Search results for: regenerative dentistry

Authors: Neelesh Papineni

Abstract:

Aim: Piezosurgery is a relatively new technique for osteotomy and osteoplasty that uses ultrasonic vibration. The conventional method of treating periodontal cases are by conventional surgeries. However, in this advancing field the use of motor-driven instruments is being considered less invasive. Out of these motor-driven instruments, piezo-electric device has been introduced to the field of periodontics and oral implantology. This article discusses about the wide range of application of piezo-electric device in periodontology, its advantages over conventional surgical therapies and other motor-driven instruments. Results: Piezo- electric has shown better results in aspect of osteotomy, osteoplasty, implants, and any procedure which includes conserving the bone. Also piezo-electric does not cause any kind of damage to the surrounding soft tissue and eliminates the risk of bone necrosis which is a risk factor in other motor driven instruments. Conclusion: In this era of modern dentistry , a successful periodontal and implant surgery requires a sound osseous support. This review gives a pictorial representation about the wide range of application of piezo-electric device in periodontology.

Keywords: piezo-electric, osteotomy, osteoplasty, implantology

Procedia PDF Downloads 337

Authors: Gi-Hoon Yang, JongHan Ha, MyungGu Yeo, JaeYoon Lee, SeungHyun Ahn, Hyeongjin Lee, HoJun Jeon, YongBok Kim, Minseong Kim, GeunHyung Kim

Abstract:

Tissue engineering scaffolds must be biocompatible and biodegradable, provide adequate mechanical strength and cell attachment site for proliferation and differentiation. Furthermore, the scaffold morphology (such as pore size, porosity and pore interconnectivity) plays an important role. The electrospinning process has been widely used to fabricate micro/nano-sized fibres. Electrospinning allows for the fabrication of non-woven meshes containing micro- to nano-sized fibers providing high surface-to-volume area for cell attachment. Due to its advantageous characteristics, electrospinning is a useful method for skin, cartilage, bone, and nerve regeneration. In this study, we fabricated PCL scaffolds (SP) consisting of spiral-like struts using 3D melt-plotting system and micro/nanofibers using direct electrospinning writing. By altering the conditions of the conventional melt-plotting method, spiral-like struts were generated. Then, micro/nanofibers were deposited selectively. The control scaffold composed of perpendicular PCL struts was fabricated using the conventional melt-plotting method to compare the cellular activities. The effect on the attached cells (osteoblast-like cells (MG63)) was evaluated depending on the bending instability of the struts. The SP scaffolds showed enhanced biological properties such as initial cell attachment, proliferation and osteogenic differentiation. These results suggest that the SP scaffolds has potential as a bioengineered substitute for soft and hard tissue regeneration.

Keywords: cell attachment, electrospinning, mechanical strength, melt-plotting

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Authors: Esra Turker, Umit Hakan Yildiz, Ahu Arslan Yildiz

Abstract:

The key of regenerative medicine is mimicking natural three dimensional (3D) microenvironment of tissues by utilizing appropriate biomaterials. In this study, a synthetic biodegradable polymer; poly (L-lactide-co-ε-caprolactone) (PLLCL) and a natural polymer; collagen was used to mimic the biochemical structure of the natural extracellular matrix (ECM), and by means of electrospinning technique the real physical structure of ECM has mimicked. PLLCL/Collagen biocomposite scaffolds enables cell attachment, proliferation and nutrient transport through fabrication of micro to nanometer scale nanofibers. Biocomposite materials are commonly preferred due to limitations of physical and biocompatible properties of natural and synthetic materials. Combination of both materials improves the strength, degradation and biocompatibility of scaffold. Literature studies have shown that collagen is mostly solved with heavy chemicals, which is not suitable for cell culturing. To overcome this problem, a new approach has been developed in this study where polyvinylpyrrolidone (PVP) is used as co-electrospinning agent. PVP is preferred due to its water solubility, so PLLCL/collagen biocomposite scaffold can be easily and rapidly produced. Hydrolytic and enzymatic biodegradation as well as mechanical strength of scaffolds were examined in vitro. Cell adhesion, proliferation and cell morphology characterization studies have been performed as well. Further, on-chip drug screening analysis has been performed over 3D tumor models. Overall, the developed biocomposite scaffold was used for 3D tumor model formation and obtained results confirmed that developed model could be used for drug screening studies to predict clinical efficacy of a drug.

Keywords: biomaterials, 3D cell culture, drug screening, electrospinning, lab-on-a-chip, tissue engineering

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Authors: Ezinne M. Igwe

Abstract:

Statements of facts have been made about Nollywood, a segment of the Nigerian film industry that has in recent times become phenomenal due largely to its quantity of production and specific production style. In the face of recent transformations reshaping the industry, matters have been arising which have not been given due academic attention from an industry player perspective. While re-addressing such issues like structure, policy and informality, this study benefits from a new perspective – that of a community member adopting participant observation to research into a familiar culture. With data drawn from an extensive ethnographic study of the industry, this paper examines these matters with an emphasis on structure and the industry’s overall political economy. Drawing from discourses on the new and old Nollywood labels and other current matters arising within the industry such as the MOPICON bill redraft, corporate financing and possibilities of regeneration, this paper examines structure and power struggle within Nollywood. These are championing regenerative processes that bring about formalization, professionalism and the quest for a transnational presence, which have only been superficially evaluated. Focused essentially on Nollywood’s political economy, this study critically analyses the transforming face of an informal industry, the consistent quest for structure, quality and standard, and issues of corporate sponsorship as possible trends of regeneration. It evaluates them as indicators of regeneration, questioning the possibilities of their sustenance in an industry experiencing increased interactions with the formal economy and an influx of young professionals. With findings that make sustained regeneration both certain (due to increased formal economy interaction) and uncertain (due to the dysfunctionality of the society and its political system), it concludes that the transforming face of the industry suggests impending gentrification of the industry.

