systematic review / Efectividad de la fibrina rica en leucocitos y plaquetas en el tratamiento de osteonecrosis mandibular relacionada con medicamentos: una revisión sistemática

Background: Medication-related osteonecrosis of the jaw (MRONJ) is a rare, but significant adverse event primarily associated with the intake of antiresorptive and antiangiogenic medications. Although antiresorptive and antiangiogenic the-rapies improve life expectancy, particularly in cancer patients, MRONJ may hamper the patient's quality of life due to pain, discomfort, anxiety, depression, speech impairment, difficulty in swallowing and eating, frequent medical and dental evaluations and treatments, and the possibility of treatment discontinuation. Leukocyte­ and Platelet-rich Fibrin (L-PRF) is an autologous platelet aggregate that promotes wound healing by stimulating re-epithelialization, angiogenesis, and extracellular matrix production. Aim: The present systematic review aimed to compare the results in the published literature on whether L-PRF is an effective and predictable adjuvant to surgical debridement of necrotic bone for improving the healing efficacy in patients with MRONJ. Materials and Methods: The PubMed, Scopus, Cochrane, Science Direct, LILACS, and Web of Science databases were searched using the predetermined MeSH terms and eligibility criteria, and the search yielded a total of five articles. Two studies were retrospective, and three studies were case series. Results: Seventeen participants received a combination of surgical debridement, L-PRF membrane, and antibiotics. Complete wound healing was observed in 70% of the participants, and most of them healed without any complications. Conclusions: L-PRF as an adjuvant to surgical debridement of necrosed bone appears to have a positive association with the healing outcome in patients with MRONJ.

Introducción: La osteonecrosis mandibular relacionada con medicamentos (ONMRM) es un evento adverso raro pero significativo asociado principalmente con la ingesta de medicamentos antirresortivos y antiangiogénicos. Aunque las terapias antirresortivas y antiangiogénicas mejoran la esperanza de vida, particularmente en pacientes con cáncer, la ONMRM puede obstaculizar la calidad de vida del paciente debido a dolor, incomodidad, ansiedad, depresión, discapacidad del habla, dificultad para tragar y comer, evaluaciones y tratamientos médicos y dentales frecuentes, y la posibilidad de interrupción del tratamiento. La fibrina rica en plaquetas y leucocitos (L-PRF) es un agregado de plaquetas autólogo que promueve la curación de heridas al estimular la reepitelización, la angiogénesis y la producción de la matriz extracelular. Objetivo: La presente revisión sistemática tuvo como objetivo comparar los resultados en la literatura publicada sobre si L-PRF es un adyuvante efectivo y predecible al desbridamiento quirúrgico del hueso necrótico para mejorar la eficacia curativa en pacientes con ONMRM. Materiales y Métodos: Las bases de datos de PubMed, Scopus, Cochrane, ScienceDirect, LILACS y Web of Science se registraron utilizando los términos DeCS/MeSH predeterminados y los criterios de elegibilidad, y la búsqueda arrojó un total de cinco artículos. Dos estudios fueron retrospectivos, y tres estudios fueron series de casos. Resultado: Diecisiete participantes recibieron una combinación de desbridamiento quirúrgico, membrana L-PRF y antibióticos. Se observó curación completa de heridas en el 70% de los participantes, y la mayoría de ellos se curaron sin ninguna complicación. Conclusión: L-PRF como adyuvante para el desbridamiento quirúrgico del hueso necrótico parece tener una asociación positiva con el resultado de curación en pacientes con ONMRM.

Antifibrotic Effect of Baicalin on Arecoline Induced Human Oral Fibroblast: An In-Vitro Study.

