EXPLORING THE ROLE AND IMPACT OF MICROSCALE PHENOMENA ON ELECTRODE, MICRODEVICE, AND CELLULAR FUNCTION

Microfluidic technologies enable the development of portable devices to perform multiple high-resolution unit operations with small sample and reagent volumes, low fabrication cost, facile operation, and quick response times. Microfluidic platforms are expected to effectively interpret both wanted and unwanted phenomena; however, a comprehensive evaluation of the unwanted phenomena has not been appropriately investigated in the literature. This work explored an attenuative evaluation of unwanted phenomena, also called here as secondary phenomena, in a unique approach. Upon electric field utilization within microfluidic devices, electrode miniaturization improves device sensitivity. However, electrodes in contact with medium solution can yield byproducts that can change medium properties such as pH as well as bulk ion concentration and eventually target cell viability. While electrode byproducts are sometimes beneficial; but, this is not always the case. Two strategies were employed to protect cells from the electrode byproducts: (i) coating the electrodes with hafnium oxide (HfO2), and (ii) stabilization of the cell membrane using a low concentration of Triton X-100 surfactant. Our results showed that both strategies are a plausible way to selectively isolate cell and reduce the risk of contamination from electrode byproducts. The design of a medium solution is also critical to minimize unwanted cell-medium interaction. Surfactants are frequently added to cell-medium solutions to improve sensitivity and reproducibility without disrupting protein composition of cell membranes or cell viability. In non-electrokinetic systems, surfactants have been shown to reduce interfacial tensions and prevent analyte sticking. However, the impacts of surfactant interactions with cell membranes have not previously been explored in electrokinetic systems. This work indicated the dynamic surfactant interactions with cell membranes which altered the cell membrane integrity. It is important that the effects of the chemical interactions between cells to be fully explored and to be separately attributed to reported cellular responses to accurate catalog properties and engineer reliable microfluidic electrokinetic devices. Finally, a comprehensive level of understanding led us to utilize dielectrophoresis in its full capacity as a tool to monitor the state and progression of virus infection as well as anti-viral activities of regenerative compound. Glycine was utilized as potential antiviral compounds to reduce porcine parvovirus (PPV) infection in porcine kidney (PK-13) cells. Our results demonstrate that the glycine altered the virus-host interactions during virus assembly. Thus, elucidating the mechanisms of these novel antiviral compounds is crucial to their development as potential therapeutic drugs

Bone mineral density and bone metabolism in patients with Duchenne muscular dystrophy

 ABSTRACTObjective: Poor bone health with related morbidity is a major problem with Duchene Muscular Dystrophy (DMD). Decreased mobility and long-term corticosteroid therapy are involved in poor bone health in DMD. This study investigates bone mineral density and bone metabolism in 30 steroid treated DMD patients and also comparison of mentioned factors between ambulated and non-ambulated patients.Materials & Methods: In this cross-sectional study 30 boys (21 patients ambulate and 9 non-ambulate) with documented DMD, according to genetic analysis, were enrolled. Demographic characteristics, neurologic exam findings, muscle function score, corticosteroid dose and duration and food frequency questionnaire   were recorded. Bone mineral density was measured with dual- energy X-ray absorptiometry (DEXA) on lumbar spine and left proximal femur. Serum 25-hydroxyvitamin D, calcium, phosphorus and parathyroid hormone (PTH) levels were measured.Results: Osteoporosis was found in 86.7% patients. Mean bone density in the lumbar spine was -1.5±0.24 and -1.4±0.27 in ambulates and non-ambulates respectively (P=0.7). Mean bone density at proximal femur was -3.4±0.2 in ambulates and -3.4±0.3 in non-ambulates (p =0.48).Intra-groups statistical analysis showed significant difference between bone mineral density at lumbar spine and proximal femur in both mentioned groups (P<0.05). Vitamin D deficiency was detected in 13 patients (43.3%) and its serum level was significantly lower in non-ambulates compared with ambulates.Conclusion: High prevalence of vitamin D deficiency and osteoporosis was found in DMD patients although osteoporosis severity was not affected by muscle function and ambulation. So it seems that vitamin D supplementation can improve vitamin D status and osteoporosis in these patients, especially in non-ambulates.