Keywords: formalisation, MOPICON, Nollywood, structure

Procedia PDF Downloads 246

Authors: Ziyad S. Haidar

Abstract:

Background: Saliva plays a major role in maintaining oral, dental, and general health and well-being; where it normally bathes the oral cavity acting as a clearing agent. This becomes more apparent when the amount and quality of saliva are significantly reduced due to medications, salivary gland neoplasms, disorders such as Sjögren’s syndrome, and especially ionizing radiation therapy for tumors of the head and neck, the 5th most common malignancy worldwide, during which the salivary glands are included within the radiation field/zone. Clinically, patients affected by salivary gland dysfunction often opt to terminate their radiotherapy course prematurely as they become malnourished and experience a significant decrease in their QoL. Accordingly, the formulation of a radio-protection/-prevention modality and development of an alternative Rx to restore damaged salivary gland tissue is eagerly awaited and highly desirable. Objectives: Assess the pre-clinical radio-protective effect and reparative/regenerative potential of layer-by-layer self-assembled lipid-polymer-based core-shell nanocapsules designed and fine-tuned for the sequential (ordered) release of dual cytokines, following a single local administration (direct injection) into a murine sub-mandibular salivary gland model of irradiation. Methods: The formulated core-shell nanocapsules were characterized by physical-chemical-mechanically pre-/post-loading with the drugs, followed by optimizing the pharmaco-kinetic profile. Then, nanosuspensions were administered directly into the salivary glands, 24hrs pre-irradiation (PBS, un-loaded nanocapsules, and individual and combined vehicle-free cytokines were injected into the control glands for an in-depth comparative analysis). External irradiation at an elevated dose of 18Gy was exposed to the head-and-neck region of C57BL/6 mice. Salivary flow rate (un-stimulated) and salivary protein content/excretion were regularly assessed using an enzyme-linked immunosorbent assay (3-month period). Histological and histomorphometric evaluation and apoptosis/proliferation analysis followed by local versus systemic bio-distribution and immuno-histochemical assays were then performed on all harvested major organs (at the distinct experimental end-points). Results: Monodisperse, stable, and cytocompatible nanocapsules capable of maintaining the bioactivity of the encapsulant within the different compartments with the core and shell and with controlled/customizable pharmaco-kinetics, resulted, as is illustrated in the graphical abstract (Figure) below. The experimental animals demonstrated a significant increase in salivary flow rates when compared to the controls. Herein, salivary protein content was comparable to the pre-irradiation (baseline) level. Histomorphometry further confirmed the biocompatibility and localization of the nanocapsules, in vivo, into the site of injection. Acinar cells showed fewer vacuoles and nuclear aberration in the experimental group, while the amount of mucin was higher in controls. Overall, fewer apoptotic activities were detected by a Terminal deoxynucleotidyl Transferase (TdT) dUTP Nick-End Labeling (TUNEL) assay and proliferative rates were similar to the controls, suggesting an interesting reparative and regenerative potential of irradiation-damaged/-dysfunctional salivary glands. The Figure below exemplifies some of these findings. Conclusions: Biocompatible, reproducible, and customizable self-assembling layer-by-layer core-shell delivery system is formulated and presented. Our findings suggest that localized sequential bioactive delivery of dual cytokines (in specific dose and order) can prevent irradiation-induced damage via reducing apoptosis and also has the potential to promote in situ proliferation of salivary gland cells; maxSALIVA is scalable (Good Manufacturing Practice or GMP production for human clinical trials) and patent-pending.

Keywords: cancer, head and neck, oncology, drug development, drug delivery systems, nanotechnology, nanoncology

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Authors: John Scanlon, Antony Raymond, Randeep Aujla, Peter D’Alessandro, Satyen Gohil

Abstract:

Background: The incidence of significant acute chondral injuries with patella dislocation is around 10-15%. It is accepted that chondral procedures should only be performed in the presence of joint stability Methods:Patients were identified from surgeon/hospital logs. Patient demographics, lesion size and location, surgical procedure, patient reported outcome measures, post-operative MR imaging, and complications were recorded. PROMs and patient satisfaction was obtained. Results:20 knees (18 patients) were included. Mean age was 18.6 years (range; 11-39), and the mean follow-up was 16.6 months (range; 2-70). The defect locations were the lateral femoral condyle (9/20; 45%), patella (9/20; 45%), medial femoral condyle (1/20; 5%) and the trochlea (1/20; 5%). The mean defect size was 2.6cm2. Twelve knees were treated with cartilage fixation, 5 with microfracture, and 3 with OATS. At follow up, the overall mean Lysholm score was 77.4 (± 17.1), with no chondral regenerative procedure being statistically superior. There was no difference in Lysholm scores between those patients having acute medial patellofemoral ligament reconstruction versus medial soft tissue plication (p=0.59). Five (25%) knees required re-operation (one arthroscopic arthrolysis; one patella chondroplasty; two removal of loose bodies; one implant adjustment). Overall, 90% responded as being satisfied with surgery. Conclusion: Our aggressive pathway to identify and treat acute cartilage defects with early operative intervention and patella stabilisation has shown high rates of satisfaction and Lysholm scores. The full range of chondral restoration options should be considered by surgeons managing these patients.

Keywords: patella dislocation, chondral restoration, knee, patella stabilisation

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Authors: Odeyemi Ifeoluwayemi, Maha Joushua, Fulani Omoyeni

Abstract:

In the search for sustainability, over time, architectural approaches to design have moved from just nature inspired design to the study of nature’s principles to produce effective designs that solve the issue of sustainability. Nature has established materials, shapes and processes that are effective right from a minor scale to an enormous scale. A branch of human knowledge that studies nature is called biology. Biology helps us to grasp and understand nature. Biomimicry is a new way of viewing and valuing nature, based not on what we can extract from the natural world but on what we can learn from it. Life has sustained on the earth for the last 3.85 billion years, and it is necessary for us to find out how life has been able to stay sustained for that long. The building must teach the society new ecological morals, thus, a better understanding of how nature works can usefully inspire architectural designs to resolve issues that have already been resolved by nature. This will not only help in creating a healthy environment but will also produce positive environmental impacts. Biomimetic Architecture connects and reproduces the ideologies found in nature in order to create built environment which benefit people and other living creatures as well as preserving it for the future. Understanding the bioprocess would lead to the establishment of ecological approaches that serve as a platform for creating a built environment that goes beyond sustaining current settings but also mimic nature’s regenerative ecosystem. This paper aims to explain these design methods under the name of biomimicry and biomimetic architecture by reviewing literature and research works done by examining these approaches classified as forms, processes and ecosystems. It is expected that this research will provide information that would, therefore, lead to the creation of buildings that are eco-friendly and provide greater comfort to the populaces.

Keywords: biomimetic architecture, biomimicry, ecological design, nature

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Authors: Jerome Osentowski

Abstract:

The objective at Central Rocky Mountain Permaculture Institute has been to put in practice a sustainable way of life while growing food, medicine, and providing education. This has been done by applying methods of farming such as agroforestry, forest farming, and perennial polycultures. These methods have been found to be regenerative to the environment through carbon sequestration, soil-building, climate change mitigation, and the provision of food security. After 30 years of implementing carbon farming methods, the results are agro-diversity, self-sustaining systems, and a consistent provision of food and medicine. These results are exhibited through polyculture plantings in an outdoor forest garden spanning roughly an acre containing about 200 varieties of fruits, nuts, nitrogen-fixing trees, and medicinal herbs, and two indoor forest garden greenhouses (one Mediterranean and one Tropical) containing about 50 varieties of tropical fruits, beans, herbaceous plants and more. While the climate zone outside the greenhouse is 6, the tropical forest garden greenhouse retains an indoor climate zone of 11 with near-net-zero energy consumption through the use of a climate battery, allowing the greenhouse to serve as a year-round food producer. The effort to source food from the forest gardens is minimal compared to annual crop production. The findings at Central Rocky Mountain Permaculture Institute conclude that agroecological methods are not only beneficial but necessary in order to revive and regenerate the environment and food security.

