Biological Interaction Between Human Gingival Fibroblasts and Vascular Endothelial Cells for Angiogenesis: A Co-culture Perspective

Advancement in cell culture protocols, multidisciplinary research approach, and the need of clinical implication to reconstruct damaged or diseased tissues has led to the establishment of three-dimensional (3D) test systems for regeneration and repair. Regenerative therapies, including dental tissue engineering, have been pursued as a new prospect to repair and rebuild the diseased/lost oral tissues. Interactions between the different cell types, growth factors, and extracellular matrix components involved in angiogenesis are vital in the mechanisms of new vessel formation for tissue regeneration. In vitro pre-vascularization is one of the leading scopes in the tissue-engineering field. Vascularization strategies that are associated with co-culture systems have proved that there is communication between different cell types with mutual beneficial effects in vascularization and tissue regeneration in two-dimensional or 3D cultures. Endothelial cells with different cell populations, including osteoblasts, smooth muscle cells, and fibroblasts in a co-culture have shown their ability to advocate pre-vascularization. In this review, a co-culture perspective of human gingival fibroblasts and vascular endothelial cells is discussed with the main focus on vascularization and future perspective of this model in regeneration and repair.

White mineral trioxide aggregate mixed with calcium chloride dihydrate: chemical analysis and biological properties

OBJECTIVES: This study aimed to evaluate the chemical and biological properties of fast-set white mineral trioxide aggregate (FS WMTA), which was WMTA combined with calcium chloride dihydrate (CaCl₂·2H₂O), compared to that of WMTA. MATERIALS AND METHODS: Surface morphology, elemental, and phase analysis were examined using scanning electron microscope (SEM), energy dispersive X-ray microanalysis (EDX), and X-ray diffraction (XRD), respectively. The cytotoxicity and cell attachment properties were evaluated on human periodontal ligament fibroblasts (HPLFs) using methyl-thiazol-diphenyltetrazolium (MTT) assay and under SEM after 24 and 72 hours, respectively. RESULTS: Results showed that the addition of CaCl₂·2H₂O to WMTA affected the surface morphology and chemical composition. Although FS WMTA exhibited a non-cytotoxic profile, the cell viability values of this combination were lesser than WMTA, and the difference was significant in 7 out of 10 concentrations at the 2 time intervals (p < 0.05). HPLFs adhered over the surface of WMTA and at the interface, after 24 hours of incubation. After 72 hours, there were increased numbers of HPLFs with prominent cytoplasmic processes. Similar findings were observed with FS WMTA, but the cells were not as confluent as with WMTA. CONCLUSIONS: The addition of CaCl₂·2H₂O to WMTA affected its chemical properties. The favorable biological profile of FS WMTA towards HPLFs may have a potential impact on its clinical application for repair of perforation defects.

Angiogenic Potential of Extracellular Matrix of Human Amniotic Membrane

Combination between tissue engineering and other fields has brought an innovation in the area of regenerative medicine which ultimate aims are to repair, improve, and produce a good tissue construct. The availability of many types of scaffold, both synthetically and naturally have developed into many outstanding end products that have achieved the general objective in tissue engineering. Interestingly, most of this scaffold emulates extracellular matrix (ECM) characteristics. Therefore, ECM component sparks an interest to be explored and manipulated. The ECM featured in human amniotic membrane (HAM) provides a suitable niche for the cells to adhere, grow, proliferate, migrate and differentiate, and could possibly contribute to the production of angiogenic micro-environment indirectly. Previously, HAM scaffold has been widely used to accelerate wound healing, treat bone related and ocular diseases, and involved in cardiovascular repair. Also, it has been used in the angiogenicity study, but with a different technical approach. In addition, both side of HAM could be used in cellularised and decellularised conditions depending on the objectives of a particular research. Therefore, it is of paramount importance to investigate the behavior of ECM components especially on the stromal side of HAM and further explore the angiogenic potential exhibited by this scaffold.

Effect of perivitelline fluid from horseshoe crab on the expression of COL1A1 in dental pulp stem cells

Perivitelline fluid, extracted from the fertilized eggs of horseshoe crabs, has been reported to play a vital role in supporting embryogenesis as well as cell proliferation. The present study aims to evaluate the effect of PVF on the expression of COL1A1 in human dental pulp stem cells (DPSCs). The cells were grouped into two; untreated (control) and treated with a single dose of PVF (0.019 mg/ml). Gene expression was quantified for COL1A1 on day 1, 3 and 7 using reverse transcriptase PCR. The expression of COL1A1 on day 3 of treated group with PVF was the highest though there was a decline of COL1A1 expression on day 7. Mann Whitney test was utilized to determine the significance of COL1A1 expression between treated and untreated groups. Significant difference in the expression of COL1A1 was observed between the treated and untreated groups on day 3 though there was no significance in the expression on day 7. The present study indicates that PVF may have the potential to increase cell proliferation in human DPSCs.

