Using the reactive/transport dispersive models to simulate a monolithic anion exchanger: Experimental parameter determination, simultaneous model evaluation, and validation.

Selecting an adequate model to represent the mass transfer mechanisms occurring in a chromatographic process is generally complicated, which is one of the reasons why monolithic chromatography is scarcely simulated. In this study, the chromatographic separation of model proteins bovine serum albumin (BSA), ß-lactoglobulin-A, and ß-lactoglobulin-B on an anion exchange monolith was simulated based on experimental parameter determination, simultaneous model testing, and validation under three statistical criteria: retention time, dispersion accuracies, and Pearson correlation coefficient. Experimental characterization of morphologic, physicochemical, and kinetic parameters was performed through volume balances, pressure drop analysis, breakthrough curve analysis, and batch adsorptions. Free Gibbs energy indicated a spontaneous adsorption process for proteins and counterions. Dimensionless numbers were estimated based on height equivalent to a theoretical plate analysis, finding that pore diffusion controlled ß-lactoglobulin separation, whereas adsorption/desorption kinetics was the dominant mechanism for BSA. The elution profiles were modeled using the transport dispersive model and the reactive dispersive model coupled with steric mass action (SMA) isotherms because these models allowed to consider most of the mass transport mechanisms that have been described. RDM-SMA presented the most accurate simulations at pH 6.0 and at low (250 mM) and high (400 mM) NaCl concentrations. This simulation will be used as reference to forecast the purification of these proteins from bovine whey waste and to extrapolate this methodology to other monolith-based separations using these three statistical criteria that have not been used previously for this purpose.

The role of lipids in exosome biology and intercellular communication: Function, analytics and applications.

Exosomes are extracellular vesicles that in recent years have received special attention for their regulatory functions in numerous biological processes. Recent evidence suggests a correlation between the composition of exosomes in body fluids and the progression of some disorders, such as cancer, diabetes and neurodegenerative diseases. In consequence, numerous studies have been performed to evaluate the composition of these vesicles, aiming to develop new biomarkers for diagnosis and to find novel therapeutic targets. On their part, lipids represent one of the most important components of exosomes, with important structural and regulatory functions during exosome biogenesis, release, targeting and cellular uptake. Therefore, exosome lipidomics has emerged as an innovative discipline for the discovery of novel lipid species with biomedical applications. This review summarizes the current knowledge about exosome lipids and their roles in exosome biology and intercellular communication. Furthermore, it presents the state-of-the-art analytical procedures used in exosome lipidomics while emphasizing how this emerging discipline is providing new insights for future applications of exosome lipids in biomedicine.

Production and purification of bacterial membrane vesicles for biotechnology applications: Challenges and opportunities.

Bacterial membrane vesicles (BMVs) are bi-layered nanostructures derived from Gram-negative and Gram-positive bacteria. Among other pathophysiological roles, BMVs are critical messengers in intercellular communication. As a result, BMVs are emerging as a promising technology for the development of numerous therapeutic applications. Despite the remarkable progress in unveiling BMV biology and functions in recent years, their successful isolation and purification have been limited. Several challenges related to vesicle purity, yield, and scalability severely hamper the further development of BMVs for biotechnology and clinical applications. This review focuses on the current technologies and methodologies used in BMV production and purification, such as ultracentrifugation, density-gradient centrifugation, size-exclusion chromatography, ultrafiltration, and precipitation. We also discuss the current challenges related to BMV isolation, large-scale production, storage, and stability that limit their application. More importantly, the present work explains the most recent strategies proposed for overcoming those challenges. Finally, we summarize the ongoing applications of BMVs in the biotechnological field.

A monolithic stationary phase with dendritic nanostructures for the separation of PEGylated proteins.

