Ciencias Exactas y Ciencias de la Salud
Permanent URI for this collectionhttps://hdl.handle.net/11285/551039
Pertenecen a esta colección Tesis y Trabajos de grado de las Maestrías correspondientes a las Escuelas de Ingeniería y Ciencias así como a Medicina y Ciencias de la Salud.
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- “Determination of interaction properties between PEGylated proteins and a modified resin by Isothermal Titration Calorimetry (ITC) and FTIR”(Instituto Tecnológico y de Estudios Superiores de Monterrey, 2018-05-25) Magaña, Paulyna; Aguilar-Jiménez, Oscar Alejandro; Gonzalez-Valdez, José Guillermo; Ramos de la Peña, Ana MayelaPEGylated proteins are an increasing important class of therapeutic drugs due to their improved pharmacokinetic characteristics and solubility over their corresponding native forms. PEGylation is the covalent attachment of one or more polyethylene glycol (PEG) molecules to a protein. Despite the many advantages of PEGylated drugs, one of the major challenges is the purification step after the chemical reaction. The main purpose of this project is to determine the nature of chemical interactions between a modified resin with PEG 5000 g/mol and PEGylated proteins that results in a previously demonstrated ability of such resins for the resolution of PEGylated proteins. A chromatographical separation of PEGylated proteins was additionally demonstrated for lysozyme using the modified resin Sepharose 6B-PEG5000 previously reported for PEGylated RNase A. Fourier Transform Infrared (FTIR) spectroscopy provided insight of the resin modification. The interaction thermodynamics associated with PEGylated proteins in hydrophobic interaction chromatography (HIC) with modified resin was carried out in with an ITC (Isothermal Titration Calorimetry) analysis. The specific enthalpy (∆G) was found to be exothermic for both proteins in potassium phosphate buffer with 1.5 M ammonium sulfate at 25ºC. MonoPEGylated proteins showed large negative entropy (-T∆S) values, related to the enhanced hydrophobic interaction between PEG5000 molecules from the resin and PEGylated protein forms. In addition, binding constants (K) of PEGylated proteins to modified resin were slightly higher compared to unmodified proteins.
- Biogenic silica as a novel carrier to encapsulate isorhamnetin using a microfluidic device(Instituto Tecnológico y de Estudios Superiores de Monterrey, 2018-05-14) Mancera-Andrade, Elena Ivonne; Parra-Saldivar, Roberto; Iqbal, Hafiz M. N.; González-Valdez, José GuillermoDiatoms have the peculiarity to synthesize amorphous silica around the cell wall. Frustules (empty silica shells) have the advantages of being biocompatible, biodegradable, nontoxic and rich with OH groups on the surface. Frustules have been used in diverse fields, but recently their application in the biomedical field has been investigated. Drug delivery systems (DDS) have been studied to improve the therapeutic effect of different drugs, especially hydrophobic drugs. Different encapsulation methodologies have been used to load the drug in a carrier such as drop-wise methodology or solvent evaporation. However, a reproducible methodology that reduces handling error must be explored. In the present work, a microfluidic device is used as a novel encapsulation technique for solid particles and hydrophobic drugs. A novel microfluidic device fabrication technique was used: ESCARGOT (Embedded SCAffold RemovinG Open Technology). Isorhamnetin was used as a model drug which has a hydrophobic nature. Three different concentrations were studied: 20, 60 and 100 μg/mL, and three different resident times in the device (0.4, 1 and 2 minutes). The highest encapsulation efficiency (EE%) and loading capacity (LC%) were 17.92% and 1.63% respectively. According to the statistical analysis, the optimum conditions to obtain a maximum (EE%) were 2 minutes and 20 μg/mL. The isorhamnetin release behavior was observed with a burst release in the first hour with 48.26%, while the total amount of drug was delivered in three hours. The feasibility of frustules as carriers and the microfluidic device as a mixer was successfully accessed. This methodology could be used as a standardization technique to obtain reproducible results. Further studies with frustule surface functionalization need to be performed to improve EE%.
- Characterizing the role of mitochondrial Ca2+ overload and permeability transition during the BCR-dependent activation of murine B lymphocytes(2017-12-05) Torres Quintanilla, Alejandro Daniel; González Castillo, Elena Cristina; García Rivas, Gerardo; Maravillas Montero, José Luis; Torre Amione, GuillermoMitochondria have proved their role as major players for immunometabolism, regulating cell function by metabolic signals. Indeed, mitochondrial function during lymphocyte activation goes beyond ATP supply, as recent evidence shows that mitochondrial Ca2+ uptake and mitochondrial ROS (mROS) production regulate lymphocyte activation. Furthermore, the adaptive immune response depends on the differentiation of lymphocytes into various subsets. Whether metabolic signals determine lymphocyte differentiation has just began to be explored. Interestingly, recent evidence shows that mitochondrial signals determine B lymphocyte differentiation into memory cells. During lymphocyte activation, Ca2+ uptake into mitochondria has proven to be an important mechanism of regulation. However, if mitochondrial Ca2+ uptake capacity impacts B cell differentiation and activation is not yet determined. Here, we have generated an in vitro model of B lymphocyte mitochondrial Ca2+ overload to explore whether this mechanism is involved in B cell activation. Primary mice B lymphocytes were subjected to activation or thapsigargin treatment to induce Ca2+ overload. The effect on the expression of activation surface marker CD69 was explored using flow cytometry. Mitochondrial contribution was explored by Ca2+ uptake inhibition using Ru360 or treatment with the mitochondrial antioxidant MitoTEMPO. Mitochondrial membrane potential, Ca2+ uptake, and content were determined. Results show that our in vitro model effectively induces mitochondrial Ca2+ overload, as seen by higher Ca2+ content and mitochondrial membrane depolarization. Moreover, mitochondrial Ca2+ overload contributes to the expression of CD69 via mROS production. Lastly, results suggest that during BCR-dependent activation, mitochondria take up Ca2+ to increase mROS production that can alter the signaling cascade. The in vitro model generated herein, should allow for further exploration of whether mitochondrial Ca2+ overload impacts B cell differentiation.