Hydroxamate-Based Histone Deacetylase Inhibitors as Potential Mediators to Induce Dentine Regeneration by Human Dental Pulp Cell

Human dental pulp cells (hDPCs) have shown their plasticity when treated with the hydroxamate-based histone deacetylase (HDAC) inhibitor members, Trichostatin A (TSA), and suberoylanilide hydroxamic acid (SAHA). However, a comparison of their potency to stimulate odontoblast-like differentiation and mineralization has not been reported. The aim of our study was to confirm and compare these TSA and SAHA effects. Primary hDPCs cultured with/without various TSA or SAHA concentrations were evaluated using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), ALP activity, alizarin red staining, and scratch wound healing assays. The inhibitory effect of TSA and SAHA on inhibiting the activity of HDAC was evaluated by HDAC activity assay. Odontoblast-related gene expression was determined using RT-qPCR. The MTT assay indicated that TSA or SAHA did not affect hDPC viability. TSA or SAHA treatment-induced odontoblast-like differentiation as evidenced by a significant increase in alkaline phosphatase activity and mineral deposition after 400 nM TSA or 1 μM SAHA treatment. A significant increase in nuclear factor I C, kruppel like factor 4, dentin matrix acidic phosphoprotein 1, dentin sialophosphoprotein, collagen type I alpha 1 chain, alkaline phosphatase (ALPL), integrin-binding sialoprotein, bone gamma-carboxyglutamate protein, vascular endothelial growth factor A, and cyclin-dependent kinase inhibitor 1A gene expression analyzed by RT-qPCR, at 24, 72 h, 7, and 10 days of treatment. The activity of HDAC in hDPCs culture was significantly inhibited after 72 h TSA and SAHA treatment. The scratch wound healing assay displayed enhanced cell migration at 72 h after TSA or SAHA treatment. Our findings demonstrated that TSA and SAHA have similar stimulatory effects in inducing HDPC odontogenic differentiation and mineralization and propose another potential use of TSA and SAHA to promote dentin regeneration.

Download Full-text

The Selective Histone Deacetylase Inhibitor MI192 Enhances the Osteogenic Differentiation Efficacy of Human Dental Pulp Stromal Cells

International Journal of Molecular Sciences ◽

10.3390/ijms22105224 ◽

2021 ◽

Vol 22 (10) ◽

pp. 5224

Author(s):

Kenny Man ◽

Liam Lawlor ◽

Lin-Hua Jiang ◽

Xuebin B. Yang

Keyword(s):

Alkaline Phosphatase ◽

Osteogenic Differentiation ◽

Stromal Cells ◽

Dental Pulp ◽

Selective Inhibitor ◽

Epigenetic Reprogramming ◽

Type I ◽

Human Dental Pulp ◽

Pre Treatment ◽

Dose Dependent

The use of human dental pulp stromal cells (hDPSCs) has gained increasing attention as an alternative stem cell source for bone tissue engineering. The modification of the cells’ epigenetics has been found to play an important role in regulating differentiation, with the inhibition of histone deacetylases 3 (HDAC3) being linked to increased osteogenic differentiation. This study aimed to induce epigenetic reprogramming using the HDAC2 and 3 selective inhibitor, MI192 to promote hDPSCs osteogenic capacity for bone regeneration. MI192 treatment caused a time–dose-dependent change in hDPSC morphology and reduction in viability. Additionally, MI192 successfully augmented hDPSC epigenetic functionality, which resulted in increased histone acetylation and cell cycle arrest at the G2/M phase. MI192 pre-treatment exhibited a dose-dependent effect on hDPSCs alkaline phosphatase activity. Quantitative PCR and In-Cell Western further demonstrated that MI192 pre-treatment significantly upregulated hDPSCs osteoblast-related gene and protein expression (alkaline phosphatase, bone morphogenic protein 2, type I collagen and osteocalcin) during osteogenic differentiation. Importantly, MI192 pre-treatment significantly increased hDPSCs extracellular matrix collagen production and mineralisation. As such, for the first time, our findings show that epigenetic reprogramming with the HDAC2 and 3 selective inhibitor MI192 accelerates the osteogenic differentiation of hDPSCs, demonstrating the considerable utility of this MSCs engineering approach for bone augmentation strategies.

Download Full-text

The calcium dynamics of human dental pulp stem cells stimulated with tricalcium silicate-based cements determine their differentiation and mineralization outcome

Scientific Reports ◽

10.1038/s41598-020-80096-5 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Elanagai Rathinam ◽

Srinath Govindarajan ◽

Sivaprakash Rajasekharan ◽

Heidi Declercq ◽

Dirk Elewaut ◽

...

