Candida albicans is an opportunistic fungal pathogen, which causes a plethora of superficial, as well as invasive, infections in humans. The ability of this fungus in switching from commensalism to active infection is attributed to its many virulence traits. Biofilm formation is a key process, which allows the fungus to adhere to and proliferate on medically implanted devices as well as host tissue and cause serious life-threatening infections. Biofilms are complex communities of filamentous and yeast cells surrounded by an extracellular matrix that confers an enhanced degree of resistance to antifungal drugs. Moreover, the extensive plasticity of the C. albicans genome has given this versatile fungus the added advantage of microevolution and adaptation to thrive within the unique environmental niches within the host. To combat these challenges in dealing with C. albicans infections, it is imperative that we target specifically the molecular pathways involved in biofilm formation as well as drug resistance. With the advent of the -omics era and whole genome sequencing platforms, novel pathways and genes involved in the pathogenesis of the fungus have been unraveled. Researchers have used a myriad of strategies including transcriptome analysis for C. albicans cells grown in different environments, whole genome sequencing of different strains, functional genomics approaches to identify critical regulatory genes, as well as comparative genomics analysis between C. albicans and its closely related, much less virulent relative, C. dubliniensis, in the quest to increase our understanding of the mechanisms underlying the success of C. albicans as a major fungal pathogen. This review attempts to summarize the most recent advancements in the field of biofilm and antifungal resistance research and offers suggestions for future directions in therapeutics development.
Culture expanded chondrocytes isolated from non-load bearing region of osteoarthritic (OA) joint has been used to construct tissue engineered cartilage for treatment purposes. The aim of the study was to compare the histological properties of the cartilage tissue and morphological properties of the chondrocytes isolated from less and severely affected OA knee. Human articular cartilage was obtained as redundant tissue from consented patients with late-stage OA undergoing total knee replacement surgery at Universiti Kebangsaan Malaysia Medical Centre (UKMMC). Articular cartilage was graded according to Dougados and Osteoarthritis Research Society International (OARSI) classification. Articular cartilage was classified into less affected (LA; Grade 0-1) and severely affected (SA; Grade 2-3). Cartilage tissue from less and severely affected region was stained with Safranin O staining. Isolated chondrocytes from each group were cultured until passage 4 (P4). Their growth patterns, cell areas, and circularity were compared. LA-cartilage tissue shows uniform spread of safranin O staining indicating intact extracellular matrix (ECM) component. However, SA-cartilage shows significant reduction and unstable staining due to its degraded ECM. LA-chondrocytes showed an aggregated growth compared to SA-chondrocyte that remains monolayer. Moreover, LA-chondrocytes have significantly higher cell area with wider spreading at passage 0 and 4 compared to SA-chondrocytes. It was also found that chondrocyte circularity increased with passage, and circularity of LAchondrocytes was significantly higher than that of the SA-chondrocytes at passage 3. This study demonstrated the considerable difference in the cellular properties for less and severely affected chondrocytes and implication of these differences in cell-based therapy needed to be explored.
Electrospinning is a simple and versatile technique to fabricate continuous fibers with diameter ranging from micrometers to a few nanometers. To date, the number of polymers that have been electrospun has exceeded 200. In recent years, electrospinning has become one of the most popular scaffold fabrication techniques to prepare nanofiber mesh for tissue engineering applications. Collagen, the most abundant extracellular matrix protein in the human body, has been electrospun to fabricate biomimetic scaffolds that imitate the architecture of native human tissues. As collagen nanofibers are mechanically weak in nature, it is commonly cross-linked or blended with synthetic polymers to improve the mechanical strength without compromising the biological activity. Electrospun collagen nanofiber mesh has high surface area to volume ratio, tunable diameter and porosity, and excellent biological activity to regulate cell function and tissue formation. Due to these advantages, collagen nanofibers have been tested for the regeneration of a myriad of tissues and organs. In this review, we gave an overview of electrospinning, encompassing the history, the instrument settings, the spinning process and the parameters that affect fiber formation, with emphasis given to collagen nanofibers' fabrication and application, especially the use of collagen nanofibers in skin tissue engineering.