BACKGROUND: Baicalin is a flavonoid obtained from the Chinese herb Scutellaria baicalensis, which has a wide varieties of health benefits and scope to be studied for its therapeutic potential in oral fibrosis. AIM: The aim of the study was to investigate the antifibrotic effect of a Baicalin in arecoline induced human oral fibroblast in vitro setting. MATERIAL AND METHODS: Arecoline and ethanolic extracts of Baicalin were commercially purchased from Sigma-Aldrich. Human oral fibroblasts were cultured and characterized with specific fibroblast markers, and cells were stimulated with arecoline. An MTT assay (3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide) was executed to determine the half-maximal inhibitory concentration of arecoline and Baicalin. Arecoline-induced cells (25µg/ml) were treated with a non-toxic dose of Baicalin (proliferative dose of 25µg/ml). Cytokine (CCL2, CXCL-8, IL17, IL-beta, and IL-6) and fibrotic marker genes were studied by reverse transcription-polymerase chain reaction (RT-PCR). The inhibitory effect of Baicalin was studied to prove its antifibrotic properties. RESULTS: Arecoline significantly upregulated all inflammatory and fibrotic markers. On treatment with 25µg/ml of Baicalin, all inflammatory and fibrotic markers were inhibited. Arecoline affects fibroblast morphology, supporting the fact that arecoline is cytotoxic to cells. CONCLUSION: Baicalin can be used as an antifibrotic herb to treat OSMF.

Feasibility of Immediate Implant Placement in Maxillary First Premolars: Prediction of Implant Locations Using Restorations- A Radiographic Study.

BACKGROUND: Maxillary premolars have a unique anatomical location. This is an CBCT based study where the suitability of maxillary premolars for immediate implant placement (IIP) is evaluated. Based on prosthetically driven treatment treatment planning a simple classification system is put forth. MATERIALS AND METHODS: 150 CBCTs of maxillary first premolars were analysed in BlueskyBio software. The topographic position of the tooth was determined by analysing the dimensions of the buccal and lingual cortical plates, the distance between the bucco-lingual plates and the residual bone height from the root apex to the floor of the sinus. Virtual placement of an implant was carried out such that the implant would be positioned 1 mm apical to the buccal bone crest, would engage 3 mm of bone apical to the root apex, and would have a trajectory so that the abutment access was from the central fossa. Four categories were identified and the classification was proposed. RESULTS: It was observed that 74% of cases had buccal bone<1mm,26% had buccal bone >1mm. 79% cases had an average distance >3mm between root apex and maxillary sinus, 21% had an average distance of root apex and maxillary sinus <3mm. The categorizations of implant placement were as follows -Type 1- 24%, Type 2- 56.6%, Type 3-43.3%, Type 4- 0%. CONCLUSIONS: In majority of maxillary 1st premolars an IIP is possible with the implants to be placed in the palatal sockets or the furcation area. In cases were the buccal plate thickness is inadequate, simultaneous grafting should be considered between the implant position and buccal plate.

Linking inflammation and angiogenesis with fibrogenesis: Expression of FXIIIA, MMP-9, and VEGF in oral submucous fibrosis.

OBJECTIVES: Interplay of Factor XIIIa (FXIIIa), a transglutaminase, responsible for cross-linking of matrix proteins, Matrix Metalloproteinase-9 (MMP-9), a gelatinase, and Vascular Endothelial Growth Factor (VEGF), an angiogenic inducer, were studied in relation to fibrogenesis and disease progression in oral submucous fibrosis (OSMF). MATERIAL AND METHODS: Immunohistochemical expression of markers was studied in 60 formalin-fixed paraffin-embedded tissue blocks of OSMF and 20 normal oral mucosal tissues. FXIIIa was studied quantitatively while MMP-9 and VEGF were assessed semi-quantitatively. Expression was compared with histopathological grades of OSMF. RESULTS: FXIIIa expression significantly increased in OSMF (p-value 0.000). However, expression decreased and cells became quiescent with increasing grades (p-value 0.000). MMP-9 (p-value epithelium 0.011, p-value connective tissue 0.000) and VEGF expression (p-value epithelium 0.000, connective tissue 0.000) increased in OSMF. A negative correlation between FXIIIa and MMP-9 (-0.653) in early grade (p-value of 0.021) and a positive correlation between FXIIIa and VEGF (0.595) (p-value of 0.032) was found in the moderate grade OSMF. Regression analysis showed a significant association (p<0.01) of FXIIIa in OSMF and with increasing grades of OSMF. CONCLUSION: FXIIIa may play a crucial role in initiation of fibrosis in OSMF. MMP-9 may have a diverse role to play in OSMF as a regulator of fibrosis. VEGF may show an angio-fibrotic switch and contribute to fibrosis in OSMF. These cytokines may show altered function and can contribute to fibrosis and chronicity of disease due to changes in the microenvironment. Tissue stiffness in OSMF itself creates an environment that enhances the chronicity of the disease.