Keywords: agroecology, agroforestry, carbon farming, carbon sequestration, climate battery, food security, forest farming, forest garden, greenhouse, near-net-zero, perennial polycultures

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Authors: Qiuyue Peng, Vladimir Zachar

Abstract:

The application of adipose-derived stromal/stem cells (ASCs) in regenerative medicine is gaining more awareness due to their advanced translational potential and abundant source preparations. However, ASC-based translation has been confounded by high subpopulation heterogeneity, causing ambiguity about its precise therapeutic value. Some phenotypes defined by a unique combination of positive and negative surface markers have been found beneficial to the required roles. Therefore, the immunophenotypic repertoires of cultured ASCs and temporal changes of distinct subsets were investigated in this study. ASCs from three donors undergoing cosmetic liposuction were cultured in standard culturing methods, and the co-expression patterns based on the combination of selected markers at passages 1, 4, and 8 were analyzed by multi-chromatic flow cytometry. The results showed that the level of heterogeneity of subpopulations of ASCs became lower by in vitro expansion. After a few passages, most of the CD166⁺/CD274⁺/CD271⁺ based subpopulations converged to CD166 single positive cells. Meanwhile, these CD29⁺CD201⁺ double-positive cells, in combination with CD36/Stro-1 expression or without, feathered only the major epitopes and maintained prevailing throughout the whole process. This study suggested that, upon in vitro expansion, the phenotype repertoire of ASCs redistributed and stabilized in a way that cells co-expressing exclusively the strong markers remained dominant. These preliminary findings provide a general overview of the distribution of heterogeneous subsets residents within human ASCs during expansion in vitro. It is a critical step to fully characterize ASCs before clinical application, although the biological effects of heterogeneous subpopulations still need to be clarified.

Keywords: adipose-derived stromal/stem cells, heterogeneity, immunophenotype, subpopulations

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Authors: Ainur Mukhambetova, Miras Karzhauov, Vyacheslav Ogay

Abstract:

Introduction: Mesenchymal stem cells (MSCs) have the capacity to be differentiated into several cell lineages and are a promising source for cell therapy and tissue engineering. However, currently most MSCs culturing protocols use media supplemented with fetal bovine serum (FBS), which limits their application in clinic due to the possibility of zoonotic infections, contamination and immunological reactions. Consequently, formulating effective serum free culture medium becomes one of the important problems in contemporary cell biotechnology. Objectives: The aim of this study was to define an optimal serum-free medium for culturing of periosteum derived MSCs. Materials and methods: The MSCs were extracted from human periosteum and transferred to the culture flasks pretreated with CELLstart™. Immunophenotypic characterization, proliferation and in vitro differentiation of cells grown on STEM PRO® MSC SFM were compared to the cells cultured in the standard FBS containing media. Chromosome analysis and flow cytometry were also performed. Results: We have shown that cells were grown on STEM PRO® MSC SFM retained all the morphological, immunophenotypic (CD73, CD90, CD105, vimentin and Stro-1) and cell differentiation characteristics specific to MSCs. Chromosome analysis indicated no anomalies in the chromosome structure. Flow cytometry showed a high expression of cell adhesion molecules CD44 (98,8%), CD90 (97,4%), CD105 (99,1%). In addition, we have shown that cell is grown on STEM PRO® MSC SFM have higher proliferation capacity compared to cell expanded on standard FBS containing the medium. Conclusion: We have shown that STEM PRO® MSC SFM is optimal for culturing periosteum derived human MSCs which subsequently can be safely used in cell therapy.

Keywords: cell technologies, periosteum-derived MSCs, regenerative medicine, serum-free medium

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Authors: Reza Fardid, Aliyeh Alipour

Abstract:

Introduction: Coronary heart disease (CHD) is the most common and deadliest diseases. A coronary angiography is an important tool for the diagnosis and treatment of this disease. Because angiography is performed by exposure to ionizing radiation, it can lead to harmful effects. Ionizing radiation induces double-stranded breaks in DNA, which is a potentially life-threatening injury. The purpose of the present study is an investigation of the phosphorylation of histone H2AX in the location of the double-stranded break in Peripheral blood lymphocytes as an indication of Biological effects of radiation on angiography patients. Materials and Methods: This method is based on measurement of the phosphorylation of histone (gamma-H2AX, gH2AX) level on serine 139 after formation of DNA double-strand break. 5 cc of blood from 24 patients with angiography were sampled before and after irradiation. Blood lymphocytes were removed, fixed and were stained with specific ϒH2AX antibodies. Finally, ϒH2AX signal as an indicator of the double-strand break was measured with Flow Cytometry Technique. Results and discussion: In all patients, an increase was observed in the number of breaks in double-stranded DNA after irradiation (20.15 ± 14.18) compared to before exposure (1.52 ± 0.34). Also, the mean of DNA double-strand break was showed a linear correlation with DAP. However, although induction of DNA double-strand breaks associated with radiation dose in patients, the effect of individual factors such as radiosensitivity and regenerative capacity should not be ignored. If in future we can measure DNA damage response in every patient angiography and it will be used as a biomarker patient dose, will look very impressive on the public health level. Conclusion: Using flow cytometry readings which are done automatically, it is possible to detect ϒH2AX in the number of blood cells. Therefore, the use of this technique could play a significant role in monitoring patients.

Keywords: coronary angiography, DSB of DNA, ϒH2AX, ionizing radiation

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Authors: Masaaki Ii, Takashi Saito, Yasuhiko Tabata, Shintaro Nemoto

Abstract:

Background: Clinical trials of autologous adipose-derived stem cell (AdSC) therapy for ischemic heart diseases (IHD) are now on-going. We have investigated the hypothesis that combination of AdSCs and statin, an agent with pleiotropic effects, could augment the therapeutic effect on myocardial infarction (MI). Methods and Results: Human AdSC functions with different doses of simvastatin-conjugated nanoparticle (STNP) uptake were evaluated by in vitro assays. STNP promoted the migration activity without changing the proliferation activity, and also up-regulated growth factors. Next, MI was induced by LAD ligation in nude mice, and the mice were assigned in the following groups 3 days after MI: 1) PBS (control), 2) NP-AdSCs (50000 cells), 3) STNP, and 4) STNP-AdSCs (50000 cells). Cardiac functional recovery assessed by echocardiography was improved at 4 weeks after surgery in STNP-AdSC group. Masson’s trichrome-stained sections revealed that LV fibrosis length was reduced, and the number of TUNEL-positive cardiomyocytes was less in STNP-AdSC group. Surprisingly, a number of de novo endogenous Nkx-2.5/GATA4 positive immature cardiomyocytes as well as massive vascular formation were observed in outer layer of infarcted myocardium despite of a few recruited/retained transfused STNP-AdSCs 4 weeks after MI in STNP-AdSC group. Finally, massive myocardial regeneration was observed 8 weeks after MI. Conclusions: Intravenously injected small number of statin nanoparticle-loaded hAdSCs exhibited a potent therapeutic effect inducing endogenous cardiac tissue regeneration.