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Dental Pulp ◽

Maximum Amplitude ◽

Tricalcium Silicate ◽

Dental Pulp Stem Cells ◽

Alizarin Red ◽

Mineralization Potential ◽

Intracellular Ca ◽

Human Dental Pulp

AbstractCalcium (Ca2+) signalling plays an indispensable role in dental pulp and dentin regeneration, but the Ca2+ responses of human dental pulp stem cells (hDPSCs) stimulated with tricalcium silicate-based (TCS-based) dental biomaterials remains largely unexplored. The objective of the present study was to identify and correlate extracellular Ca2+ concentration, intracellular Ca2+ dynamics, pH, cytotoxicity, gene expression and mineralization ability of human dental pulp stem cells (hDPSCs) stimulated with two different TCS-based biomaterials: Biodentine and ProRoot white MTA. The hDPSCs were exposed to the biomaterials, brought in contact with the overlaying medium, with subsequent measurements of extracellular Ca2+ and pH, and intracellular Ca2+ changes. Messenger RNA expression (BGLAP, TGF-β, MMP1 and BMP2), cytotoxicity (MTT and TUNEL) and mineralization potential (Alizarin red and Von Kossa staining) were then evaluated. Biodentine released significantly more Ca2+ in the α-MEM medium than ProRoot WMTA but this had no cytotoxic impact on hDPSCs. The larger Biodentine-linked Ca2+ release resulted in altered intracellular Ca2+ dynamics, which attained a higher maximum amplitude, faster rise time and increased area under the curve of the Ca2+ changes compared to ProRoot WMTA. Experiments with intracellular Ca2+ chelation, demonstrated that the biomaterial-triggered Ca2+ dynamics affected stem cell-related gene expression, cellular differentiation and mineralization potential. In conclusion, biomaterial-specific Ca2+ dynamics in hDPSCs determine differentiation and mineralization outcomes, with increased Ca2+ dynamics enhancing mineralization.

Download Full-text

The Role of Nephronectin on Proliferation and Differentiation in Human Dental Pulp Stem Cells

Stem Cells International ◽

10.1155/2017/2546261 ◽

2017 ◽

Vol 2017 ◽

pp. 1-14

Author(s):

Jia Tang ◽

Takashi Saito

Keyword(s):

Stem Cells ◽

Mrna Expression ◽

Dental Pulp ◽

Expression Profiles ◽

Dental Pulp Stem Cells ◽

Type I ◽

Fish Scale ◽

Alizarin Red ◽

Matrix Mineralization ◽

Human Dental Pulp

Aim. The purpose of the current study was to investigate the effects of nephronectin (Npnt) in human dental pulp stem cells (hDPSCs). Methodology. Npnt was coated to nontissue culture-treated polystyrene (non-PS) plates. The presence of immobilized protein on the surface was detected by polyclonal rabbit primary anti-Npnt antibody. Then the cell number was counted and compared with PBS-, bovine serum albumin- (BSA-), fish scale type I collagen- (FCOL1-), and human fibronectin- (Fn-) coated wells. Cell proliferation was assessed using CCK-8 assay. Cell morphology was observed under light microscopy and fluorescence microscopy. Lastly, the mRNA expression profiles of integrins, dentin sialophosphoprotein (DSPP), bone sialoprotein (BSP), and mineralization capacity of hDPSCs were investigated by real time RT-PCR and alizarin red staining, respectively. Results. Npnt mediates hDPSC adhesion and spreading partially via the Arg-Gly-Asp (RGD) motif. Npnt enhanced the mRNA expression of ITGA1, ITGA4, ITGA7, and ITGB1 on day five. Npnt downregulated DSPP but significantly upregulated BSP mRNA expression at day 28. Further, Npnt and FCOL1 accelerated the matrix mineralization in hDPSCs. Conclusions. The current findings implicate that Npnt would be favorable to recruit hDPSCs and conducive to mineralization in hDPSCs. The combination of Npnt with hDPSCs may offer a promising approach for hard tissue regeneration.