Separation of PEGylated protein mixtures into individual species is a challenging procedure, and many efforts have been focused on creating novel chromatographic supports for this purpose. In this study, a new monolithic stationary phase with hyperbranched nanostructures was chemically synthesized. For this, monoliths with a support matrix of poly (glycidyl methacrylate-co-ethylene dimethacrylate) and ethylenediamine chemistry were modified with third-generation dendrons with butyl-end groups. The new monolith was analyzed by infrared spectroscopy, confirming the dendron with butyl ligands and exhibited low mass transfer resistance as observed by breakthrough frontal analysis. This support was able to separate mono-PEG ribonuclease A from the PEGylation mixture, indicated by a single band (∼30 kDa) in the electrophoretic analysis. Moreover, the separation of mono-PEGylated positional isomers was probably observed, as the protein with ∼30 kDa was found in two separate peaks. Interestingly, the dendronized monolith allowed the separation of the reaction mixture into individual PEGylated species when using high ammonium sulfate concentrations (2 M). A correlation between the PEGylation degree and the strength of the hydrophobic interactions on the monolith was observed. This chromatographic approach combines the natural branched architecture of dendrons and the higher capabilities of the monoliths enhancing the hydrophobic surface area, and therefore the interaction between the PEGylated proteins and ligands. Thus, the novel support represents a novel platform for the purification of PEGylated from non-PEGylated proteins with biotechnological applications.

HIV Stigma Mechanisms Scale: Factor Structure, Reliability, and Validity in Mexican Adults.

We aimed to validate the HIV Stigma Mechanisms Scale (HIV-SMS) in a sample of Mexican adults living with HIV, which differentiates between sources and mechanisms of stigma. Adults (n = 362) with a median age of 32 years old completed a web-based version in Spanish of the HIV-SMS as well as sociodemographic and HIV-related characteristics questionnaire. Exploratory factor analyses with weighted least squares and oblique rotation were performed to assess the construct validity of the scale. The Spanish translation for the Mexican population of the HIV-SMS has adequate internal consistency (Ω = 0.86) and demonstrated a structure similar to the original scale. After excluding the items related to community and social workers, a five-factor solution with internalized, promulgated, and anticipated stigma from family and healthcare workers showed adequate construct validity. The HIV-SMS is a valid and sensitive scale that can be used in a Mexican adult population living with HIV.

RESUMEN: El objetivo de este estudio fue validar la Escala de Mecanismos de Estigma de VIH (EME-VIH) en una muestra de adultos mexicanos que viven con VIH. Esta escala distingue entre fuentes y mecanismos de estigma. 362 adultos con una edad media de 32 años completaron vía web una versión en español de la EME-VIH así como preguntas acerca de sus características sociodemográficas y cuestiones relacionadas con el VIH. Se realizaron análisis factoriales exploratorios de mínimos cuadrados ponderados con rotación oblicua para evaluar la validez de constructo de la escala. La traducción al español de la EME-VIH para población mexicana tiene consistencia interna adecuada (Ω = 0.86) y muestra una estructura similar a la escala original. Después de excluir los ítems relacionados con trabajadores comunitarios y sociales, se encontró una solución con validez de constructo adecuada de cinco factores: estigma internalizado, promulgado y anticipado ejercido por la familia y personal de salud. La EME-VIH es una escala válida y sensible que puede usarse en población adulta mexicana que vive con VIH.

Biological nanoparticles: Relevance as novel target drug delivery systems and leading chromatographic isolation approaches.

Nanotechnology is one of the most promising technologies of the 21st century, and it is now presenting an enormous impact on target drug delivery. In this context, the recent use of natural vesicle-like nanoparticles such as extracellular vesicles (i.e., exosomes, microvesicles, and apoptotic bodies) and virus-like particles is rendering encouraging results mostly because these delivery systems present cargo versatility, favorable body circulating advantages, biocompatibility, immunogenicity, and the capacity to be modified superficially to increase their affinity to a certain target or to control their entrance to the cell. However, some of the biggest challenges toward their clinical implementation are poorly standardized processing operations due to their inherent heterogeneity and expensive, long-lasting, and difficult to scale isolation procedures that can also affect the stability of the particles. Under these circumstances, chromatographic procedures represent an attractive and favorable alternative to overcome their downstream processing. Moreover, even when standardized chromatographic purification protocols are still in development, great achievements have been made using size exclusion, ionic exchange, hydrophobic interaction, and affinity protocols, mostly because of the correct harnessing of the nanovesicle membrane properties. In this sense, this review focuses on presenting the current understanding on the most promising therapeutic biological nanoparticles and the chromatographic isolation approaches employed in their recovery, providing at the same time recent findings and a general overview of the aspects that might impact the outcome of chromatographic techniques for this application.