Keywords: statin, drug delivery system, stem cells, cardiac regeneration

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Authors: Ekaterina Zubkova, Irina Beloglazova, Iurii Stafeev, Konsyantin Dergilev, Yelena Parfyonova, Mikhail Menshikov

Abstract:

Mesenchymal stromal cells from adipose tissue (MSC) currently are widely used in regenerative medicine to restore the function of damaged tissues, but that is significantly hampered by their heterogeneity. One of the modern approaches to overcoming this obstacle is the polarization of cell subpopulations into a specific phenotype under the influence of cytokines and other factors that activate receptors and signal transmission to cells. We polarized MSC with factors affecting the inflammatory signaling and functional properties of cells, followed by verification of their expression profile and ability to affect the polarization of macrophages. RT-PCR evaluation showed that cells treated with LPS, interleukin-17, tumor necrosis factor α (TNF α), primarily express pro-inflammatory factors and cytokines, and after treatment with polyninosin polycytidic acid and interleukin-4 (IL4) anti-inflammatory factors and some proinflammatory factors. MSC polarized with pro-inflammatory cytokines showed a more robust pro-angiogenic effect in fibrin gel bead 3D angiogenesis assay. Further, we evaluated the possibility of paracrine effects of MSCs on the polarization of intact macrophages. Polarization efficiency was assesed by expression of M1/M2 phenotype markers CD80 and CD206. We showed that conditioned media from MSC preincubated in the presence of IL-4 cause an increase in CD206 expression similar to that observed in M2 macrophages. Conditioned media from MSC polarized in the presence of LPS or TNF-α increased the expression of CD80 antigen in macrophages, similar to that observed in M1 macrophages. In other cases, a pronounced paracrine effect of MSC on the polarization of macrophages was not detected. Thus, our study showed that the polarization of MSC along the pro-inflammatory or anti-inflammatory pathway allows us to obtain cell subpopulations that have a multidirectional modulating effect on the polarization of macrophages. (RFBR grants 20-015-00405 and 18-015-00398.)

Keywords: angiogenesis, cytokines, mesenchymal, polarization, inflammation

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Authors: Nety Trisnawaty, Mirna Febriani

Abstract:

The use of stainless steel wires in the field of dentistry is widely used, especially for orthodontic and prosthodontic treatment using stainless steel wire. The oral cavity is the ideal environment for corrosion, which can be caused by saliva. Prevention of corrosion on stainless steel wires can be done by using an organic or non-organic corrosion inhibitor. One of the organic inhibitors that can be used to prevent corrosion is black tea leaves extracts. To explain the comparison of chromium ions release for stainlees steel between artificial saliva and black tea leaves extracts. In this research we used artificial saliva, black tea leaves extracts, stainless steel wire and using Atomic Absorption Spectrophometric testing machine. The samples were soaked for 1, 3, 7 and 14 days in the artificial saliva and black tea leaves extracts. The results showed the difference of chromium ion release soaked in artificial saliva and black tea leaves extracts on days 1, 3, 7 and 14. Statistically, calculation with independent T-test with p < 0,05 showed a significant difference. The longer the duration of days, the more ion chromium were released. The conclusion of this study shows that black tea leaves extracts can inhibit the corrosion rate of stainless steel wires.

Keywords: chromium ion, stainless steel, artificial saliva, black tea leaves extracts

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Authors: Vanessa Cho, Janet Hsiao, Nigel King, Robert Anthonappa

Abstract:

Background: The personal protective equipment (PPE) requirements for health care workers (HCWs) have changed significantly during the COVID-19 pandemic. Aim: To ascertain, using eye-tracking technology, what children notice the most when seeing HCWs in various PPE. Design: A Tobii nano pro-eye-tracking camera tracked 156 children's visual attention while they viewed photographs of HCWs in various PPEs. Eye Movement analysis with Hidden Markov Models (EMHMM) was employed to analyse 624 recordings using two approaches, namely (i) data-driven where children's fixation determined the regions of interest (ROIs), and (ii) fixed ROIs where the investigators predefined the ROIs. Results: Two significant eye movement patterns, namely distributed(85.2%) and selective(14.7%), were identified(P<0.05). Most children fixated primarily on the face regardless of the different PPEs. Children fixated equally on all PPE images in the distributed pattern, while a strong preference for unmasked faces was evident in the selective pattern (P<0.01). Conclusion: Children as young as 2.5 years used a top-down visual search behaviour and demonstrated their face processing ability. Most children did not show a strong visual preference for a specific PPE, while a minority preferred PPE with distinct facial features, namely without masks and loupes.

Keywords: COVID-19, PPE, dentistry, pediatric

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Authors: Ziyad S. Haidar

Abstract:

