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Differentiation capacity of mouse dental pulp stem cells into osteoblasts and osteoclasts

Kermani S, Megat Abdul Wahab R, Zarina Zainol Abidin I, Zainal Ariffin Z, Senafi S, Hisham Zainal Ariffin S

Cell J, 2014 Feb 3;16(1):31-42.
PMID: 24518973

Our research attempted to show that mouse dental pulp stem cells (DPSCs) with characters such as accessibility, propagation and higher proliferation rate can provide an improved approach for generate bone tissues. With the aim of finding and comparing the differentiation ability of mesenchymal stem cells derived from DPSCs into osteoblast and osteoclast cells; morphological, molecular and biochemical analyses were conducted.
Fulltext In vitro chondrogenesis transformation study of mouse dental pulp stem cells

Zainal Ariffin SH, Kermani S, Megat Abdul Wahab R, Senafi S, Zainal Ariffin Z, Abdul Razak M

ScientificWorldJournal, 2012;2012:827149.
PMID: 22919354 DOI: 10.1100/2012/827149

A major challenge in the application of mesenchymal stem cells in cartilage reconstruction is that whether the cells are able to differentiate into fully mature chondrocytes before grafting. The aim of this study was to isolate mouse dental pulp stem cells (DPSC) and differentiate them into chondrocytes. For this investigation, morphological, molecular, and biochemical analyses for differentiated cells were used. To induce the chondrocyte differentiation, DPSC were cultured in chondrogenic medium (Zen-Bio, Inc.). Based on morphological analyses using toluidine blue staining, proteoglycan products appear in DPSC after 21 days of chondrocyte induction. Biochemical analyses in differentiated group showed that alkaline phosphatase activity was significantly increased at day 14 as compared to control (P < 0.05). Cell viability analyses during the differentiation to chondrocytes also showed that these cells were viable during differentiation. However, after the 14th day of differentiation, there was a significant decrease (P < 0.05) in the viability proportion among differentiated cells as compared to the control cells. In RT-PCR molecular analyses, mouse DPSC expressed Cd146 and Cd166 which indicated that these cells belong to mesenchymal stem cells. Coll I and Coll II markers showed high expression after 14 and 21 days, respectively. In conclusion, this study showed that DPSC successfully differentiated into chondrocytes.
Fulltext Cellular and molecular changes in orthodontic tooth movement

Zainal Ariffin SH, Yamamoto Z, Zainol Abidin IZ, Megat Abdul Wahab R, Zainal Ariffin Z

ScientificWorldJournal, 2011;11:1788-803.
PMID: 22125437 DOI: 10.1100/2011/761768

Tooth movement induced by orthodontic treatment can cause sequential reactions involving the periodontal tissue and alveolar bone, resulting in the release of numerous substances from the dental tissues and surrounding structures. To better understand the biological processes involved in orthodontic treatment, improve treatment, and reduce adverse side effects, several of these substances have been proposed as biomarkers. Potential biological markers can be collected from different tissue samples, and suitable sampling is important to accurately reflect biological processes. This paper covers the tissue changes that are involved during orthodontic tooth movement such as at compression region (involving osteoblasts), tension region (involving osteoclasts), dental root, and pulp tissues. Besides, the involvement of stem cells and their development towards osteoblasts and osteoclasts during orthodontic treatment have also been explained. Several possible biomarkers representing these biological changes during specific phenomenon, that is, bone remodelling (formation and resorption), inflammation, and root resorption have also been proposed. The knowledge of these biomarkers could be used in accelerating orthodontic treatment.
Analyses of basal media and serum for in vitro expansion of suspension peripheral blood mononucleated stem cell

Zainal Ariffin SH, Mohamed Rozali NA, Megat Abdul Wahab R, Senafi S, Zainol Abidin IZ, Zainal Ariffin Z

Cytotechnology, 2016 Aug;68(4):675-86.
PMID: 26231833 DOI: 10.1007/s10616-014-9819-8