Effects of perivitelline fluid obtained from horseshoe crab on the proliferation and genotoxicity of dental pulp stem cells

Perivitelline fluid [PVF] of the horseshoe crab embryo has been reported to possess an important role during embryogenesis by promoting cell proliferation. This study aims to evaluate the effect of PVF on the proliferation, chromosome aberration [CA] and mutagenicity of the dental pulp stem cells [DPSCs]. This is an in vitro experimental study. PVF samples were collected from horseshoe crabs from beaches in Malaysia and the crude extract was prepared. DPSCs were treated with different concentrations of PVF crude extract in an 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide [MTT] assay [cytotoxicity test]. We choose two inhibitory concentrations [IC[50] and IC[25]] and two PVF concentrations which produced more cell viability compared to a negative control [100%] for further tests. Quantitative analysis of the proliferation activity of PVF was studied using the AlamarBlue[Registered sign] assay for 10 days. Population doubling times [PDTs] of the treatment groups were calculated from this assay. Genotoxicity was evaluated based on the CA and Ames tests. Statistical analysis was carried out using independent t test to calculate significant differences in the PDT and mitotic indices in the CA test between the treatment and negative control groups. Significant differences in the data were P<0.05. A total of four PVF concentrations retrieved from the MTT assay were 26.887 mg/ml [IC[50]], 14.093 mg/ml [IC[25]], 0.278 mg/ml [102% cell viability] and 0.019 mg/ml [102.5% cell viability]. According to the AlamarBlue[Registered sign] assay, these PVF groups produced comparable proliferation activities compared to the negative [untreated] control. PDTs between PVF groups and the negative control were insignificantly different [P>0.05]. No significant aberrations in chromosomes were observed in the PVF groups and the Ames test on the PVF showed the absence of significant positive results. PVF from horseshoe crabs produced insignificant proliferative activity on treated DPSCs. The PVF was non-genotoxic based on the CA and Ames tests

Effects of different processing methods of human amniotic membrane on the quality of extracted RNA

The aim of this study was to determine the efficiency of different human amniotic membrane (HAM) processing methods on the concentration, purity and integrity of RNA. Two different techniques (Technique 1 and Technique 2) were employed for the processing of HAM, which differed in terms of washing solution, sample storage conditions and processing time. Based on preservation of HAM, three groups were formed under each technique. In Technique 1, the groups were fresh frozen 1 (F1), glycerol preserved (GP) and gamma irradiated glycerol preserved (IGP); where else in Technique 2, the groups were fresh frozen 2 (F2), 50% glycerol/Dulbecco’s modified Eagle medium (DMEM) cryopreserved HAM diluted with phosphate buffered saline (GB) and 50% glycerol/DMEM cryopreserved HAM diluted with diethylprocarbonate water (GD). Total RNA was extracted from the samples and their concentration, purity and integrity were examined. The F2 sample of which there was no pre-washing step and involved direct sample storage at -80ºC, shorter processing time and chilled processing conditions had yielded better quality of RNA compared to the others.

Keloid Scarring: Understanding the Genetic Basis, Advances, and Prospects

Keloid disease is a fibroproliferative dermal tumor with an unknown etiology that occurs after a skin injury in genetically susceptible individuals. Increased familial aggregation, a higher prevalence in certain races, parallelism in identical twins, and alteration in gene expression all favor a remarkable genetic contribution to keloid pathology. It seems that the environment triggers the disease in genetically susceptible individuals. Several genes have been implicated in the etiology of keloid disease, but no single gene mutation has thus far been found to be responsible. Therefore, a combination of methods such as association, gene-gene interaction, epigenetics, linkage, gene expression, and protein analysis should be applied to determine keloid etiology.

Genotoxicity assessment of locally produced dental nanocomposite using Comet assay

The aim of this study was to determine the genotoxicity of a locally produced nanocomposite by Universiti Sains Malaysia, Malaysia using Comet assay. Stem cells from human exfoliated deciduous teeth (SHED) were treated with the nanocomposite at five different concentrations (0.006, 0.0125, 0.025, 0.05, and 0.1 mg/ml) along with concurrent negative (medium alone) and positive control (zinc sulfate heptahydrate) and incubated at 37°C for 24 hours in an incubator at 5% CO2. The tail moment was used to assess the extent of DNA damage. The tail moment for the group of SHED treated with nanocomposite (for all the five different concentrations) was not statistically significant as compared to the negative control, suggesting that the locally produced dental nanocomposite did not induce any DNA damage. Hence, it can be concluded that the locally produced nanocomposite is non-genotoxic on stem cells from human exfoliated deciduous teeth.

Cytogenetics: Past, Present And Future

Fifty years have elapsed since the discovery of the number of human chromosomes in 1956. Newer techniques have been developed since then, ranging from the initial conventional banding techniques to the currently used molecular array comparative genomic hybridisation. With a combination of these conventional and molecular techniques, cytogenetics has become an indispensable tool for the diagnosis of various genetic disorders, paving the way for possible treatment and management. This paper traces the history and evolution of cytogenetics leading up to the current state of technology.