Background: Saliva plays a major role in maintaining oral and dental health (consequently, general health and well-being). Where it normally bathes the oral cavity and acts as a clearing agent. This becomes more apparent when the amount and quality of salivare significantly reduced due to medications, salivary gland neoplasms, disorders such as Sjögren’s syndrome, and especially ionizing radiation therapy for tumors of the head and neck, the fifth most common malignancy worldwide, during which the salivary glands are included within the radiation field or zone. Clinically, patients affected by salivary gland dysfunction often opt to terminate their radiotherapy course prematurely because they become malnourished and experience a significant decrease in their quality of life. Accordingly, the development of an alternative treatment to restore or regenerate damaged salivary gland tissue is eagerly awaited. Likewise, the formulation of a radioprotection modality and early damage prevention strategy is also highly desirable. Objectives: To assess the pre-clinical radio-protective effect as well as the reparative/regenerative potential of layer-by-layer self-assembled lipid-polymer-based core-shell nanocapsules designed and fine-tuned in this experimental work for the sequential (ordered) release of dual cytokines, following a single local administration (direct injection) into a murine sub-mandibular salivary gland model of irradiation. Methods: The formulated core-shell nanocapsules were characterized by physical-chemical-mechanically pre-/post-loading with the drugs (in solution and powder formats), followed by optimizing the pharmaco-kinetic profile. Then, nanosuspensions were administered directly into the salivary glands, 24hrs pre-irradiation (PBS, un-loaded nanocapsules, and individual and combined vehicle-free cytokines were injected into the control glands for an in-depth comparative analysis). External irradiation at an elevated dose of 18Gy (revised from our previous 15Gy model) was exposed to the head-and-neck region of C57BL/6 mice. Salivary flow rate (un-stimulated) and salivary protein content/excretion were regularly assessed using an enzyme-linked immunosorbent assay (3-month period). Histological and histomorphometric evaluation and apoptosis/proliferation analysis followed by local versus systemic bio-distribution and immuno-histochemical assays were then performed on all harvested major organs (at the distinct experimental end-points). Results: Monodisperse, stable, and cytocompatible nanocapsules capable of maintaining the bioactivity of the encapsulant within the different compartments with the core and shell and with controlled/customizable pharmaco-kinetics, resulted, as is illustrated in the graphical abstract (Figure) below. The experimental animals demonstrated a significant increase in salivary flow rates when compared to the controls. Herein, salivary protein content was comparable to the pre-irradiation (baseline) level. Histomorphometry further confirmed the biocompatibility and localization of the nanocapsules, in vivo, into the site of injection. Acinar cells showed fewer vacuoles and nuclear aberration in the experimental group, while the amount of mucin was higher in controls. Overall, fewer apoptotic activities were detected by a Terminal deoxynucleotidyl Transferase (TdT) dUTP Nick-End Labeling (TUNEL) assay and proliferative rates were similar to the controls, suggesting an interesting reparative and regenerative potential of irradiation-damaged/-dysfunctional salivary glands. The Figure below exemplifies some of these findings. Conclusions: Biocompatible, reproducible, and customizable self-assembling layer-by-layer core-shell delivery system is formulated and presented. Our findings suggest that localized sequential bioactive delivery of dual cytokines (in specific dose and order) can prevent irradiation-induced damage via reducing apoptosis and also has the potential to promote in situ proliferation of salivary gland cells; maxSALIVA is scalable (Good Manufacturing Practice or GMP production for human clinical trials) and patent-pending.

Keywords: saliva, head and neck cancer, nanotechnology, controlled drug delivery, xerostomia, mucositis, biopolymers, innovation

Procedia PDF Downloads 56

Authors: Paria Koraei, Meysam Ahmadzadeh, Reza Shahriarirad, Keivan Ranjbar, Farzane Farahmand

Abstract:

The rapid spread of the coronavirus disease 2019 (COVID-19) has enlisted dental offices in high-risk places. A cross-sectional survey was carried out among the general population aged 17 and above in Shiraz, Fars, Iran. The patients' demographic features were recorded and analyzed against questions regarding fear, anxiety, and attitude toward dental appointments. The Dental Anxiety Scale (DAS) questionnaire was used for the evaluation of anxiety levels. Among a total of 760 participants, the average age was 32.33±11.27 years, while 242 (31.8%) were male and 357 (47%) were single. Based on the anxiety scores, 336 (44.2%) participants had mild anxiety, 215 (28.3%) had moderate anxiety, 74 (9.7%) had high, and 135 (17.8%) had severe anxiety. The majority of participants used masks (93.8%) and disinfectants (79.9%) during dental visits, and the most emphasized policies during dental visits were using disposable face masks by the personnel (83.6%) and controlling the number of visitors in the office (81.8%). Only 226 (29.7%) reported that they hadn’t postponed their dental visits. Our findings demonstrated that anxiety in people increases toward dental treatments during the COVID-19 pandemic, resulting in decreased dental appointments. Dentally anxious people were more likely to be female and married couples. Being female and older age are possible risk factors for developing a fear of dental procedures.

Keywords: Covid-19, anxiety, fear, dentistry, dental care

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Authors: Soumya S., Manju Nidagodu Jayakumar, Vellore Kannan Gopinath

Abstract:

Dental pulp inflammation resulting from dental caries often leads to a pathologic condition known as irreversible pulpitis and the currently managed by root canal treatment. Extirpation of the entire pulp tissue is done during this procedure, and the canal space is filled with synthetic materials. Recent studies in the stem cell biology state that some portion of the irreversibly inflamed pulp tissue could be viable with progenitor cells, having the properties similar to that of Mesenchymal stem cells. Hence, we aim to isolate Dental Pulp Stem Cells (DPSCs) from patients diagnosed with severe irreversible pulpitis and characterize the cells for the MSC specific markers. The pulp tissue was collected from the dental clinic and subjected to collagenase/dispase digestion. The isolated cells were expanded in culture, and the phenotypic characterization was done using flow cytometry. MSC specific markers such as CD-90, CD-73, and CD-105 were analysed along with negative markers such as CD-14 and CD-45. The isolated cells expressed positive expression for CD markers with CD90 and CD105 ( > 95%) and CD73 (19%). The cells did not express the negative markers CD-14 and CD-45. The commercially available DPSCs from vital extracted teeth, preferably molar/wisdom teeth with large pulp cavity or incomplete root growth in young patients (aged 15-30 years) showed more than 90% expression for all the CD markers such as CD-90, 73 and 105, whereas negative for CD-14 and CD-45. The DPSCs isolated from inflamed pulp tissue showed a less expression for CD-73 compared to the commercially available DPSCs whereas, as the other two markers were found to show similar percentage of positive expression. This could be attributed to the fact that the pulp population is very heterogeneous and we used the pooled tissue from different patients. Hence the phenotypic characterization and comparison with the commercially available DPSCs proved that the inflamed pulp tissue is a good source of MSC like cells which can be utilized further for regenerative application.

Keywords: collagenase/dispase, dental pulp stem cells, flow cytometry, irreversible pulpitis

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Authors: Matej Buzgo, Erico Himawan, Ksenija JašIna, Aiva Simaite

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Electrospinning is a versatile and efficient technology for producing nanofibers for biomedical applications. One of the most common polymers used for the preparation of nanofibers for regenerative medicine and drug delivery applications is polycaprolactone (PCL). PCL is a biocompatible and bioabsorbable material that can be used to stimulate the regeneration of various tissues. It is also a common material used for the development of drug delivery systems by blending the polymer with small active molecules. However, for many drug delivery applications, e.g. cancer immunotherapy, PCL biodegradation rate that may exceed 9 months is too long, and faster nanofiber dissolution is needed. In this paper, we investigate the dissolution and small molecule release rates of PCL blends with two hydrophilic polymers: polyethylene oxide (PEO) or polyvinylpyrrolidone (PVP). We show that adding hydrophilic polymer to the PCL reduces the water contact angle, increases the dissolution rate, and strengthens the interactions between the hydrophilic drug and polymer matrix that further sustain its release. Finally using this method, we were also able to increase the nanofiber degradation rate when PCL-PEO and PCL-PVP were used as a shell in the electrospun core-shell nanofibers and spread up the release of active proteins from their core. Electrospinning can be used for the preparation of the core-shell nanofibers, where active ingredients are encapsulated in the core and their release rate is regulated by the shell. However, such fibers are usually prepared by coaxial electrospinning that is an extremely low-throughput technique. An alternative is emulsion electrospinning that could be upscaled using needleless blades. In this work, we investigate the possibility of using emulsion electrospinning for encapsulation and sustained release of the growth factors for the development of the organotypic skin models. The core-shell nanofibers were prepared using the optimized formulation and the release rate of proteins from the fibers was investigated for 2 weeks – typical cell culture conditions.

