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.

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.

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.

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.

Linking inflammation and angiogenesis with fibrogenesis: Expression of FXIIIA, MMP-9, and VEGF in oral submucous fibrosis / Vinculación de la inflamación y la angiogénesis con la fibrogénesis: expresión de FXIIIA, MMP-9 y VEGF en la fibrosis submucosa oral

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. (AU)

Objetivos Se estudió la interacción del factor XIIIa (FXIIIa), una transglutaminasa responsable de los entrecruzamientos de las proteínas de la matriz, la metaloproteinasa de matriz-9 (MMP-9), una gelatinasa y el factor de crecimiento endotelial vascular (VEGF), un inductor angiogénico, en relación con la fibrogénesis y la progresión de la enfermedad en la fibrosis submucosa oral (OSMF). Material y métodos Se estudió la expresión inmunohistoquímica de marcadores en 60 bloques de tejido de OSMF fijados con formalina e incluidos en parafina y 20 tejidos de mucosa oral normales. FXIIIa se estudió cuantitativamente mientras que MMP-9 y VEGF se evaluaron semicuantitativamente. La expresión se comparó con los grados histopatológicos de OSMF. Resultados La expresión de FXIIIa aumentó significativamente en OSMF (valor de p 0,000). Sin embargo, la expresión disminuyó y las células se volvieron inactivas a medida que aumentaban los grados (valor de p 0,000). MMP-9 (valor de p epitelio 0,011, tejido conectivo valor de p 0,000) y expresión de VEGF (valor de p epitelio 0,000, tejido conectivo 0,000) aumentaron en OSMF. Se encontró una correlación negativa entre FXIIIa y MMP-9 (-0,653) en grado temprano (valor de p de 0,021) y una correlación positiva entre FXIIIa y VEGF (0,595) (valor de p de 0,032) en OSMF de grado moderado. El análisis de regresión mostró una asociación significativa (p<0,01) de FXIIIa en OSMF y con grados crecientes de OSMF. Conclusión FXIIIa puede desempeñar un papel crucial en el inicio de la fibrosis en OSMF. MMP-9 puede desempeñar un papel diverso en OSMF como regulador de la fibrosis. VEGF puede mostrar un interruptor angiofibrótico y contribuir a la fibrosis en OSMF. Estas citocinas pueden mostrar una función alterada y pueden contribuir a la fibrosis y la cronicidad de la enfermedad debido a cambios en el microambiente. La rigidez del tejido en el propio OSMF crea un entorno que mejora la cronicidad de la enfermedad. (AU)

Linking inflammation and angiogenesis with fibrogenesis: Expression of FXIIIA, MMP-9, and VEGF in oral submucous fibrosis / Vinculación de la inflamación y la angiogénesis con la fibrogénesis: expresión de FXIIIA, MMP-9 y VEGF en la fibrosis submucosa oral

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. (AU)

Objetivos Se estudió la interacción del factor XIIIa (FXIIIa), una transglutaminasa responsable de los entrecruzamientos de las proteínas de la matriz, la metaloproteinasa de matriz-9 (MMP-9), una gelatinasa y el factor de crecimiento endotelial vascular (VEGF), un inductor angiogénico, en relación con la fibrogénesis y la progresión de la enfermedad en la fibrosis submucosa oral (OSMF). Material y métodos Se estudió la expresión inmunohistoquímica de marcadores en 60 bloques de tejido de OSMF fijados con formalina e incluidos en parafina y 20 tejidos de mucosa oral normales. FXIIIa se estudió cuantitativamente mientras que MMP-9 y VEGF se evaluaron semicuantitativamente. La expresión se comparó con los grados histopatológicos de OSMF. Resultados La expresión de FXIIIa aumentó significativamente en OSMF (valor de p 0,000). Sin embargo, la expresión disminuyó y las células se volvieron inactivas a medida que aumentaban los grados (valor de p 0,000). MMP-9 (valor de p epitelio 0,011, tejido conectivo valor de p 0,000) y expresión de VEGF (valor de p epitelio 0,000, tejido conectivo 0,000) aumentaron en OSMF. Se encontró una correlación negativa entre FXIIIa y MMP-9 (-0,653) en grado temprano (valor de p de 0,021) y una correlación positiva entre FXIIIa y VEGF (0,595) (valor de p de 0,032) en OSMF de grado moderado. El análisis de regresión mostró una asociación significativa (p<0,01) de FXIIIa en OSMF y con grados crecientes de OSMF. Conclusión FXIIIa puede desempeñar un papel crucial en el inicio de la fibrosis en OSMF. MMP-9 puede desempeñar un papel diverso en OSMF como regulador de la fibrosis. VEGF puede mostrar un interruptor angiofibrótico y contribuir a la fibrosis en OSMF. Estas citocinas pueden mostrar una función alterada y pueden contribuir a la fibrosis y la cronicidad de la enfermedad debido a cambios en el microambiente. La rigidez del tejido en el propio OSMF crea un entorno que mejora la cronicidad de la enfermedad. (AU)

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.

Evaluation of tumor-infiltrating T & B lymphocytes and their association and distribution in oral squamous cell carcinoma tumor microenvironment: An in vitro immunohistochemical study.