Transplantation of stem cells requires a huge amount of cells, deeming the expansion of the cells in vitro necessary. The aim of this study is to define the optimal combination of basal medium and serum for the expansion of suspension peripheral blood mononucleated stem cells (PBMNSCs) without resulting in loss in the differentiation potential. Mononucleated cells were isolated from both mice and human peripheral blood samples through gradient centrifugation and expanded in α-MEM, RPMI, MEM or DMEM supplemented with either NBCS or FBS. The suspension cells were then differentiated to osteoblast. Our data suggested that α-MEM supplemented with 10 % (v/v) NBCS gives the highest fold increase after 14 days of culture for both mice and human PBMNSCs, which were ~1.51 and ~2.01 times, respectively. The suspension PBMNSCs in the respective medium were also able to maintain osteoblast differentiation potential as supported by the significant increase in ALP specific activity. The cells are also viable during the differentiated states when using this media. All these data strongly suggested that α-MEM supplemented with 10 % NBCS is the best media for the expansion of both mouse and human suspension PBMNSCs.
Fulltext Impact of isolation method on doubling time and the quality of chondrocyte and osteoblast differentiated from murine dental pulp stem cells

Megat Abdul Wahab R, Mohamed Rozali NA, Senafi S, Zainol Abidin IZ, Zainal Ariffin Z, Zainal Ariffin SH

PeerJ, 2017;5:e3180.
PMID: 28626603 DOI: 10.7717/peerj.3180

BACKGROUND: Stem cells are normally isolated from dental pulps using the enzymatic digestion or the outgrowth method. However, the effects of the isolation method on the quality of the isolated stem cells are not studied in detail in murine models. The aim of this study was to compare the matrices secreted by osteoblast and chondrocytes differentiated from dental pulp stem cells isolated through different means.
METHOD: DPSC from murine incisors were isolated through either the outgrowth (DPSC-OG) or the enzymatic digestion (DPSC-ED) method. Cells at passage 4 were used in this study. The cells were characterized through morphology and expression of cell surface markers. The cells' doubling time when cultured using different seeding densities was calculated and analyzed using one-way ANOVA and Tukey's multiple comparison post-test. The ability of cells to differentiate to chondrocyte and osteoblast was evaluated through staining and analysis on the matrices secreted.
RESULTS: Gene expression analysis showed that DPSC-OG and DPSC-ED expressed dental pulp mesenchymal stem cell markers, but not hematopoietic stem cell markers. The least number of cells that could have been used to culture DPSC-OG and DPSC-ED with the shortest doubling time was 5 × 10(2) cells/cm(2) (11.49 ± 2.16 h) and 1 × 10(2) cells/cm(2) (10.55 h ± 0.50), respectively. Chondrocytes differentiated from DPSC-ED produced 2 times more proteoglycan and at a faster rate than DPSC-OG. FTIR revealed that DPSC-ED differentiated into osteoblast also secreted matrix, which more resembled a calvaria.
DISCUSSION: Isolation approaches might have influenced the cell populations obtained. This, in turn, resulted in cells with different proliferation and differentiation capability. While both DPSC-OG and DPSC-ED expressed mesenchymal stem cell markers, the percentage of cells carrying each marker might have differed between the two methods. Regardless, enzymatic digestion clearly yielded cells with better characteristics than outgrowth.
Fulltext Synthesis, Characterisation and Antibacterial Properties of Silicone-Silver Thin Film for the Potential of Medical Device Applications

Aizamddin MF, Mahat MM, Zainal Ariffin Z, Samsudin I, Razali MSM, Amir M'

Polymers (Basel), 2021 Nov 05;13(21).
PMID: 34771378 DOI: 10.3390/polym13213822

Silver (Ag) particles have sparked considerable interest in industry and academia, particularly for health and medical applications. Here, we present the "green" and simple synthesis of an Ag particle-based silicone (Si) thin film for medical device applications. Drop-casting and peel-off techniques were used to create an Si thin film containing 10-50% (v/v) of Ag particles. Electro impedance spectroscopy (EIS), X-ray diffraction analysis (XRD), scanning electron microscopy (SEM), energy dispersive X-ray (EDX), and tensile tests were used to demonstrate the electrical conductivity, crystallinity, morphology-elemental, and mechanical properties, respectively. The oriented crystalline structure and excellent electronic migration explained the highest conductivity value (1.40 × 10-5 S cm-1) of the 50% Ag-Si thin film. The findings regarding the evolution of the conductive network were supported by the diameter and distribution of Ag particles in the Si film. However, the larger size of the Ag particles in the Si film resulted in a lower tensile stress of 68.23% and an elongation rate of 68.25% compared to the pristine Si film. The antibacterial activity of the Ag-Si film against methicillin-resistant Staphylococcus aureus (MRSA), Bacillus cereus (B. cereus), Klebsiella pneumoniae (K. pneumoniae), and Pseudomonas aeruginosa (P. aeruginosa) was investigated. These findings support Si-Ag thin films' ability to avoid infection in any medical device application.
Fulltext Intrinsic anticarcinogenic effects of Piper sarmentosum ethanolic extract on a human hepatoma cell line