High-throughput assessment of hemoglobin polymer in single red blood cells from sickle cell patients under controlled oxygen tension.

Sickle cell disease (SCD) is caused by a variant hemoglobin molecule that polymerizes inside red blood cells (RBCs) in reduced oxygen tension. Treatment development has been slow for this typically severe disease, but there is current optimism for curative gene transfer strategies to induce expression of fetal hemoglobin or other nonsickling hemoglobin isoforms. All SCD morbidity and mortality arise directly or indirectly from polymer formation in individual RBCs. Identifying patients at highest risk of complications and treatment candidates with the greatest curative potential therefore requires determining the amount of polymer in individual RBCs under controlled oxygen. Here, we report a semiquantitative measurement of hemoglobin polymer in single RBCs as a function of oxygen. The method takes advantage of the reduced oxygen affinity of hemoglobin polymer to infer polymer content for thousands of RBCs from their overall oxygen saturation. The method enables approaches for SCD treatment development and precision medicine.

A Crest Factor Reduction Technique for LTE Signals with Target Relaxation in Power Amplifier Linearization.

The signal conditioning treatment to achieve good relation of power with radio-frequency (RF) conversion in conventional transceiver systems require precise baseband models. A developed framework is built to provide a demonstration of the modeling figures of merit with orthogonal frequency division multiplexing (OFDM) support under signal conditioning and transmission restrictions to waveforms with high peak to average power ratio (PAPR) in practical applications. Therefore, peak and average power levels have to be limited to correct high PAPR for a better suited correction power from the amplifier that can lead to compression or clipping in the signal of interest. This work presents an alternative joint crest factor reduction (CFR) algorithm to correct the performance of PAPR. A real-time field-programmable gate array (FPGA) testbed is developed to characterize and measure the behavior of an amplifier using a single-carrier 64-QAM OFDM based on long-term evolution (LTE) downlink at 2.40 GHz as stimulus, across wide modulation bandwidths. The results demonstrate that the CFR accuracy capabilities for the signal conditioning show a reliable clipping reduction to give a smooth version of the clipping signal and provide a factor of correction for the unwanted out-of-band emission validated according to the adjacent channel power ratio (ACPR), PAPR, peak power, complementary cumulative distribution function (CCDF), and error vector magnitude (EVM) figures of merit.

A Comparison of Surrogate Behavioral Models for Power Amplifier Linearization under High Sparse Data.

A good approximation to power amplifier (PA) behavioral modeling requires precise baseband models to mitigate nonlinearities. Since digital predistortion (DPD) is used to provide the PA linearization, a framework is necessary to validate the modeling figures of merit support under signal conditioning and transmission restrictions. A field-programmable gate array (FPGA)-based testbed is developed to measure the wide-band PA behavior using a single-carrier 64-quadrature amplitude modulation (QAM) multiplexed by orthogonal frequency-division multiplexing (OFDM) based on long-term evolution (LTE) as a stimulus, with different bandwidths signals. In the search to provide a heuristic target approach modeling, this paper introduces a feature extraction concept to find an appropriate complexity solution considering the high sparse data issue in amplitude to amplitude (AM-AM) and amplitude to phase AM-PM models extraction, whose penalties are associated with overfitting and hardware complexity in resulting functions. Thus, experimental results highlight the model performance for a high sparse data regime and are compared with a regression tree (RT), random forest (RF), and cubic-spline (CS) model accuracy capabilities for the signal conditioning to show a reliable validation, low-complexity, according to the peak-to-average power ratio (PAPR), complementary cumulative distribution function (CCDF), coefficients extraction, normalized mean square error (NMSE), and execution time figures of merit. The presented models provide a comparison with original data that aid to compare the dimension and robustness for each surrogate model where (i) machine learning (ML)-based and (ii) CS interpolate-based where high sparse data are present, NMSE between the CS interpolated based are also compared to demonstrate the efficacy in the prediction methods with lower convergence times and complexities.

An Experimental-Computational Approach to Quantify Blood Rheology in Sickle Cell Disease.