The Prevalence of Hepatitis C Virus (HCV) Infection and Genotypes in Patients with Hemophilia and Other Blood Coagulopathies in Mashhad, Iran

Background and Aim: Patients with blood coagulopathies treated with multiple transfusions have a high risk to acquire some viral infections such as hepatitis C. This research was aimed to identify hepatitis C virus (HCV) infection prevalence, and the viral genotypes among patients with hemophilia and other inherited coagulopathies in Mashhad, Iran. Methods: Medical records of 760 patients with inherited coagulopathies including hemophilia were reviewed in Sarvar Clinic of Mashhad. Plasma samples were subjected to detect antibodies against HCV (anti-HCV) by enzyme-linked immunosorbent assay. HCV RNA and genotypes were determined by a real-time polymerase chain reaction (PCR) method. Results: Totally 128 individuals (16.8%) including patients with hemophilia (n=116) and individuals with other coagulopathies (n=12) were found to be seropositive for anti-HCV. They comprised 122 men and six women with a mean age of 31.6 ± 10.5 years. The PCR results were available for 118 patients, of whom 86 individuals (72.9%) were found to have detectable HCV RNA in their serum. The most frequent genotypes were 1a and 3a (49.1% and 35.8%, respectively). HCV genotypes were not significantly correlated with the patients’ age (p=0.477) as well as with the serum levels of alanine aminotransferase (p=0.655) and aspartate aminotransferase (p=0.332). Conclusion: The patients with blood coagulation disorders had a greater prevalence of HCV infection in comparison with the general population in our region. The most common subgenotypes of HCV were 1a, and 3a, respectively. These results could assist professionals to choose more efficient approaches for the management of their patients. *Corresponding Author: Mohammad Reza Hedayati-Moghaddam; Email: [email protected] Please cite this article as: Badiei Z, Ahmadi-Ghezeldasht S, Sima HR, Habibi M, Khamooshi M, Azimi A, Hedayati-Moghaddam MR. The Prevalence of Hepatitis C Virus (HCV) Infection and Genotypes in Patients with Hemophilia and Other Blood Coagulopathies in Mashhad, Iran. Arch Med Lab Sci. 2021;7:1-7 (e9). https://doi.org/10.22037/amls.v7.3396

GMO perceptions among Swedish stakeholders and their implication on the acceptance of a new biotechnological advancement

With the global population expected to reach more than 10 billion by the year 2050, one of the global challenges will be in meeting the growing food demand. Coupled with the effects of climate change on the environment, the world will need plant varieties that can adapt to changing environmental conditions to meet the food production challenge. A viable option to aid food production is using genetic modification for more specific plant breeding and crop optimization. The usage of genetic modification has been highly debated since the course of its first developed commercialized crop. The consumer concerns, stringent regulations and rigorous testing genetically modified crops face, may be circumvented with the legally undefined development of new biotechnological advancements such as CRISPR. This study aims to investigate whether arguments used against GMOs from different stakeholders and antiGM NGOs in Sweden are applicable to the CRISPR technique. Through content analysis of policy documents of influential stakeholders and a literature review of the scientific literature, major concerns with GMOs are identified and their applicability to the CRISPR technique analyzed. This study finds that the most prevalent reservations against GMOs do not apply to the new biotechnology, although the applicability and in effect acceptance of CRISPR among Swedish stakeholders depends on its legal classification, which the European Commission has yet to determine. The findings in this study also indicate that the reservations against GMOs are based on misconceptions and that if CRISPR falls into the same legal category, it will be subject to the same obstacles in its implementation. The implications of its categorical classification may in addition affect a possible utilization of CRISPR among Swedish crop producers in taking part in the global food production challenge. This study concludes that while the prevalent reservations against GMOs do not apply to CRISPR, its acceptance may solely rely on its legal classification and as such there is a need for a closer look on the regulations around the various biotechnologies, as well as a need for better biotechnological communication between researchers, decision makers and consumers

GMO perceptions among Swedish stakeholders and their implication on the acceptance of a new biotechnological advancement