Keywords: electrospinning, polycaprolactone (PCL), polyethylene oxide (PEO), polyvinylpyrrolidone (PVP)

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Authors: Sat Sharma, Antonella Conflitti, Hilary Reiter

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Introduction: The study's purpose was to assess the efficacy of the oral appliance device O2Vent Optima + ExVent as compared to Optima in the treatment of OSA. Methods: A prospective, open-label study conducted at 3 sites included subjects with mild to moderate OSA (AHI ≥ 5 and ≤ 30). Screening Phase: A diagnostic in-lab PSG study was performed to confirm a diagnosis of mild to moderate OSA. Treatment I: Subjects used O2Vent Optima for 6 weeks and underwent an in-lab PSG sleep night while using the O2Vent Optima. Treatment II: Subjects used O2Vent Optima + ExVent for 6 weeks and underwent an in-lab PSG sleep night while using the O2Vent Optima + ExVent Primary Effectiveness Measure: Change in AHI between baseline vs. O2Vent Optima MAD vs. O2Vent Optima + ExVent. Results: Treatment with Optima, Optima + ExVent reduced AHI from 22.5±6.4/hr to 12.6±4.5/hr to 5.9±2.7 (p< 0.005 baseline vs. Optima and Optima + ExVent; p<0.05 Optima MAD vs. Optima + ExVent). The average reduction in AHI with Optima was 43%, and with Optima + ExVent was 72%. The lowest oxygen during sleep increased from 84.6±2.7% to 88.6±2.9% to 91.6±3.2% (p< 0.005 baseline vs. Optima and Optima + ExVent; p<0.05 Optima vs. Optima + ExVent). During the trial, patients on treatment with Optima and Optima + ExVent demonstrated no excessive adverse events or device malfunction. Conclusion: Treatment with O2Vent Optima and O2Vent Optima + ExVent significantly improved OSA compared to the baseline. An even greater benefit was observed with the addition of ExVent to the Optima in mild to moderate OSA.

Keywords: oral appliance, O2Vent, sleep dentistry, sleep apnea

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Authors: Ankush M. Dewle, Suditi Bhattacharya, Prachi R. Abhang, Savita Datar, Ajay J. Jog, Rupesh K. Srivastava, Geetanjali Tomar

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Objectives: The aim of this study was to investigate the quality of mesenchymal stem cells (MSCs) obtained from ageing gingival tissues, in order to suggest their potential role in autologous stem cell therapy for old individuals. Methods: MSCs were isolated from gingival tissues of young (18-45 years) and old (above 45 years) donors by enzymatic digestion. MSCs were analysed for cfu-f, surface marker expression by flow-cytometry and multilineage differentiation potential. The angiogenic potential was compared in a chick embryo yolk sac membrane model. The aging and differentiation markers including SA-β-galactosidase and p21 respectively were analysed by staining and flow-cytometry analysis. Additionally, osteogenic markers such as glucocorticoid receptor (GR), vitamin D receptor (VDR) were measured by flow-cytometry and RT-qPCR was performed for quantification of osteogenic gene expression. Alizarin Red S and alkaline phosphatase (ALP) activity were also quantitated. Results: Gingival MSCs (GMSCs) from both the age groups were similar in their morphology and displayed cfu-f. They had similar expression of MSC surface markers and p21, comparable rate of proliferation and differentiated to all the four lineages. GMSCs from young donors had a higher adipogenic differentiation potential as compared to the old GMSCs. Moreover, these cells did not display a significant difference in ALP activity probably due to comparable expression of GR, VDR, and osteogenic genes. Conclusions: Ageing of GMSCs occurs at a much slower rate than stem cells from other sources. Thus we suggest GMSCs as an excellent candidate for autologous stem cell therapy in degenerative diseases of elderly individuals. Clinical Significance: GMSCs could help overcome the setbacks in clinical implementation of autologous stem cell therapy for regenerative medicine in all age group of patient.

Keywords: bone regeneration, cell therapy, senescence, stem cell

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Authors: Salma Ainine, Colin Ritchie, Tracey McFee

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Background: The societal impact of antibiotic resistance is a major public health concern. The increase in the incidence of resistant bacteria can ultimately be fatal. Objective: To analyse the appropriateness of antibiotic prescribing in Dundee Dental Hospital, ultimately improving the safety and quality of patient care. Methods: Two examiners independently cross-checked approximately fifty consecutive prescriptions, and corresponding patient case notes, for three data collection cycles between August 2014–September 2015. The Scottish Dental Clinical Effectiveness Program (SDCEP) Drug Prescribing for Dentistry guidelines was the standard utilised. The criteria: clinical justification, regime justification, and review arrangements was measured, and compared to the standard. Results: Cycle one revealed 42% of antibiotic prescriptions were appropriate. Interventions included: multiple staff meetings, an introduction of a checklist attached to the prescription pack, and production of patient leaflets explaining indications for antibiotics. Cycle two and three revealed 44%, and 30% compliance, respectively. Conclusion: The results of the audit have yet to meet target standards set out in prescribing guidelines. However, steps are being taken and change has occurred on a cultural level.

Keywords: antibiotic resistance, antibiotic stewardship, dental infection, hygiene standards

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Authors: Kerry Mackereth

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Kimberle Crenshaw’s theory of intersectionality has become immensely influential in the fields of women’s and gender studies. However, intersectionality’s widespread use among feminist scholars and activists has been accompanied by critiques of its reliance upon subject categorization. These critiques are of particular import when connected to Wendy Brown’s characterization of identity politics as static 'wounded attachments'. Together, these critiques show how the gridlock model proposed by intersectionality’s primary metaphor, the traffic accident at the intersection, is useful for identifying discrimination but not for remembering historical injustices or imagining feminist and anti-racist resistance. Through the lens of New Zealand Maori film, focusing upon Once Were Warriors (1994) and Whale Rider (2002), this article examines how wounded histories need not be passively reproduced by contemporaneously oppressed groups. Instead, the metaphor of the traffic intersection should be complemented by the metaphor of the wound. Against Brown’s characterization of wounded attachments as negative, static identities, Gloria Anzaldua’s account of the borderland between the United States and Mexico as “una herida abierta”, an open wound, offers an alternative reading of the wound. Through Anzaldua’s and Hortense Spillers’ political thought, the wound is reconceptualized as not only a site of suffering but also as a regenerative space. The coexistence of deterioration and regeneration at the site of the wound underpins the narrative arc of both Once Were Warriors and Whale Rider. In both films, the respective child protagonists attempt to reconcile the pain of wounded histories with the imagination of cultural regeneration. The metaphor of the wound thus serves as an alternative theoretical resource for mapping experiences of oppression, one that enriches feminist theory by balancing the remembrance of historical grievance with the forging of hopeful political projects.