In sickle cell disease, aberrant blood flow due to oxygen-dependent changes in red cell biomechanics is a key driver of pathology. Most studies to date have focused on the potential role of altered red cell deformability and blood rheology in precipitating vaso-occlusive crises. Numerous studies, however, have shown that sickle blood flow is affected even at high oxygen tensions, suggesting a potentially systemic role for altered blood flow in driving pathologies, including endothelial dysfunction, ischemia, and stroke. In this study, we applied a combined experimental-computation approach that leveraged an experimental platform that quantifies sickle blood velocity fields under a range of oxygen tensions and shear rates. We computationally fitted a continuum model to our experimental data to generate physics-based parameters that capture patient-specific rheological alterations. Our results suggest that sickle blood flow is altered systemically, from the arterial to the venous circulation. We also demonstrated the application of this approach as a tool to design patient-specific transfusion regimens. Finally, we demonstrated that patient-specific rheological parameters can be combined with patient-derived vascular models to identify patients who are at higher risk for cerebrovascular complications such as aneurysm and stroke. Overall, this study highlights that sickle blood flow is altered systemically, which can drive numerous pathologies, and this study demonstrates the potential utility of an experimentally parameterized continuum model as a predictive tool for patient-specific care.

State-of-the-art exosome loading and functionalization techniques for enhanced therapeutics: a review.

Exosomes are a subpopulation of cell membrane-derived vesicles which play an essential role in cellular communication. In recent years, several studies have exploited the natural properties of exosomes as nanocarriers for several applications such as immunotherapy or drug delivery. Consequently, numerous techniques have been developed to improve their immunogenicity, drug loading efficiency, or targeting. Nonetheless, to date, there is no consensus on which technique results in more advantages for this purpose. In this context, this review discusses the currently used methodologies regarding traditional and engineered exosome loading and targeting techniques. Here, we focus on the advantages and disadvantages of each method while discussing some results obtained in relevant reports. Although there is a lack of evidence regarding the effects of exogenous exosomes in humans and several limitations in exosome isolation and purification techniques at the large-scale exist, the formulation of new exosome-based therapeutics is in the spotlight. Therefore, the development of more efficient functionalization techniques is required to reduce the potential risks associated with the clinical use of these vesicles.

Electrokinetically Driven Exosome Separation and Concentration Using Dielectrophoretic-Enhanced PDMS-Based Microfluidics.

Exosomes are a specific subpopulation of extracellular vesicles that have gained interest because of their many potential biomedical applications. However, exosome isolation and characterization are the first steps toward designing novel applications. This work presents a direct current-insulator-based dielectrophoretic (DC-iDEP) approach to simultaneously capture and separate exosomes by size. To do so, a microdevice consisting of a channel with two electrically insulating post sections was designed. Each section was tailored to generate different nonuniform spatial distributions of the electric field and, therefore, different dielectrophoretic forces acting on exosomes suspended in solution. Side channels were placed adjacent to each section to allow sample recovery. By applying an electric potential difference of 2000 V across the length of the main channel, dielectrophoretic size-based separation of exosomes was observed in the device. Analysis of particle size in each recovered fraction served to assess exosome separation efficiency. These findings show that iDEP can represent a first step toward designing a high-throughput, fast, and robust microdevice capable of capturing and discriminating different subpopulations of exosomes based on their size.

Recent advances and challenges in the recovery and purification of cellular exosomes.

Exosomes are nanovesicles secreted by most cellular types that carry important biochemical compounds throughout the body with different purposes, playing a preponderant role in cellular communication. Because of their structure, physicochemical properties and stability, recent studies are focusing in their use as nanocarriers for different therapeutic compounds for the treatment of different diseases ranging from cancer to Parkinson's disease. However, current bioseparation protocols and methodologies are selected based on the final exosome application or intended use and present both advantages and disadvantages when compared among them. In this context, this review aims to present the most important technologies available for exosome isolation while discussing their advantages and disadvantages and the possibilities of being combined with other strategies. This is critical since the development of novel exosome-based therapeutic strategies will be constrained to the effectiveness and yield of the selected downstream purification methodologies for which a thorough understanding of the available technological resources is needed.

Protein A chromatography: Challenges and progress in the purification of monoclonal antibodies.