This study determined the antiaging effect of stingless bee honey on the expression of extracellular matrix genes. MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt) assay was performed for determination of optimum concentration and incubation time of stingless bee honey. Gene expression of matrix metalloproteinase-1 (MMP-1) and collagen type Ⅰ (COL1A1) were analyzed using real time reverse transcriptase polymerase chain reaction technique. Incubation with stingless bee honey at concentration of 0.02% for 72 hr showed significant increase in the viability of human fibroblast cells. Stingless bee honey significantly downregulates metalloproteinase-1 gene expression in both pre-senescence and senescence fibroblast cells and upregulates collagen type Ⅰ gene expression in senescence fibroblast cells. In conclusion, stingless bee honey potentially delayed skin aging through modulation of extracellular matrix genes. PRACTICAL APPLICATIONS: Changes of the extracellular matrix regulation promote skin aging. Stingless bee honey is a good source of natural antioxidant which potentially delays skin aging. This study demonstrated that stingless bee honey beneficially increases collagen type Ⅰ expression and decreases MMP-1 expression during cellular aging of human dermal fibroblast cells.
The use of hybridisation strategy in biomaterials technology provides a powerful synergistic effect as a functional matrix. Silk fibroin (SF) has been widely used for drug delivery, and collagen (Col) resembles the extracellular matrix (ECM). This systematic review was performed to scrutinise the outcome of hybrid Col and SF for cutaneous wound healing. This paper reviewed the progress of related research based on in vitro and in vivo studies and the influence of the physicochemical properties of the hybrid in wound healing. The results indicated the positive outcome of hybridising Col and SF for cutaneous wound healing. The hybridisation of these biomaterials exhibits an excellent moisturising property, perfectly interconnected structure, excellent water absorption and retention capacity, an acceptable range of biodegradability, and synergistic effects in cell viability. The in vitro and in vivo studies clearly showed a promising outcome in the acceleration of cutaneous wound healing using an SF and Col hybrid scaffold. The review of this study can be used to design an appropriate hybrid scaffold for cutaneous wound healing. Therefore, this systematic review recapitulated that the hybridisation of Col and SF promoted rapid cutaneous healing through immediate wound closure and reepithelisation, with no sign of adverse events. This paper concludes on the need for further investigations of the hybrid SF and Col in the future to ensure that the hybrid biomaterials are well-suited for human skin.
The term microfibril angle, MFA in wood science refers to the angle between the direction of the helical windings of cellulose microfibrils in the secondary cell wall, S2 layer of fibres and tracheids and the long axis of the cell. In this study, the mean MFA of the cell walls were determined for thin samples of thickness 200.0 µm from pith and outwards, for eight ages of Acacia mangium wood. The determination of MFA was based on a diffraction pattern arising from cellulose crystal planes of the type 002 generated by x-ray diffraction and recorded using an electronic detector. The results show an inversely relationship between MFA and age of tree in Acacia mangium wood. MFA decreased from 26.13° at age 3 year-old to 0.20° at tree of age 15 year-old for the pith region. The most significant drop occurred from 16.14° at age 7 yearold to 11.30° at age 9 year-old. An inversely relationship between MFA and storage modulus E’ was evidence in Acacia mangium at age 10-year-old. The results showed that about 76.22% variation of loss modulus E” was attributed to the MFA, while about 66.4% of the variation of glass transition Tg was explained by MFA under the same experimental conditions.