With the global population expected to reach more than 10 billion by the year 2050, one of the global challenges will be in meeting the growing food demand. Coupled with the effects of climate change on the environment, the world will need plant varieties that can adapt to changing environmental conditions to meet the food production challenge. A viable option to aid food production is using genetic modification for more specific plant breeding and crop optimization. The usage of genetic modification has been highly debated since the course of its first developed commercialized crop. The consumer concerns, stringent regulations and rigorous testing genetically modified crops face, may be circumvented with the legally undefined development of new biotechnological advancements such as CRISPR. This study aims to investigate whether arguments used against GMOs from different stakeholders and antiGM NGOs in Sweden are applicable to the CRISPR technique. Through content analysis of policy documents of influential stakeholders and a literature review of the scientific literature, major concerns with GMOs are identified and their applicability to the CRISPR technique analyzed. This study finds that the most prevalent reservations against GMOs do not apply to the new biotechnology, although the applicability and in effect acceptance of CRISPR among Swedish stakeholders depends on its legal classification, which the European Commission has yet to determine. The findings in this study also indicate that the reservations against GMOs are based on misconceptions and that if CRISPR falls into the same legal category, it will be subject to the same obstacles in its implementation. The implications of its categorical classification may in addition affect a possible utilization of CRISPR among Swedish crop producers in taking part in the global food production challenge. This study concludes that while the prevalent reservations against GMOs do not apply to CRISPR, its acceptance may solely rely on its legal classification and as such there is a need for a closer look on the regulations around the various biotechnologies, as well as a need for better biotechnological communication between researchers, decision makers and consumers

GMO perceptions among Swedish stakeholders and their implication on the acceptance of a new biotechnological advancement

With the global population expected to reach more than 10 billion by the year 2050, one of the global challenges will be in meeting the growing food demand. Coupled with the effects of climate change on the environment, the world will need plant varieties that can adapt to changing environmental conditions to meet the food production challenge. A viable option to aid food production is using genetic modification for more specific plant breeding and crop optimization. The usage of genetic modification has been highly debated since the course of its first developed commercialized crop. The consumer concerns, stringent regulations and rigorous testing genetically modified crops face, may be circumvented with the legally undefined development of new biotechnological advancements such as CRISPR. This study aims to investigate whether arguments used against GMOs from different stakeholders and antiGM NGOs in Sweden are applicable to the CRISPR technique. Through content analysis of policy documents of influential stakeholders and a literature review of the scientific literature, major concerns with GMOs are identified and their applicability to the CRISPR technique analyzed. This study finds that the most prevalent reservations against GMOs do not apply to the new biotechnology, although the applicability and in effect acceptance of CRISPR among Swedish stakeholders depends on its legal classification, which the European Commission has yet to determine. The findings in this study also indicate that the reservations against GMOs are based on misconceptions and that if CRISPR falls into the same legal category, it will be subject to the same obstacles in its implementation. The implications of its categorical classification may in addition affect a possible utilization of CRISPR among Swedish crop producers in taking part in the global food production challenge. This study concludes that while the prevalent reservations against GMOs do not apply to CRISPR, its acceptance may solely rely on its legal classification and as such there is a need for a closer look on the regulations around the various biotechnologies, as well as a need for better biotechnological communication between researchers, decision makers and consumers

Induction and Suppression of Cell Lysis in an Electrokinetic Microfluidic System

The ability to strategically induce or suppress cell lysis is critical for many cellular-level diagnostic and therapeutic applications conducted within electrokinetic microfluidic platforms. The chemical and structural integrity of sub-cellular components is important when inducing cell lysis. However, metal electrodes and electrolytes participate in undesirable electrochemical reactions that alter solution composition and potentially damage protein, RNA, and DNA integrity within device microenvironments. For many biomedical applications, cell viability must be maintained even when device-imposed cell-stressing stimuli (e.g., electrochemical reaction byproducts) are present. In this work, we explored a novel and tunable method to accurately induce or suppress device-imposed artifacts on human red blood cell (RBC) lysis in non-uniform AC electric fields. For precise tunability, a dielectric hafnium oxide (HfO ) layer was used to prevent electron transfer between the electrodes and the electric double layer and thus reduce harmful electrochemical reactions. Additionally, a low concentration of Triton X-100 surfactant was explored as a tool to stabilize cell membrane integrity. The extent of hemolysis was studied as a function of time, electrode configuration (T-shaped and star-shaped), cell position, applied non-uniform AC electric field, with uncoated and HfO coated electrodes (50 nm), and absence and presence of Triton X-100 (70 μM). Tangible outcomes include a parametric analysis relying upon literature and this work to design, tune, and operate electrokinetic microdevices to intentionally induce or suppress cellular lysis without altering intracellular components. Implications are that devices can be engineered to leverage or minimize device-imposed biological artefacts extending the versatility and utility of electrokinetic diagnostics. This article is protected by copyright. All rights reserved