Antibodies for therapeutic use are being continuously approved and their demand has been steadily growing. As known, the golden standard for monoclonal antibody (mAb) purification is Protein A affinity chromatography, a technology that has gained high interest because of its great performance and capabilities. The main concerns are the elevated resins costs and their limited lifetime compared to other resins (e.g. ion exchange chromatography). Great efforts have been carried out to improve purification conditions, such as resin characterization and designing alkali/acid stable resins with a longer lifetime. Modification of Protein A ligands and alternative formats such as monoliths membranes and microshperes have been tested to increase the purification performance. New technology has been proposed to improve the large-scale separation; in addition, alternative ligands have been suggested to capture mAbs instead of Protein A ligand; however, most of the information is locked by pharmaceutical companies. This mini review summarizes and describes the advances, results, and impact on the Protein A chromatography purification processing.

A modified method of local infiltration for endoscopic stapes surgery: how I do it.

PURPOSE: To present a modified method of local infiltration (MMLI) for endoscopic stapes surgery to reduce surgical time, bleeding and complications. MATERIALS AND METHODS: This study involved 70 patients who underwent stapes surgery for otosclerosis by endoscopic and microscopic approaches. The MMLI was applied as follows: local infiltration was performed with one hand while the other hand inserted the endoscope into the ear canal to observe vasoconstriction signs on the monitor; the single site of infiltration was located at the center of the anterior conchal cartilage. Operative time, intraoperative blood loss, preservation of anatomical structures, postoperative hearing and complications were evaluated. RESULTS: The MMLI allowed for quick bleeding control and a clear and dry operative field. Operative time, intraoperative blood loss and preservation of anatomical structures were significantly reduced in the endoscopic group (P < 0.00) versus the microscopic group. The scutum was removed less frequent in the endoscopic group 7.1% versus 53.6% of the microscopic group (P < 0.00). The chorda tympani was preserved in all cases but it was more manipulated in the microscopic group 39.3% versus 9.5% of the endoscopic group (P < 0.00). No complications were observed and the hearing outcomes were significantly better than the preoperative thresholds. CONCLUSIONS: This is the first report on the use of a MMLI for endoscopic stapes surgery. Using this method, the surgeon performs the infiltration at one site and concurrently observes the vasoconstriction signs without the use of a microscope, frontal lamp or speculum. This method provides benefits in terms of operative time and complications.

Design and Construction of an ROV for Underwater Exploration.

The design of a remotely operated vehicle (ROV) with a size of 18.41 cm × 29.50 cm × 33.50 cm, and a weight of 15.64 kg, is introduced herein. The main goal is to capture underwater video by remote control communication in real time via Ethernet protocol. The ROV moves under the six brushless motors governed through a smart PID controller (Proportional + Integral + Derivative) and by using pulse-wide modulation with short pulses of 1 µs to improve the stability of the position in relation to the translational, ascent or descent, and rotational movements on three axes to capture images of 800 × 640 pixels on a video graphic array standard. The motion control, 3D position, temperature sensing, and video capture are performed at the same time, exploiting the four cores of the Raspberry Pi 3, using the threading library for parallel computing. In such a way, experimental results show that the video capture stage can process up to 42 frames per second on a Raspberry Pi 3. The remote control of the ROV is executed under a graphical user interface developed in Python, which is suitable for different operating systems, such as GNU/Linux, Windows, Android, and OS X. The proposed ROV can reach up to 100 m underwater, thus solving the issue of divers who can only reach 30 m depth. In addition, the proposed ROV can be useful in underwater applications such as surveillance, operations, maintenance, and measurement.

New Trends in Biopolymer-Based Membranes for Pervaporation.

Biopolymers are currently the most convenient alternative for replacing chemically synthetized polymers in membrane preparation. To date, several biopolymers have been proposed for such purpose, including the ones derived from animal (e.g., polybutylene succinate, polylactic acid, polyhydroxyalcanoates), vegetable sources (e.g., starch, cellulose-based polymers, alginate, polyisoprene), bacterial fermentation products (e.g., collagen, chitin, chitosan) and specific production processes (e.g., sericin). Particularly, these biopolymer-based membranes have been implemented into pervaporation (PV) technology, which assists in the selective separation of azeotropic water-organic, organic-water, organic-organic mixtures, and specific separations of chemical reactions. Thereby, the aim of the present review is to present the current state-of-the-art regarding the different concepts on preparing membranes for PV. Particular attention is paid to the most relevant insights in the field, highlighting the followed strategies by authors for such successful approaches. Finally, by reviewing the ongoing development works, the concluding remarks and future trends are addressed.