OBJECTIVE: The immune interaction between host immunity and the tumor microenvironment is complex, and a thorough understanding of tumor-infiltrating lymphocyte selection in oral cancer, including T and B cells, is urgently required. Within the tumor microenvironment, tumor cells escape immune surveillance and grow uncontrollably. The study examined the relationship and distribution of tumor-infiltrating T and B lymphocytes. STUDY DESIGN: Retrospective data of paraffin-embedded tissue samples of 47 primary oral squamous cell carcinoma (OSCC) cases were retrieved. Hematoxylin and eosin evaluation, along with all clinicopathologic data, were collected. Immunohistochemical CD3 and CD20 markers were used and evaluated for association and distribution in given OSCC cases. RESULTS: The intermediate type of inflammatory infiltrate was seen primarily in Well DIfferentiated Squamous cell Carcinoma grade and positive and negative lymph nodes. Compared with T-cell density, B-cell density showed an aggregate pattern rather than a scattered pattern, indicating a statistically significant association between T-cell and B-cell infiltrate. B-cell infiltrates were also found to have a statistically significant relationship with tertiary lymphoid structure. CONCLUSIONS: A strong, positive association and correlation exists between B- and T-lymphocyte infiltration in both the stroma and the invasive front. When compared with T-cell density, B-cell density is more predominantly in aggregates.

Evaluation of shear bond strength and translucency of zirconia-reinforced lithium silicate and lithium disilicate: An in vitro study.

Aim: To analyze the effect of various surface treatment protocols on shear bond strength between the ceramic and resin cement (RC) and influence of zirconia on the translucency of LD as compared to zirconia-reinforced lithium silicate (ZLS). Setting and Design: In-Vitro Study. Materials and Methods: Specimens (14 mm × 12 mm × 2 mm) (n = 135) and (14 mm × 12 mm × 1 mm) (n = 45) of ZLS computer-aided design/computer-aided manufacturing glass ceramic block and LD were fabricated, respectively. All the ZLS specimens were crystallized and were tested for the translucency parameter and ceramic-resin shear bond strength. Two different types of surface treatment were used on the ZLS and LD samples. The specimens were treated using the hydrofluoric acid (HF) etching or air abrasion with diamond particles (DPs). The specimens were then bonded using self-adhesive RC to a composite disc of 10 mm and thermocycling was performed. A universal testing machine was used to evaluate ceramic-resin shear bond strength after 24 h. The translucency of the specimens was evaluated using the spectrophotometer by calculating the difference in color between the readings over a black background and a white background. Statistical Analysis Used: Data were statistically analyzed using the independent sample t-test and analysis of variance with Bonferroni's correction and comparison was made between the specimens. Results: Independent sample t-test demonstrated statistically significantly higher translucency for group ZLS (61.44 ± 22) as compared to group LD (20.16 ± 8.39) (P < 0.001). Group ZLS showed statistically significant higher shear bond strength when surface treatment using HF or air abrasion with synthetic DPs was performed as compared to untreated group (3.58 ± 0.45) (P < 0.001). Moreover, air abrasion group (16.79 ± 2.11 megapascal [MPa]) demonstrated statistically significant higher shear bond strength as compared to HF etched group (8.25 ± 0.30 MPa) (P < 0.001). Furthermore, statistically significant higher shear bond strength was noted when air abrasion was done for group ZLS (16.79 ± 2.11 MPa) as compared to group LD (10.82 ± 1.92 MPa) (P < 0.001). However, on surface treatment with HF, a statistically significantly lower shear bond strength was noted for group ZLS (8.25 ± 0.30 MPa) as compared to group LD (11.29 ± 0.58 MPa) (P = 0.001). Conclusion: ZLS demonstrated higher translucency compared to LD restorations. DP abrasion of ZLS is recommended to achieve higher shear bond strength between the ceramic and RC.

In-Vitro Evaluation of Photofunctionalized Implant Surfaces in a High-Glucose Microenvironment Simulating Diabetics.

The present study aimed to assess the efficacy of photofunctionalization on commercially available dental implant surfaces in a high-glucose environment. Discs of three commercially available implant surfaces were selected with various nano- and microstructural alterations (Group 1-laser-etched implant surface, Group 2-titanium-zirconium alloy surface, Group 3-air-abraded, large grit, acid-etched surface). They were subjected to photo-functionalization through UV irradiation for 60 and 90 min. X-ray photoelectron spectroscopy (XPS) was used to analyze the implant surface chemical composition before and after photo-functionalization. The growth and bioactivity of MG63 osteoblasts in the presence of photofunctionalized discs was assessed in cell culture medium containing elevated glucose concentration. The normal osteoblast morphology and spreading behavior were assessed under fluorescence and phase-contrast microscope. MTT (3-(4,5 Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and alizarin red assay were performed to assess the osteoblastic cell viability and mineralization efficiency. Following photofunctionalization, all three implant groups exhibited a reduced carbon content, conversion of Ti4+ to Ti3+, increased osteoblastic adhesion, viability, and increased mineralization. The best osteoblastic adhesion in the medium with increased glucose was seen in Group 3. Photofunctionalization altered the implant surface chemistry by reducing the surface carbon content, probably rendering the surfaces more hydrophilic and conducive for osteoblastic adherence and subsequent mineralization in high-glucose environment.