Changes in Membrane Dielectric Properties of Porcine Kidney Cells Provide Insight into the Antiviral Activity of Glycine

The ability to monitor the status and progression of viral infections is important for development and screening of new antiviral drugs. Previous research illustrated that the osmolyte glycine (Gly) reduced porcine parvovirus (PPV) infection in porcine kidney (PK-13) cells by stabilizing the capsid protein and preventing virus capsid assembly into viable virus particles. Dielectrophoresis (DEP) was examined herein as a noninvasive, electric field- and frequency-dependent tool for real-time monitoring of PK-13 cell responses to obtain information about membrane barrier functionality and polarization. DEP responses of PK-13 cells were compared to those of PPV-infected cells in the absence and presence of the osmolyte glycine. With infection progression, PK-13 DEP spectra shifted toward lower frequencies, reducing crossover frequencies (fCO). The spherical single-shell model was used to extract PK-13 cell dielectric properties. Upon PPV infection, specific membrane capacitance increased over the time progression of virus attachment, penetration, and capsid protein production and assembly. Following glycine treatment, the DEP spectra displayed attenuated fCO and specific membrane capacitance values shifted back toward uninfected PK-13 cell values. These results suggest that DEP can be used to noninvasively monitor the viral infection cycle and screen antiviral compounds. DEP can augment traditional tools by elucidating membrane polarization changes related to drug mechanisms that interrupt the virus infection cycle

pH Gradients in Spatially Non-Uniform AC Electric Fields around the Charging Frequency; A Study of Two Different Geometries and Electrode Passivation

Dielectrophoresis (DEP), a precision nonlinear electrokinetic tool utilized within microfluidic devices, can induce bioparticle polarization that manifests as motion in the electric field; this phenomenon has been leveraged for phenotypic cellular and biomolecular detection, making DEP invaluable for diagnostic applications. As device operation times lengthen, reproducibility and precision decrease, which has been postulated to be caused by ion gradients within the supporting electrolyte medium. This research focuses on characterizing pH gradients above, at, and below the electrode charging frequency (0.2–1.4 times charging frequency) in an aqueous electrolyte solution in order to extend the parameter space for which microdevice-imposed artifacts on cells in clinical diagnostic devices have been characterized. The nonlinear alternating current (AC) electric fields (0.07 Vpp/μm) required for DEP were generated via planar T-shaped and star-shaped microelectrodes overlaid by a 70 μm high microfluidic chamber. The experiments were designed to quantify pH changes temporally and spatially in the two microelectrode geometries. In parallel, a 50 nm hafnium oxide (HfO2) thin film on the microelectrodes was tested to provide insights into the role of Faradaic surface reactions on the pH. Electric field simulations were conducted to provide insights into the gradient shape within the microelectrode geometries. Frequency dependence was also examined to ascertain ion electromigration effects above, at, and below the electrode charging frequency. The results revealed Faradaic reactions above, at, and below the electrode charging frequency. Comparison experiments further demonstrated that pH changes caused by Faradaic reactions increased inversely with frequency and were more pronounced in the star-shaped geometry. Finally, HfO2 films demonstrated frequency-dependent properties, impeding Faradaic reactions

Impacts of low concentration surfactant on red blood cell dielectrophoretic responses.

Cell dielectrophoretic responses have been extensively studied for biomarker expression, blood typing, sepsis, circulating tumor cell separations, and others. Surfactants are often added to the analytical buffer in electrokinetic cellular microfluidic systems to lower surface/interfacial tensions. In nonelectrokinetic systems, surfactants influence cell size, shape, and agglomeration; this has not been systematically documented in electrokinetic systems. In the present work, the impacts of the Triton X-100 surfactant on human red blood cells (RBCs) were explored via ultraviolet-visible spectroscopy (UV-Vis) and dielectrophoresis (DEP) to compare nonelectrokinetic and electrokinetic responses, respectively. The UV-Vis spectra of Triton X-100 treated RBCs were dramatically different from that of native RBCs. DEP responses of RBCs were compared to RBCs treated with low concentrations of Triton X-100 (0.07-0.17 mM) to ascertain surfactant effects on dielectric properties. A star-shaped electrode design was used to quantify RBC dielectric properties by fitting a single-shell oblate cell model to experimentally-derived DEP spectra. The presence of 0.07 and 0.11 mM of Triton X-100 shifted the RBC\u27s DEP spectra yielding lower crossover frequencie