Zainal Ariffin SH, Wan Omar WH, Zainal Ariffin Z, Safian MF, Senafi S, Megat Abdul Wahab R

Cancer Cell Int, 2009;9:6.
PMID: 19257877 DOI: 10.1186/1475-2867-9-6

Piper sarmentosum, locally known as kaduk is belonging to the family of Piperaceae. It is our interest to evaluate their effect on human hepatoma cell line (HepG2) for the potential of anticarcinogenic activity.
Fulltext Stem cell heterogeneity of mononucleated cells from murine peripheral blood: molecular analysis

Yazid MD, Zainal Ariffin SH, Senafi S, Zainal Ariffin Z, Megat Abdul Wahab R

ScientificWorldJournal, 2011;11:2150-9.
PMID: 22125464 DOI: 10.1100/2011/340278

The main purpose of this paper was to determine the heterogeneity of primary isolated mononucleated cells that originated from the peripheral blood system by observing molecular markers. The isolated cells were cultured in complete medium for 4 to 7 days prior to the separation of different cell types, that is, adherent and suspension. Following a total culture time of 14 days, adherent cells activated the Cd105 gene while suspension cells activated the Sca-1 gene. Both progenitor markers, Cbfa-1 and Ostf-1, were inactivated in both suspension and adherent cells after 14-day culture compared to cells cultured 3 days in designated differentiation medium. In conclusion, molecular analyses showed that primary mononucleated cells are heterogeneous, consisting of hematopoietic stem cells (suspension) and mesenchymal stem cells (adherent) while both cells contained no progenitor cells.
Fulltext Differentiation of dental pulp stem cells into neuron-like cells in serum-free medium

Zainal Ariffin SH, Kermani S, Zainol Abidin IZ, Megat Abdul Wahab R, Yamamoto Z, Senafi S, et al.

Stem Cells Int, 2013;2013:250740.
PMID: 24348580 DOI: 10.1155/2013/250740

Dental pulp tissue contains dental pulp stem cells (DPSCs). Dental pulp cells (also known as dental pulp-derived mesenchymal stem cells) are capable of differentiating into multilineage cells including neuron-like cells. The aim of this study was to examine the capability of DPSCs to differentiate into neuron-like cells without using any reagents or growth factors. DPSCs were isolated from teeth extracted from 6- to 8-week-old mice and maintained in complete medium. The cells from the fourth passage were induced to differentiate by culturing in medium without serum or growth factors. RT-PCR molecular analysis showed characteristics of Cd146(+) , Cd166(+) , and Cd31(-) in DPSCs, indicating that these cells are mesenchymal stem cells rather than hematopoietic stem cells. After 5 days of neuronal differentiation, the cells showed neuron-like morphological changes and expressed MAP2 protein. The activation of Nestin was observed at low level prior to differentiation and increased after 5 days of culture in differentiation medium, whereas Tub3 was activated only after 5 days of neuronal differentiation. The proliferation of the differentiated cells decreased in comparison to that of the control cells. Dental pulp stem cells are induced to differentiate into neuron-like cells when cultured in serum- and growth factor-free medium.
Fulltext The effects of Piper sarmentosum aqueous extracts on zebrafish (Danio rerio) embryos and caudal fin tissue regeneration

Zainol Abidin IZ, Fazry S, Jamar NH, Ediwar Dyari HR, Zainal Ariffin Z, Johari AN, et al.