In recent years, three-dimensional (3D) in vitro cell culture models have earned great attention, especially in the field of human cancer disease modelling research as they provide a promising alternative towards the conventional two-dimensional (2D) monolayer culture of cells with improved tissue organization. In 2D cell culture systems, the complexity of cells on a planar surface does not accurately reflects the in vivo cellular microenvironment. Cells propagated in 3D cell culture model, on the other hand, exhibit physiologically relevant cell-to-cell interactions and cell-to-extracellular matrix (ECM) interactions, important in maintaining a normal homeostasis and specificity of tissues. This review gives an overview on 2D models and their limitations, followed by 3D cell culture models, their advantages, drawbacks and challenges in present perspectives. The review also highlights the dissimilarities of 2D and 3D models and the applicability of 3D models in current cancer research
The current trend of burn wound care has shifted to more holistic approach of improvement in the long-term form and function of the healed burn wounds and quality of life. This has demanded the emergence of various skin substitutes in the management of acute burn injury as well as post burn reconstructions. Skin substitutes have important roles in the treatment of deep dermal and full thickness wounds of various aetiologies. At present, there is no ideal substitute in the market. Skin substitutes can be divided into two main classes, namely, biological and synthetic substitutes. The biological skin substitutes have a more intact extracellular matrix structure, while the synthetic skin substitutes can be synthesised on demand and can be modulated for specific purposes. Each class has its advantages and disadvantages. The biological skin substitutes may allow the construction of a more natural new dermis and allow excellent re-epithelialisation characteristics due to the presence of a basement membrane. Synthetic skin substitutes demonstrate the advantages of increase control over scaffold composition. The ultimate goal is to achieve an ideal skin substitute that provides an effective and scar-free wound healing.
Natural cellular microenvironment consists of spatiotemporal gradients of multiple physical (e.g. extracellular matrix stiffness, porosity and stress/strain) and chemical cues (e.g. morphogens), which play important roles in regulating cell behaviors including spreading, proliferation, migration, differentiation and apoptosis, especially for pathological processes such as tumor formation and progression. Therefore, it is essential to engineer cellular gradient microenvironment incorporating various gradients for the fabrication of normal and pathological tissue models in vitro. In this article, we firstly review the development of engineering cellular physical and chemical gradients with cytocompatible hydrogels in both two-dimension and three-dimension formats. We then present current advances in the application of engineered gradient microenvironments for the fabrication of disease models in vitro. Finally, concluding remarks and future perspectives for engineering cellular gradients are given.
Diabetes now is the most common chronic disease in the world inducing heavy burden for the people's health. Based on this, diabetes research such as islet function has become a hot topic in medical institutes of the world. Today, in medical institutes, the conventional experiment platform in vitro is monolayer cell culture. However, with the development of micro- and nano-technologies, several microengineering methods have been developed to fabricate three-dimensional (3D) islet models in vitro which can better mimic the islet of pancreases in vivo. These in vitro islet models have shown better cell function than monolayer cells, indicating their great potential as better experimental platforms to elucidate islet behaviors under both physiological and pathological conditions, such as the molecular mechanisms of diabetes and clinical islet transplantation. In this review, we present the state-of-the-art advances in the microengineering methods for fabricating microscale islet models in vitro. We hope this will help researchers to better understand the progress in the engineering 3D islet models and their biomedical applications such as drug screening and islet transplantation.
Fibrin has excellent biocompatibility and biological properties to support tissue regeneration and promote wound healing. However, the role of diluted fibrin in wound healing has yet to be elucidated as it is commonly used in high concentration. This study was aimed to examine the effects of diluted plasma-derived fibrin (PDF) on keratinocyte and fibroblast wound healing in term of cell proliferation, migration, extracellular matrix (ECM) production and soluble factor secretion. Two PDF concentrations, 10 and 20% (v/v) were tested on keratinocytes and fibroblasts indirectly co-cultured in the transwell system. The control group was cultured with 5% FBS. Results showed that PDF reduced the keratinocyte growth rate and fibroblast migration, and increased the fibroblast ECM gene expression whereby significant differences were found between the 20% PDF group and the 5% FBS group. Similar trend was seen for the 10% PDF group but the differences were not significant. Comparison of the soluble factors between the PDF groups demonstrated that the level of growth-related oncogene alpha, interleukin-8 and epithelial neutrophil-activating peptide-78 were significantly higher in the 10% PDF group, whilst interleukin-1 alpha and granulocyte-macrophage colony stimulating factor were significantly more concentrated in the 20% PDF group. Our results suggested that PDF selectively elevated the expression of collagen type 1 and collagen type 3 in fibroblasts but slowed down the migration in concentration-dependent manner. These novel findings provide new insight into the role of PDF in wound healing and may have important implications for the use of fibrin in skin tissue engineering.