Download Full-text

Trichostatin A, a histone deacetylase inhibitor, modulates unloaded-induced skeletal muscle atrophy

Journal of Applied Physiology ◽

10.1152/japplphysiol.01031.2014 ◽

2015 ◽

Vol 119 (4) ◽

pp. 342-351 ◽

Cited By ~ 16

Author(s):

Sylvie Dupré-Aucouturier ◽

Josiane Castells ◽

Damien Freyssenet ◽

Dominique Desplanches

Keyword(s):

Gene Expression ◽

Skeletal Muscle ◽

Histone Deacetylase ◽

Muscle Atrophy ◽

Soleus Muscle ◽

Histone Deacetylase Inhibitor ◽

Trichostatin A ◽

Skeletal Muscle Atrophy ◽

Type I ◽

Deacetylase Inhibitor

Skeletal muscle atrophy is commonly associated with immobilization, ageing, and catabolic diseases such as diabetes and cancer cachexia. Epigenetic regulation of gene expression resulting from chromatin remodeling through histone acetylation has been implicated in muscle disuse. The present work was designed to test the hypothesis that treatment with trichostatin A (TSA), a histone deacetylase inhibitor, would partly counteract unloading-induced muscle atrophy. Soleus muscle atrophy (−38%) induced by 14 days of rat hindlimb suspension was reduced to only 25% under TSA treatment. TSA partly prevented the loss of type I and IIa fiber size and reversed the transitions of slow-twitch to fast-twitch fibers in soleus muscle. Unloading or TSA treatment did not affect myostatin gene expression and follistatin protein. Soleus protein carbonyl content remained unchanged, whereas the decrease in glutathione vs. glutathione disulfide ratio and the increase in catalase activity (biomarkers of oxidative stress) observed after unloading were abolished by TSA treatment. The autophagy-lysosome pathway (Bnip3 and microtubule-associated protein 1 light chain 3 proteins, Atg5, Gabarapl1, Ulk1, and cathepsin B and L mRNA) was not activated by unloading or TSA treatment. However, TSA suppressed the rise in muscle-specific RING finger protein 1 (MuRF1) caused by unloading without affecting the forkhead box (Foxo3) transcription factor. Prevention of muscle atrophy by TSA might be due to the regulation of the skeletal muscle atrophy-related MuRF1 gene. Our findings suggest that TSA may provide a novel avenue to treat unloaded-induced muscle atrophy.

Download Full-text

Histone Deacetylase Inhibition with Valproic Acid Downregulates Osteocalcin Gene Expression in Human Dental Pulp Stem Cells and Osteoblasts: Evidence for HDAC2 Involvement

Stem Cells ◽

10.1002/stem.1544 ◽

2014 ◽

Vol 32 (1) ◽

pp. 279-289 ◽

Cited By ~ 73

Author(s):

Francesca Paino ◽

Marcella La Noce ◽

Virginia Tirino ◽

Pasqualina Naddeo ◽

Vincenzo Desiderio ◽

...

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Valproic Acid ◽

Histone Deacetylase ◽

Dental Pulp ◽

Dental Pulp Stem Cells ◽

Histone Deacetylase Inhibition ◽

Osteocalcin Gene ◽

Human Dental Pulp

Download Full-text

A Prototype Skin Substitute, Made of Recycled Marine Collagen, Improves the Skin Regeneration of Sheep

Animals ◽

10.3390/ani11051219 ◽

2021 ◽

Vol 11 (5) ◽

pp. 1219

Author(s):

Luca Melotti ◽

Tiziana Martinello ◽

Anna Perazzi ◽

Ilaria Iacopetti ◽

Cinzia Ferrario ◽

...

Keyword(s):

Gene Expression ◽

Wound Healing ◽

Sea Urchin ◽

Healing Process ◽

Skin Wound ◽

Skin Substitute ◽

Large Animal ◽

Type I ◽

Skin Wound Healing ◽

Skin Adnexa

Skin wound healing is a complex and dynamic process that aims to restore lesioned tissues. Collagen-based skin substitutes are a promising treatment to promote wound healing by mimicking the native skin structure. Recently, collagen from marine organisms has gained interest as a source for producing biomaterials for skin regenerative strategies. This preliminary study aimed to describe the application of a collagen-based skin-like scaffold (CBSS), manufactured with collagen extracted from sea urchin food waste, to treat experimental skin wounds in a large animal. The wound-healing process was assessed over different time points by the means of clinical, histopathological, and molecular analysis. The CBSS treatment improved wound re-epithelialization along with cell proliferation, gene expression of growth factors (VEGF-A), and development of skin adnexa throughout the healing process. Furthermore, it regulated the gene expression of collagen type I and III, thus enhancing the maturation of the granulation tissue into a mature dermis without any signs of scarring as observed in untreated wounds. The observed results (reduced inflammation, better re-epithelialization, proper development of mature dermis and skin adnexa) suggest that sea urchin-derived CBSS is a promising biomaterial for skin wound healing in a “blue biotechnologies” perspective for animals of Veterinary interest.

