Revisiting Genetics of Cleft Lip with or without Cleft Palate and Cleft Palate Only: A Narrative Review

@#Cleft lip with or without cleft palate (CLP) and cleft palate only (CP) are the most common orofacial deformities observed in humans where almost 1 in 700 to 1 in 2,000 babies born each year are affected worldwide. This condition occurs when the specific and independently derived facial primordial fails to fuse together, hence forming the cleft of the lip and palate or the palate alone. These orofacial abnormalities can be divided into syndromic and non-syndromic where the deformities are either associated with other disorders or present on their own, respectively. It is important to understand every step in the lip and palate development during the embryonic stage to pinpoint the exact problem affecting the normal development of the human face. With current technologies, more genes are identified to be associated with and cause CLP and CP in humans. Therefore, this review aims to elaborate on the latest updates on the genetics of CLP and CP. Polymorphism in some of the genes has been associated with the incidence of these anomalies. Identification of these genes provides new knowledge on how these craniofacial abnormalities occur and hopefully will enable earlier treatment of these deformities to be implemented.

FLAXSEED (LINUM USITATISSIMUM) EXTRACT ACTIVITY ON HUMAN ORAL FIBROBLASTS (HOrF) CELL LINE

@#Natural products have demonstrated various activities beneficial to general health. Flaxseed (Linum usitatissimum) has been reported in many studies for its antimicrobial, antioxidant, and anti-inflammatory effects. Additionally, flaxseed extracts have skin wound healing activity and potential for treating oral ulcers. L. usitatissimum was extracted using 70% ethanol via soxhlet method and gas chromatography mass spectrum (GCMS) was used to analyze the components of L. usitatissimum extract. The crude flaxseed oil were applied to human oral fibroblasts (HOrF), and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay was used to assess the cell viability after 24, 48 and 72 hours. Scratched HOrF cells were treated with crude flaxseed oil and healing was monitored per wound healing assay. GC-MS indicate that the major components present in L. usitatissimum oil extract are linolic, palmitic and oleic acid. L. usitatissimum crude oil extract showed high proliferation effect on HOrF cells at 24 and 48 hours, while the highest proliferation effect was recorded at 72 hours post-treatment. The wound healing assay results showed that healing activity of HOrF cells occurred as soon as 18 hours post-treatment when treated with L. usitatissimum crude oil extract. L. usitatissimum crude oil extract has proliferating and healing effects on HOrF cell line. Therefore, it can be considered as a potential promising oral wound healing agent.

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.

Effect of perivitelline fluid from horseshoe crab on the expression of cell cycle regulatory genes in human 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 expression of cell cycle regulatory genes from human dental pulp stem cells (DPSCs) between different cell passages viz. 4, 5, 6. The cells were treated with a single dose of PVF (26.89 mg/ml) PVF. Gene expression was quantified for CDKNA2A, PTEN, MDM2 and TP53 genes using reverse transcriptase PCR. CDKN2A and MDM2 expression for treated and untreated DPSCs, expressed a similar pattern of expression. The higher expression of CDKN2A showed that the treatment increased cell proliferation and prevented cell senescence. DPSCs with PVF treatment showed increased expression of MDM2 at passage 4 and drastically declined expression at passage 5 and slightly increased at passage 6. TP53 expression of DPSCs treated group showed a higher expression compared to untreated group. On the other hand, the expression of PTEN in DPSCs treated group started to increase from passage 5 to 6. However, on the whole, the PTEN expression was higher than the untreated group in all the passages studied here. The results showed that PVF could enhance cell cycle regulatory gene expression in DPSCs as indicated by the higher expression of all the genes considered in this study at different cell passages in the treated group compared to the untreated group. Mann Whitney test was utilized to determine the significance of cell cycle regulatory genes expression between treated and untreated group. Significant difference in expression of genes between the treated and untreated groups were found at all passages except for CDKN2A gene whereby, its expression was not significantly different at passage 5 though it did express slightly higher in PVF treated DPSCs.

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.

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

Absence of nucleotide alteration in region of exon 34 of NOTCH1 and NOTCH2 receptor genes analysed in oral cancer samples: a preliminary observation

Oral cancer is one of the common cancer cases identified in the developing countries. Genetic mutation and overexpression of certain genes and proteins have been associated in the development of this cancer. Notch signalling pathway is normally involved in controlling the development process of vertebrates and invertebrates; however, deregulation of this pathway was found to be responsible in the formation of certain cancers including oral cancers. Activation of this pathway requires binding of the ligands to its receptors. Four NOTCH receptors (NOTCH 1, 2, 3 and 4) have been identified in mammals. Disruptions within these molecules might interfere with the normal functions of Notch signalling pathway. Hence, this study was conducted to detect mutations of NOTCH1 and NOTCH2 receptor genes which might be occurring in the oral cancer cases obtained from the local population. DNA extracted from fresh-frozen tissue biopsy of the tongue and buccal mucosa from 10 confirmed cases of oral cancer were subjected for polymerase chain reaction (PCR) amplification using the specific sets of primers. The PCR products were sent for sequencing before final results were analysed. Due to time and cost limitation, only two out of four NOTCH receptor genes; NOTCH1 and NOTCH2, were used in this analysis. The results revealed absence of nucleotide changes for both NOTCH receptor genes amplified from these oral cancer samples. More samples and further analysis looking into other regions in these genes are required to conclude the involvement of NOTCH receptor genes mutation in causing oral cancer.