The aim of this systematic review was to address the question: Do different irrigating protocols have an impact on the dislocation resistance of mineral trioxide aggregate (MTA)-based materials? The review was performed using a well-defined search strategy in three databases (PubMed, Scopus, Web of Science) to include laboratory studies performed between January 1995 and May 2017, in accordance with PRISMA guidelines. Two reviewers analysed the papers, assessed the risk of bias and extracted data on teeth used, sample size, size of root canal preparation, type of MTA-based material, irrigants, canal filling method, storage method and duration, region of roots and the parameters of push-out testing (slice thickness, plunger dimensions and plunger loading direction), the main results and dislocation resistance values (in MPa). From 255 studies, 27 were included for full-text analysis. Eight papers that met the inclusion criteria were included in this review. There was a wide variation in dislocation resistance due to differences in irrigation sequence, time and concentration of irrigants, storage method and duration, and the parameters of push-out bond strength testing. A meta-analysis was not done but qualitative synthesis of the included studies was performed. No definitive conclusion could be drawn to evaluate the effect of irrigation protocols on dislocation resistance of MTA-based materials. Recommendations have been provided for standardized testing methods and reporting of future studies, so as to obtain clinically relevant information and to understand the effects of irrigating protocols on root canal sealers and their interactions with the dentine walls of root canals.
Fabrication of composite scaffolds is one of the strategies proposed to enhance the functionality of tissue-engineered scaffolds for improved tissue regeneration. By combining multiple elements together, unique biomimetic scaffolds with desirable physical and mechanical properties can be tailored for tissue-specific applications. Despite having a highly porous structure, the utility of electrospun fibers (EF) as scaffold is usually hampered by their insufficient mechanical strength. In this study, we attempted to produce a mechanically competent scaffold with cell-guiding ability by fabricating aligned poly lactic-co-glycolic acid (PLGA) fibers on decellularized human amniotic membrane (HAM), known to possess favorable tensile and wound healing properties. Decellularization of HAM in 18.75 μg/mL of thermolysin followed by a brief treatment in 0.25 M sodium hydroxide efficiently removed the amniotic epithelium and preserved the ultrastructure of the underlying extracellular matrix. The electrospinning of 20% (w/v) PLGA 50:50 polymer on HAM yielded beadless fibers with straight morphology. Subsequent physical characterization revealed that EF-HAM scaffold with a 3-min fabrication had the most aligned fibers with the lowest fiber diameter in comparison with EF-HAM 5- and 7-min scaffolds. Hydrated EF-HAM scaffolds with 3-min deposition had a greater tensile strength than the other scaffolds despite having thinner fibers. Nevertheless, wet HAM and EF-HAMs regardless of the fiber thicknesses had a significantly lower Young's modulus, and hence, a higher elasticity compared with dry HAM and EF-HAMs. Biocompatibility analysis showed that the viability and migration rate of skeletal muscle cells on EF-HAMs were similar to control and HAM alone. Skeletal muscle cells seeded on HAM were shown to display random orientation, whereas cells on EF-HAM scaffolds were oriented along the alignment of the electrospun PLGA fibers. In summary, besides having good mechanical strength and elasticity, EF-HAM scaffold design decorated with aligned fiber topography holds a promising potential for use in the development of aligned tissue constructs.