Implementing a Chaotic Cryptosystem by Performing Parallel Computing on Embedded Systems with Multiprocessors.

Profiling and parallel computing techniques in a cluster of six embedded systems with multiprocessors are introduced herein to implement a chaotic cryptosystem for digital color images. The proposed encryption method is based on stream encryption using a pseudo-random number generator with high-precision arithmetic and data processing in parallel with collective communication. The profiling and parallel computing techniques allow discovery of the optimal number of processors that are necessary to improve the efficiency of the cryptosystem. That is, the processing speed improves the time for generating chaotic sequences and execution of the encryption algorithm. In addition, the high numerical precision reduces the digital degradation in a chaotic system and increases the security levels of the cryptosystem. The security analysis confirms that the proposed cryptosystem is secure and robust against different attacks that have been widely reported in the literature. Accordingly, we highlight that the proposed encryption method is potentially feasible to be implemented in practical applications, such as modern telecommunication devices employing multiprocessors, e.g., smart phones, tablets, and in any embedded system with multi-core hardware.

Knockout of Lmod2 results in shorter thin filaments followed by dilated cardiomyopathy and juvenile lethality.

Leiomodin 2 (Lmod2) is an actin-binding protein that has been implicated in the regulation of striated muscle thin filament assembly; its physiological function has yet to be studied. We found that knockout of Lmod2 in mice results in abnormally short thin filaments in the heart. We also discovered that Lmod2 functions to elongate thin filaments by promoting actin assembly and dynamics at thin filament pointed ends. Lmod2-KO mice die as juveniles with hearts displaying contractile dysfunction and ventricular chamber enlargement consistent with dilated cardiomyopathy. Lmod2-null cardiomyocytes produce less contractile force than wild type when plated on micropillar arrays. Introduction of GFP-Lmod2 via adeno-associated viral transduction elongates thin filaments and rescues structural and functional defects observed in Lmod2-KO mice, extending their lifespan to adulthood. Thus, to our knowledge, Lmod2 is the first identified mammalian protein that functions to elongate actin filaments in the heart; it is essential for cardiac thin filaments to reach a mature length and is required for efficient contractile force and proper heart function during development.

Short chain nitrocompounds as a treatment of layer hen manure and litter; effects on in vitro survivability of Salmonella, generic E. coli and nitrogen metabolism.

The current study was conducted to assess the bactericidal effectiveness of several nitrocompounds against pathogens in layer hen manure and litter. Evidence from an initial study indicated that treatment of layer hen manure with 12 mM nitroethane decreased populations of generic E. coli and total coliforms by 0.7 and 2.2 log10 colony forming units (CFU) g-1, respectively, after 24 h aerobic incubation at ambient temperature when compared to untreated populations. Salmonella concentrations were unaffected by nitroethane in this study. In a follow-up experiment, treatment of 6-month-old layer hen litter (mixed with 0.4 mL water g-1) with 44 mM 2-nitroethanol, 2-nitropropanol or ethyl nitroacetate decreased an inoculated Salmonella typhimurium strain from its initial concentration (3 log10 CFU g-1) by 0.7 to 1.7 log10 CFU g-1 after 6 h incubation at 37°C in covered containers. After 24 h incubation, populations of the inoculated S. Typhmiurium in litter treated with 44 mM 2-nitroethanol, 2-nitropropanol, ethyl nitroacetate or nitroethane were decreased more than 3.2 log10 CFU g-1 compared to populations in untreated control litter. Treatment of litter with 44 mM 2-nitroethanol, 2-nitropropanol, ethyl nitroacetate decreased rates of ammonia accumulation more than 70% compared to untreated controls (0.167 µmol mL-1 h-1) and loses of uric acid (< 1 µmol mL-1) were observed only in litter treated with 44 mM 2-nitropropanol, indicating that some of these nitrocompounds may help prevent loss of nitrogen in treated litter. Results warrant further research to determine if these nitrocompounds can be developed into an environmentally sustainable and safe strategy to eliminate pathogens from poultry litter, while preserving its nitrogen content as a nutritionally valuable crude protein source for ruminants.