Sci Rep, 2020 08 25;10(1):14165.
PMID: 32843675 DOI: 10.1038/s41598-020-70962-7

In Malaysia, Piper sarmentosum or 'kaduk' is commonly used in traditional medicines. However, its biological effects including in vivo embryonic toxicity and tissue regenerative properties are relatively unknown. The purpose of this study was to determine zebrafish (Danio rerio) embryo toxicities and caudal fin tissue regeneration in the presence of P. sarmentosum aqueous extracts. The phytochemical components and antioxidant activity of the extract were studied using GC-MS analysis and DPPH assay, respectively. Embryo toxicity tests involving survival, heartbeat, and morphological analyses were conducted to determine P. sarmentosum extract toxicity (0-60 µg/mL); concentrations of 0-400 µg/mL of the extract were used to study tissue regeneration in the zebrafish caudal fin. The extract contained several phytochemicals with antioxidant activity and exhibited DPPH scavenging activity (IC50 = 50.56 mg/mL). Embryo toxicity assays showed that a concentration of 60 μg/mL showed the highest rates of lethality regardless of exposure time. Slower embryogenesis was observed at 40 µg/mL, with non-viable embryos first detected at 50 µg/mL. Extracts showed significant differences (p
Advocating Electrically Conductive Scaffolds with Low Immunogenicity for Biomedical Applications: A Review

Ahmad Ruzaidi DA, Mahat MM, Shafiee SA, Mohamed Sofian Z, Mohmad Sabere AS, Ramli R, et al.

Polymers (Basel), 2021 Oct 02;13(19).
PMID: 34641210 DOI: 10.3390/polym13193395

Scaffolds support and promote the formation of new functional tissues through cellular interactions with living cells. Various types of scaffolds have found their way into biomedical science, particularly in tissue engineering. Scaffolds with a superior tissue regenerative capacity must be biocompatible and biodegradable, and must possess excellent functionality and bioactivity. The different polymers that are used in fabricating scaffolds can influence these parameters. Polysaccharide-based polymers, such as collagen and chitosan, exhibit exceptional biocompatibility and biodegradability, while the degradability of synthetic polymers can be improved using chemical modifications. However, these modifications require multiple steps of chemical reactions to be carried out, which could potentially compromise the end product's biosafety. At present, conducting polymers, such as poly(3,4-ethylenedioxythiophene) poly(4-styrenesulfonate) (PEDOT: PSS), polyaniline, and polypyrrole, are often incorporated into matrix scaffolds to produce electrically conductive scaffold composites. However, this will reduce the biodegradability rate of scaffolds and, therefore, agitate their biocompatibility. This article discusses the current trends in fabricating electrically conductive scaffolds, and provides some insight regarding how their immunogenicity performance can be interlinked with their physical and biodegradability properties.
Fulltext Gene expression profiles for in vitro human stem cell differentiation into osteoblasts and osteoclasts: a systematic review

Zainal Ariffin SH, Lim KW, Megat Abdul Wahab R, Zainal Ariffin Z, Rus Din RD, Shahidan MA, et al.

PeerJ, 2022;10:e14174.
PMID: 36275474 DOI: 10.7717/peerj.14174

BACKGROUND: There have been promising results published regarding the potential of stem cells in regenerative medicine. However, the vast variety of choices of techniques and the lack of a standard approach to analyse human osteoblast and osteoclast differentiation may reduce the utility of stem cells as a tool in medical applications. Therefore, this review aims to systematically evaluate the findings based on stem cell differentiation to define a standard gene expression profile approach.
METHODS: This review was performed following the PRISMA guidelines. A systematic search of the study was conducted by retrieving articles from the electronic databases PubMed and Web of Science to identify articles focussed on gene expression and approaches for osteoblast and osteoclast differentiation.
RESULTS: Six articles were included in this review; there were original articles of in vitro human stem cell differentiation into osteoblasts and osteoclasts that involved gene expression profiling. Quantitative polymerase chain reaction (qPCR) was the most used technique for gene expression to detect differentiated human osteoblasts and osteoclasts. A total of 16 genes were found to be related to differentiating osteoblast and osteoclast differentiation.
CONCLUSION: Qualitative information of gene expression provided by qPCR could become a standard technique to analyse the differentiation of human stem cells into osteoblasts and osteoclasts rather than evaluating relative gene expression. RUNX2 and CTSK could be applied to detect osteoblasts and osteoclasts, respectively, while RANKL could be applied to detect both osteoblasts and osteoclasts. This review provides future researchers with a central source of relevant information on the vast variety of gene expression approaches in analysing the differentiation of human osteoblast and osteoclast cells. In addition, these findings should enable researchers to conduct accurately and efficiently studies involving isolated human stem cell differentiation into osteoblasts and osteoclasts.

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