Far-flung evolution in tissue engineering enabled the development of bioactive and biodegradable materials to generate biocomposite nanofibrous scaffolds for bone repair and replacement therapies. Polymeric bioactive nanofibers are to biomimic the native extracellular matrix (ECM), delivering tremendous regenerative potentials for drug delivery and tissue engineering applications. It's been known from few decades that Zinc oxide (ZnO) nanoparticles are enhancing bone growth and providing proliferation of osteoblasts when incorporated with hydroxyapatite (HAp). We attempted to investigate the interaction between the human foetal osteoblasts (hFOB) with ZnO doped HAp incorporated biocomposite poly(L-lactic acid)-co-poly(ε-caprolactone) and silk fibroin (PLACL/SF) nanofibrous scaffolds for osteoblasts mineralization in bone tissue regeneration. The present study, we doped ZnO with HAp (ZnO(HAp) using the sol-gel ethanol condensation technique. The properties of PLACL/SF/ZnO(HAp) biocomposite nanofibrous scaffolds enhanced with doped and blended ZnO/HAp were characterized using Scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), Contact angle and Tensile studies to determine the morphology, functionality, wettability and stability. The in vitro study results showed that the addition of ZnO and HAp enhances the secretion of bone mineral matrix (98%) with smaller fiber diameter (139.4 ± 27 nm) due to the presence of silk fibroin showing potential tensile properties (322.4%), and increased the proliferation of osteoblasts for bone tissue regeneration.
Microbial infection has been shown to involve in oral carcinogenesis; however, the underlying mechanisms remain poorly understood. The present study aimed to characterize the growth of oral microorganisms as both monospecies and polymicrobial biofilms and determine the effects of their products on oral keratinocytes. Candida albicans (ALC3), Actinomyces naeslundii (AN) and Streptococcus mutans (SM) biofilms or a combination of these (TRI) were grown in flow-cell system for 24 h. The biofilms were subjected to fluorescent in situ hybridization using species-specific probes and analysed using confocal laser scanning microscopy. The effluent derived from each biofilm was collected and incubated with malignant (H357) and normal (OKF6) oral keratinocytes to assess extracellular matrix adhesion, epithelial-mesenchymal transition (EMT) and cytokines expression. Incubation of OKF6 with ALC3 and TRI effluent significantly decreased adhesion of the oral keratinocyte to collagen I, whereas incubation of H357 with similar effluent increased adhesion of the oral keratinocyte to laminin I, significantly when compared with incubation with artificial saliva containing serum-free medium (NE; P < 0.05). In OKF6, changes in E-cadherin and vimentin expression were not consistent with EMT although there was evidence of a mesenchymal to epithelial transition in malignant oral keratinocytes incubated with AN and SM effluent. A significant increase of pro-inflammatory cytokines expression, particularly interleukin (IL)-6 and IL-8, was observed when H357 was incubated with all biofilm effluents after 2- and 24-h incubation when compared with NE (P < 0.05). In conclusion, C.albicans, A.naeslundii and S.mutans form polymicrobial biofilms which differentially modulate malignant phenotype of oral keratinocytes.
Extracellular matrix (ECM) based bioscaffolds prepared by decellularization has increasingly emerged in tissue engineering application because it has structural, biochemical, and biomechanical cues that have dramatic effects upon cell behaviors. Therefore, we developed a closed sonication decellularization system to prepare ideal bioscaffolds with minimal adverse effects on the ECM. The decellularization was achieved at 170 kHz of ultrasound frequency in 0.1% and 2% Sodium Dodecyl Sulphate (SDS) solution for 10 hours. The immersion treatment as control was performed to compare the decellularization efficiency with our system. Cell removal and ECM structure were determined by histological staining and biochemical assay. Biomechanical properties were investigated by the indentation testing to test the stiffness, a residual force and compression of bioscaffolds. Additionally, in vivo implantation was performed in rat to investigate host tissue response. Compared to native tissues, histological staining and biochemical assay confirm the absence of cellularity with preservation of ECM structure. Moreover, sonication treatment has not affected the stiffness [N/mm] and a residual force [N] of the aortic scaffolds except for compression [%] which 2% SDS significantly decreased compared to native tissues showing higher SDS has a detrimental effect on ECM structure. Finally, minimal inflammatory response was observed after 1 and 5 weeks of implantation. This study reported that the novelty of our developed closed sonication system to prepare ideal bioscaffolds for tissue engineering applications.