Keywords: gender theory, historical grievance, intersectionality, New Zealand film, postcolonialism

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Authors: Wei-Wen Hu, Yong-Zhi Zhong

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Depositing conductive polypyrrole (PPy) onto elastic polydimethylsiloxane (PDMS) substrate can obtain a highly stretchable conductive film, which can be used to construct a bioreactor to cyclically stretch and electrically stimulate surface cells. However, how to completely harvest these stimulated muscle tissue to repair damaged muscle is a challenge. To address this concern, N-isopropylacrylamide (NIPAAm), a monomer of temperature-sensitive polymer, was added during the polymerization of pyrrole on PDMS so that the resulting P(Py-co-NIPAAm)/PDMS should own both conductivity and thermo-sensitivity. Therefore, cells after stimulation can be completely harvested as cell sheets by reducing temperature. Mouse skeletal myoblast, C2C12 cells, were applied to examine our hypothesis. In electrical stimulation, C2C12 cells on P(Py-co-NIPAAm)/PDMS demonstrated the best myo-differentiation under the electric field of 1 V/cm. Regarding cyclic stretching, the strain equal to or higher than 9% can highly align C2C12 perpendicular to the stretching direction. The Western blotting experiments demonstrated that the cell sheets harvested by cooling reserved more extracellular matrix (ECM) than cells collected by the traditional trypsin digestion method. Immunostaining of myosin heavy chain protein (MHC) indicated that both mechanical and electrical stimuli effectively increased the number of myotubes and the differentiation ratio, and the myotubes can be aligned by cyclic stretching. Stimulated cell sheets can be harvested by cooling, and the alignment of myotubes was still maintained. These results suggested that the deposition of P(Py-co-NIPAAm) on PDMS can be applied to harvest intact cell sheets after cyclic stretching and electrical stimulation, which increased the feasibility of bioreactor for the application of tissue engineering and regenerative medicine.

Keywords: bioreactor, cell sheet, conductive polymer, cyclic stretching, electrical stimulation, muscle tissue engineering, myogenesis, thermosensitive hydrophobicity

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Authors: Thirupathi Thippani, Kothandaraman Ramanujam

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Current research on energy storage has been paid to metal-air batteries, because of attractive alternate energy source for the future. Metal – air batteries have the probability to significantly increase the power density, decrease the cost of energy storage and also used for a long time due to its high energy density, low-level pollution, light weight. The performance of these batteries mostly restricted by the slow kinetics of the oxygen reduction reaction (ORR) and oxygen evolution reaction (OER) on cathode during battery discharge and charge. The ORR and OER are conventionally carried out with precious metals (such as Pt) and metal oxides (such as RuO₂ and IrO₂) as catalysts separately. However, these metal-based catalysts are regularly undergoing some difficulties, including high cost, low selectivity, poor stability and unfavorable to environmental effects. So, in order to develop the active, stable, corrosion resistance and inexpensive bi-functional catalyst material is mandatory for the commercialization of zinc-air rechargeable battery technology. We have attempted and synthesized non-precious metal (NPM) catalysts comprising cobalt and N-doped multiwalled carbon nanotubes (N-MWCNTs-Co) were synthesized by the solid-state pyrolysis (SSP) of melamine with Co₃O₄. N-MWCNTs-Co acts as an excellent electrocatalyst for both the oxygen reduction reaction (ORR) and the oxygen evolution reaction (OER), and hence can be used in secondary metal-air batteries and in unitized regenerative fuel cells. It is important to study the OER and ORR at high concentrations of KOH as most of the metal-air batteries employ KOH concentrations > 4M. In the first 16 cycles of the zinc-air battery while using N-MWCNTs-Co, 20 wt.% Pt/C or 20 wt.% IrO₂/C as air electrodes. In the ORR regime (the discharge profile of the zinc-air battery), the cell voltage exhibited by N-MWCNTs-Co was 44 and 83 mV higher (based on 5th cycle) in comparison to of 20 wt.% Pt/C and 20 wt.% IrO₂/C respectively. To demonstrate this promise, a zinc-air battery was assembled and tested at a current density of 0.5 Ag⁻¹ for charge-discharge 100 cycles.

Keywords: oxygen reduction reaction (ORR), oxygen evolution reaction(OER), non-platinum, zinc air battery

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Authors: Ben Nadler

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Cell adhesion plays a vital role in many cell activities. The motivation to model cell adhesion is to study important biological processes, such as cell spreading, cell aggregation, tissue formation, and cell adhesion, which are very challenging to study by experimental methods alone. This study provides important insight into cell adhesion, which can lead to improve regenerative medicine and tissue formation techniques. In this presentation the biological cells adhesion is mediated by receptors–ligands binding and the diffusivity of the receptor on the cell membrane surface. The ability of receptors to diffuse on the cell membrane surface yields a very unique and complicated adhesion mechanism, which is exclusive to cells. The phospholipid bilayer, which is the main component in the cell membrane, shows fluid-like behavior associated with the molecules’ diffusivity. The biological cell is modeled as a fluid-like membrane with negligible bending stiffness enclosing the cytoplasm fluid. The in-plane mechanical behavior of the cell membrane is assumed to depend only on the area change, which is motivated by the fluidity of the phospholipid bilayer. In addition, the presence of receptors influences on the local mechanical properties of the cell membrane is accounted for by including stress-free area change, which depends on the receptor density. Based on the physical properties of the receptors and ligands the attraction between the receptors and ligands is modeled as a charged-nonpolar which is a noncovalent interaction. Such interaction is a short-range type, which decays fast with distance. The mobility of the receptor on the cell membrane is modeled using the diffusion equation and Fick’s law is used to model the receptor–receptor interactions. The resultant interaction force, which includes receptor–ligand and receptor–receptor interaction, is decomposed into tangential part, which governs the receptor diffusion, and normal part, which governs the cell deformation and adhesion. The formulation of the governing equations and numerical simulations will be presented. Analysis of the adhesion characteristic and properties are discussed. The roles of various thermomechanical properties of the cell, receptors and ligands on the cell adhesion are investigated.