Oral Microbial and Molecular Cross Talk between SARS-CoV-2 and Diabetes Mellitus - A Mini Review.

The oral microbiome has long been considered a measure of overall systemic health. It is often significantly altered in case of chronic inflammation or any other systemic infection. Therefore, a shift in oral microbiota and oral health is bound to be observed in diabetics infected with the coronavirus. The prognosis of COVID-19 in a diabetic individual is often worse than that in a healthy individual. The increased pathogenicity of coronavirus in diabetics is due to the peculiar ways in which it interacts with specific physiological mechanisms in a diabetic patient and vice versa. Diabetes Mellitus Type-II (DM -II) is one of the most frequently associated co-morbidities in a COVID-19 patient, and therefore it is even more pertinent that their interrelationship is understood. It is essential to recognize the above-mentioned interactions and consider their implications while treating susceptible patients. This article attempts to review and summarize the said vital interactions. Additionally, it attempts to guide and prepare oral health professionals on what to expect and how to treat diabetic patients in a future where coronavirus is, as unfortunate as it is, a regularity and not a rarity.

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.

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.

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 'One Customized Abutment One Time' concept - An innovative technique for optimizing immediacy and digital protocols.

Disconnection and reconnection of abutments multiple times have known to affect the mucosal barrier around implants leading to marginal bone loss. This clinical report describes a novel technique that amalgamates the benefits of digital technologies encompassing the fabrication of surgical guides for implant placement, customized hybrid zirconia abutments and all ceramic lithium disilicate crowns prior to implant placement. A correct 3-dimensional implant positioning along with immediate placement of the definitive hybrid customized abutment and a lithium disilicate crown has the potential to reduce treatment time, visits and costs while delivering optimal esthetic outcomes.

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.

Comparative Evaluation of Marginal Bone Levels, ISQ Trends, and Implant Survival Rates Between Conventional Drilling and Osteotome Technique Using Implants of Varied Lengths: A Split-Mouth Randomized Controlled Clinical Trial.

PURPOSE: To assess marginal bone loss (MBL) and implant stability when implant site preparation is performed with conventional drilling and the osteotome technique in the posterior maxilla. MATERIALS AND METHODS: In total, 30 patients (mean age: 46.97 + 7.48 years) receiving 60 implants were enrolled in this study. In each patient, implant site preparation was done using either conventional drilling (conventional group; n = 30) or the osteotome technique (osteotome group; n = 30). The implant sites were further divided into groups based on the implant length used (implant length < 10 mm, implant length ≥ 10 mm). Marginal bone levels and implant stability quotient (ISQ) values were evaluated at the time of crown insertion and 1 year later. Independent t test and paired t test were used for intergroup and intragroup comparison, respectively. RESULTS: The osteotome group showed statistically significant higher initial ISQ (ISQi) and final ISQ (ISQf) values (ISQi: 61 ± 3.6; ISQf: 64.08 ± 3.7) compared to the conventional group (ISQi: 58.01 ± 4.6; ISQf: 61.32 ± 4.8). Statistically significant higher mean MBL was noted in the conventional group (-0.33 ± 0.12 mm) compared to the osteotome group (-0.26 ± 0.10 mm). Higher MBL was noted in the osteotome group (-0.32 ± 0.09 mm) compared to the conventional group (-0.30 ± 0.14 mm) for implants shorter than 10 mm. For implants ≥ 10 mm in length, significantly higher MBL was noted in the conventional group (-0.37 ± 0.09 mm) compared to the osteotome group (-0.19 ± .06 mm). CONCLUSIONS: Osteotome technique could be used as an alternative to conventional drilling, especially when implants longer than 10 mm are planned in the posterior maxilla.

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.