Tissues become more rigid during tumorigenesis and have been identified as a driving factor for tumor growth. Here, we highlight the concept of tissue rigidity, contributing factors that increase tissue rigidity, and mechanisms that promote tumor growth initiated by increased tissue rigidity. Various factors lead to increased tissue rigidity, promoting tumor growth by activating focal adhesion kinase (FAK) and Rho-associated kinase (ROCK). Consequently, result in recruitment of cancer-associated fibroblasts (CAFs), epithelial-mesenchymal transition (EMT) and tumor protection from immunosurveillance. We also discussed the rationale for targeting tumor tissue rigidity and its potential for cancer treatment.
Diopside was synthesized from biowaste (Eggshell) by sol-gel combustion method at low calcination temperature and the influence of two different fuels (urea, l-alanine) on the phase formation temperature, physical and biological properties of the resultant diopside was studied. The synthesized materials were characterized by heating microscopy, FTIR, XRD, BET, SEM and EDAX techniques. BET analysis reveals particles were of submicron size with porosity in the nanometer range. Bone-like apatite deposition ability of diopside scaffolds was examined under static and circulation mode of SBF (Simulated Body Fluid). It was noticed that diopside has the capability to deposit HAP (hydroxyapatite) within the early stages of immersion. ICP-OES analysis indicates release of Ca, Mg, Si ions and removal of P ions from the SBF, but in different quantities from diopside scaffolds. Cytocompatability studies on human bone marrow stromal cells (hBMSCs) revealed good cellular attachment on the surface of diopside scaffolds and formation of extracellular matrix (ECM). This study suggests that the usage of eggshell biowaste as calcium source provides an effective substitute for synthetic starting materials to fabricate bioproducts for biomedical applications.
Hepatoprotective and curative activities of aqueous extract of decoction containing 10 Chinese medicinal herbs (HPE-XA-08) were evaluated in Sprague-Dawley albino rats with liver damage induced by thioacetamide (TAA). These activities were assessed by investigating the liver enzymes level and also histopathology investigation. Increases in alkaline phosphatase (ALP) and gamma-glutamyl transferase (GGT) levels were observed in rats with cirrhotic liver. No significant alterations of the liver enzymes were observed following treatment with HPE-XA-08. Histopathology examination of rats treated with HPE-XA-08 at 250 mg/kg body weight, however, exhibited moderate liver protective effects. Reduced extracellular matrix (ECM) proteins within the hepatocytes were noted in comparison to the cirrhotic liver. The curative effects of HPE-XA-08 were observed with marked decrease in the level of ALP (more than 3x) and level of GGT (more than 2x) in cirrhotic rat treated with 600 mg/kg body weight HPE-XA-08 in comparison to cirrhotic rat treated with just water diluent. Reversion of cirrhotic liver to normal liver condition in rats treated with HPE-XA-08 was observed. Results from the present study suggest that HPE-XA-08 treatment assisted in the protection from liver cirrhosis and improved the recovery of cirrhotic liver.
Platelet rich plasma clot- releasate (PRCR) shows significant influence on tissue regeneration in clinical trials. Although, the mechanism of PRCR effect on fibroblast differentiation has been studied on 2D culture system, a detailed investigation is needed to establish the role of PRCR in cell seeded in 3D scaffolds. Therefore, a study was conducted to evaluate the influence of PRCR in fibroblasts (DFB) differentiation and extracellular matrix formation on both 3D and 2D culture systems. Cell viability was measured using MTT assay and DFB differentiation was evaluated by determining the expression levels of nucleostamin and alpha smooth muscle actin (α-SMA), using indirect immunostaining and Western blotting. The expression levels of extracellular matrix genes (collagen-I, collagen-III, fibronectin and laminin) and focal adhesion formation gene (integrin beta-1) were measured using Real-time PCR. The PRCR at 10% showed significant effect on cells viability compared with 5% and 20% in both culture environments. The decrease in the expression levels of nucleostamin and the increase in α-SMA signify the DFB differentiation to myofibroblast-like cells that was prominently greater in 3D compared to 2D culture. In 3D culture systems, the total collage production, expression levels of the extracellular matrix gene and the focal adhesion gene were increased significantly compared to 2D culture. In conclusion, 3D culture environments enhances the proliferative and differentiation effects of PRCR on DFB, thereby potentially increases the efficacy of DFB for future tissue engineering clinical application.