Keywords: cell adhesion, cell membrane, receptor-ligand interaction, receptor diffusion

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Authors: Vivek Thorat, Suhasini Desai

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The hybrid electric vehicle is widely accepted as a promising short to mid-term technical solution due to noticeably improved efficiency and low emissions at competitive costs. Retro fitment of hybrid components into a conventional vehicle for achieving better performance is the best solution so far. But retro fitment includes major modifications into a conventional vehicle with a high cost. This paper focuses on the development of a P3x hybrid prototype with rear wheel drive parallel hybrid electric Light Commercial Vehicle (LCV) with minimum and low-cost modifications. This diesel Hybrid LCV is different from another hybrid with regard to the powertrain. The additional powertrain consists of continuous contact helical gear pair followed by chain and sprocket as a coupler for traction motor. Vehicle powertrain which is designed for the intended high-speed application. This work focuses on targeting of design, development, and packaging of this unique parallel diesel-electric vehicle which is based on multimode hybrid advantages. To demonstrate the practical applicability of this transmission with P3x hybrid configuration, one concept prototype vehicle has been build integrating the transmission. The hybrid system makes it easy to retrofit existing vehicle because the changes required into the vehicle chassis are a minimum. The additional system is designed for mainly five modes of operations which are engine only mode, electric-only mode, hybrid power mode, engine charging battery mode and regenerative braking mode. Its driving performance, fuel economy and emissions are measured and results are analyzed over a given drive cycle. Finally, the output results which are achieved by the first vehicle prototype during experimental testing is carried out on a chassis dynamometer using MIDC driving cycle. The results showed that the prototype hybrid vehicle is about 27% faster than the equivalent conventional vehicle. The fuel economy is increased by 20-25% approximately compared to the conventional powertrain.

Keywords: P3x configuration, LCV, hybrid electric vehicle, ROMAX, transmission

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Authors: Aulina R. Rahmi, Farid Yuristiawan, Annisa Ibifadillah, Ummu H. Amri, Hidayati Gunawan

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Glass ionomer cement is one of the filling material that often used on the field of dentistry because it is relatively less expensive and mostly available. Restoration materials could undergo changes in their clinical properties such as changes in roughness of the restoration`s surface. An increase of surface roughness accelerates bacterial colonization and plaque maturation. In the oral cavity, GIC was exposed to various substances, such as toothpaste, an oral care product used during toothbrushing. One of the popular toothpaste is whitening toothpaste. Abrasive and chemical agents such as hydrogen peroxide in whitening toothpaste could increase the surface roughness of restorative materials. Objective: To determine the differences on the surface roughness of glass ionomer cement that was brushed with whitening and conventional toothpaste. Method: This study was done using experimental laboratory method with pre and post test design. There were 36 samples which were divided into 2 groups. The first group was brushed with whitening toothpaste and the second group was brushed with conventional toothpaste, each for 2 minutes. Surface roughness value of the specimens was measured by using Roughness Tester test. Result: The data was analyzed by using independent t-test and the result of this study showed there was a significant difference between the surface of glass ionomer cement which was brushed with whitening and conventional toothpaste (p=0,000). Conclusion: Glass ionomer cement that was brushed with whitening toothpaste produced more roughness than conventional toothpaste.

Keywords: glass ionomer cement, surface roughness, toothpaste, roughness tester

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Authors: Aliya Sekenova, Vyacheslav Ogay

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The derivation of human induced pluripotent stem cells (iPSCs) from somatic cells by direct reprogramming opens wide perspectives in the regenerative medicine. It means the possibility to develop the personal and, consequently, any immunologically compatible cells for applications in cell-based therapy. The purpose of our study was to develop the technology for the production of NK cells from T cell-derived induced pluripotent stem cells (TiPSCs) for subsequent application in adoptive cancer immunotherapy. Methods: In this study iPSCs were derived from peripheral blood T cells using Sendai virus vectors expressing Oct4, Sox2, Klf4 and c-Myc. Pluripotent characteristics of TiPSCs were examined and confirmed with alkaline phosphatase staining, immunocytochemistry and RT-PCR analysis. For NK cell differentiation, embryoid bodies (EB) formed from (TiPSCs) were cultured in xenogeneic serum-free medium containing human serum, IL-3, IL-7, IL-15, SCF, FLT3L without using M210-B4 and AFT-024 stromal feeder cells. After differentiation, NK cells were characterized with immunofluorescence analysis, flow cytometry and cytotoxicity assay. Results: Here, we for the first time demonstrate that TiPSCs can effectively differentiate into functionally active NK cells without M210-B4 and AFT-024 xenogeneic stroma cells. Immunofluorescence and flow cytometry analysis showed that EB-derived cells can differentiate into a homogeneous population of NK cell expressing high levels of CD56, CD45 and CD16 specific markers. Moreover, these cells significantly express killing activation receptors such as NKp44 and NKp46. In the comparative analysis, we observed that NK cells derived using feeder-free culture system have more high killing activity against K-562 tumor cells, than NK cells derived by feeder-dependent method. Thus, we think that our obtained data will be useful for the development of large-scale production of NK cells for translation into cancer immunotherapy.

Keywords: induced pluripotent stem cells, NK cells, T cells, cell diffentiation, feeder cell-free culture system

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Authors: Roxane A. Legaie, Kjiana E. Schwab, Caroline E. Gargett

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Studies looking at the changes in gene expression from RNAseq data often make use of linear models. It is also common practice to focus on a subset of data for a comparison of interest, leaving aside the samples not involved in this particular comparison. This work shows the importance of including all observations in the modeling process to better estimate variance parameters, even when the samples included are not directly used in the comparison under test. The human endometrium is a dynamic tissue, which undergoes cycles of growth and regression with each menstrual cycle. The mesenchymal stem cells (MSCs) present in the endometrium are likely responsible for this remarkable regenerative capacity. However recent studies suggest that MSCs also plays a role in the pathogenesis of endometriosis, one of the most common medical conditions affecting the lower abdomen in women in which the endometrial tissue grows outside the womb. In this study we compared gene expression profiles between MSCs and non-stem cell counterparts (‘non-MSC’) obtained from women with (‘E’) or without (‘noE’) endometriosis from RNAseq. Raw read counts were used for differential expression analysis using a linear model with the limma-voom R package, including either all samples in the study or only the samples belonging to the subset of interest (e.g. for the comparison ‘E vs noE in MSC cells’, including only MSC samples from E and noE patients but not the non-MSC ones). Using the full dataset we identified about 100 differentially expressed (DE) genes between E and noE samples in MSC samples (adj.p-val < 0.05 and |logFC|>1) while only 9 DE genes were identified when using only the subset of data (MSC samples only). Important genes known to be involved in endometriosis such as KLF9 and RND3 were missed in the latter case. When looking at the MSC vs non-MSC cells comparison, the linear model including all samples identified 260 genes for noE samples (including the stem cell marker SUSD2) while the subset analysis did not identify any DE genes. When looking at E samples, 12 genes were identified with the first approach and only 1 with the subset approach. Although the stem cell marker RGS5 was found in both cases, the subset test missed important genes involved in stem cell differentiation such as NOTCH3 and other potentially related genes to be used for further investigation and pathway analysis.

Keywords: differential expression, endometriosis, linear model, RNAseq

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