Download Full-text

Effects of different aperture-sized type I collagen/silk fibroin scaffolds on the proliferation and differentiation of human dental pulp cells

Regenerative Biomaterials ◽

10.1093/rb/rbab028 ◽

2021 ◽

Vol 8 (4) ◽

Author(s):

Shihui Jiang ◽

Zhaoxia Yu ◽

Lanrui Zhang ◽

Guanhua Wang ◽

Xiaohua Dai ◽

...

Keyword(s):

Silk Fibroin ◽

Dental Pulp ◽

Type I Collagen ◽

Type I ◽

Dental Pulp Cells ◽

Human Dental Pulp Cells ◽

Alp Activity ◽

Proliferation And Differentiation ◽

Human Dental Pulp ◽

Pulp Cells

Abstract This study aimed at evaluate the effects of different aperture-sized type I collagen/silk fibroin (CSF) scaffolds on the proliferation and differentiation of human dental pulp cells (HDPCs). The CSF scaffolds were designed with 3D mapping software Solidworks. Three different aperture-sized scaffolds (CSF1–CSF3) were prepared by low-temperature deposition 3D printing technology. The morphology was observed by scanning electron microscope (SEM) and optical coherence tomography. The porosity, hydrophilicity and mechanical capacity of the scaffold were detected, respectively. HDPCs (third passage, 1 × 105 cells) were seeded into each scaffold and investigated by SEM, CCK-8, alkaline phosphatase (ALP) activity and HE staining. The CSF scaffolds had porous structures with macropores and micropores. The macropore size of CSF1 to CSF3 was 421 ± 27 μm, 579 ± 36 μm and 707 ± 43 μm, respectively. The porosity was 69.8 ± 2.2%, 80.1 ± 2.8% and 86.5 ± 3.3%, respectively. All these scaffolds enhanced the adhesion and proliferation of HDPCs. The ALP activity in the CSF1 group was higher than that in the CSF3 groups (P < 0.01). HE staining showed HDPCs grew in multilayer within the scaffolds. CSF scaffolds significantly improved the adhesion and ALP activity of HDPCs. CSF scaffolds were promising candidates in dentine-pulp complex regeneration.

Download Full-text

Evaluation of Resin-Based Material Containing Copaiba Oleoresin (Copaifera Reticulata Ducke): Biological Effects on the Human Dental Pulp Stem Cells

Biomolecules ◽

10.3390/biom10070972 ◽

2020 ◽

Vol 10 (7) ◽

pp. 972

Author(s):

Roberta Souza D’Almeida Couto ◽

Maria Fernanda Setubal Destro Rodrigues ◽

Leila Soares Ferreira ◽

Ivana Márcia Alves Diniz ◽

Fernando de Sá Silva ◽

...

Keyword(s):

Stem Cells ◽

Dental Pulp ◽

Biological Effects ◽

Dental Pulp Stem Cells ◽

Nodule Formation ◽

Alizarin Red ◽

Pulp Capping ◽

Hsp 27 ◽

Dental Pulp Capping ◽

Human Dental Pulp

The purpose of this study was to analyze in vitro the biological effects on human dental pulp stem cells triggered in response to substances leached or dissolved from two experimental cements for dental pulp capping. The experimental materials, based on extracts from Copaifera reticulata Ducke (COP), were compared to calcium hydroxide [Ca(OH)2] and mineral trioxide aggregate (MTA), materials commonly used for direct dental pulp capping in restorative dentistry. For this, human dental pulp stem cells were exposed to COP associated or not with Ca(OH)2 or MTA. Cell cytocompatibility, migration, and differentiation (mineralized nodule formation (Alizarin red assay) and gene expression (RT-qPCR) of OCN, DSPP, and HSP-27 (genes regulated in biomineralization events)) were evaluated. The results showed that the association of COP reduced the cytotoxicity of Ca(OH)2. Upregulations of the OCN, DSPP, and HSP-27 genes were observed in response to the association of COP to MTA, and the DSPP and HSP-27 genes were upregulated in the Ca(OH)2 + COP group. In up to 24 h, cell migration was significantly enhanced in the MTA + COP and Ca(OH)2 + COP groups. In conclusion, the combination of COP with the currently used materials for dental pulp capping [Ca(OH)2 and MTA] improved the cell activities related to pulp repair (i.e., cytocompatibility, differentiation, mineralization, and migration) including a protective effect against the cytotoxicity of Ca(OH)2.

Download Full-text