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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- Analysis of cellular senescence in Spinocerebellar ataxia type 7 using bioinformatics and an inducible cell model(Instituto Tecnológico y de Estudios Superiores de Monterrey, 2022-06-15) Ruiz Esparza Palacios, Vanessa; GONZALEZ MELJEM, JOSE MARIO; 316764; González Meljem, José Mario; puemcuervo; Pérez Méndez, Óscar Armando; García Aguirre, Ian Alaín; School of Engineering and Sciences; Campus Monterrey; Magaña Aguirre, Jonathan JavierSpinocerebellar ataxia type 7 (SCA7) is an autosomal dominant inherited disorder manifested by the inability to coordinate balance, gait, and speech. Some experimental evidence suggests that nuclear inclusion of mutant ataxin-7 (ATX7) is part of the molecular basis of this disease and the origin of oxidative stress. In this thesis, we made a bioinformatic analysis that supports the hypothesis that SCA7 could share some mechanisms that are core features of senescent cells. Senescence is a permanent state of cell-cycle arrest, and core features of this phenotype include the expression of anti-proliferative molecules, the activation of a chronic DNA damage response, altered metabolic rates, and many others. We sought to establish a senescence induction protocol in a human fibroblast cell line as a positive control of senescence and found that induction with H2O2 at 1200 μM resulted in 79% of positive SA-β-gal cells. Afterwards, we were able to validate an inducible cell model of MIO-M1 cells, which were stably transduced to express a mutant ATXN7 gene carrying 64 CAG repeats and 10 CAG repeats. Administration of doxycycline (dox) induced the expression of the mutated protein causing the formation of nuclear aggregates. Furthermore, in a preliminary SA-β-gal assay, we found activity of this enzyme in 64 CAG cells, suggesting the presence of senescence features after the induction of the mutated protein. Based on our findings, we propose that the oxidative stress generated by the accumulation of the mutated protein could be leading to a senescence phenotype. Future evaluation of the Senescence-associated secretory phenotype (SASP) and markers of DNA damage will bring better understanding of the possible role of senescence in SCA7.
- Bioinformatic analysis of the expression pattern of an intron-retaining isoform of the Agrp transcript in arcuate nucleus neurons of mice(Instituto Tecnológico y de Estudios Superiores de Monterrey, 2021-12-07) Gómez Montalvo, Jesús; GONZALEZ MELJEM, JOSE MARIO; 316764; González Meljem, José Mario; puemcuervo, emipsanchez; Treviño Alvarado, Víctor Manuel; de Obeso Fernández del Valle, Álvaro; School of Engineering and Sciences; Campus Monterrey; Avendaño Vázquez, Selma EréndiraKnown to be the regulatory center of hunger and satiety, the arcuate nucleus (ARC) harbors two neuronal populations that exert opposite effects on the regulation of food intake. Namely, AgRP and POMC neurons respectively trigger hunger and satiety. Although both neuronal groups are born within the same time interval in the developing hypothalamus of mouse, their peptidergic identities are established at distinct developmental timepoints. While the POMC identity is established as early as the embryonic period, the complete maturation of the AgRP peptidergic identity extends to the postnatal period. Previously, through RT-PCR, our group detected the presence of an Agrp transcript isoform that retained introns 3 and 4 (Agrp-i3,4) in the early postnatal, but not in the adult mouse hypothalamus. In this thesis project, the expression pattern of Agrp-i3,4 is analyzed in public RNA-seq datasets from ARC neurons of mice at different postnatal developmental stages. To identify intron retention events, iREAD was used, and to quantify the proportion of Agrp transcripts that retained introns 3 and 4, ASpli was employed. Using this bioinformatics approach, the largest proportion of Agrp-i3,4 was detected in ARC samples of P12 mice and there was a trend towards decreased retention of Agrp introns 3 and 4 at later developmental stages. Agrp-i3,4 was detected in poly-A RNA extracted from whole AgRP neurons, but not in the ribosomal fraction. On the other hand, food deprivation appeared to exert distinct effects on the proportion of the Agrp-i3,4 transcript depending on the duration of the fast. While in weaned mice fasted for 16 hours Agrp-i3,4 showed a slight increase, in adult mice fasted for 38 hours Agrp-i3,4 appeared to decrease. Unlike fasting, leptin treatment did not exert any effect on the retention of Agrp introns 3 and 4. Of note, IR was found in Agrp but not in other genes that characterize the ARC, such as Npy or Pomc. Taken together, the results presented in this thesis suggest that increased IR of the Agrp transcript may correlate with a lower maturation degree of the ARC during a time interval in which the AgRP peptidergic identity has not yet been fully established.
- On the improvement of in vitro senescence studies by applying microfluidic technology and mathematical modelling(Instituto Tecnológico y de Estudios Superiores de Monterrey, 2021-06-09) Rios Maciel, Mauricio Andrés; González Meljem, José Mario; puelquio; Álvarez, Mario Moisés; Trujillo de Santiago, Grissel; Escuela de Ingeniería y Ciencias; Campus MonterreyCellular senescence can be defined as an irreversible cell cycle arrest, which has been termed as an ‘antagonistic pleiotropy’ because of the duality of its role in aging. While the early-life on-set of the phenotype prevents tumorigenesis and plays an important part on the immune system-mediated removal of damaged cells, accumulation of senescent cells late in life is linked to the development of age-related diseases; thus, senescence is a target for anti-aging therapies. However, the senescent phenotype is very complex and heterogenous, making in vitro approaches desirable for the study of their individual components. In vitro culture of senescence presents a complex combination of different types of cells, including senescent, non-senescent, arrested and apoptotic, making it challenging to study the process in a complete manner. In addition, senescent cells have complex intercellular interactions, as well as ECM interactions. Recent approaches have used microfluidic technology alongside the incorporation of three-dimensional hydrogel-based matrixes to better mimic the microenvironment of the cell in vitro to make it more physiologically relevant. In this work we focus on the preliminary work for a further translation of traditional in vitro senescence techniques to a 3D and microfluidic environment. We assessed, analyzed and verified the behavior of endothelial HMEC-1 cells when cultured on GelMA-based hydrogel, as well as the effects of the matrix in senescence induction and assessment methods. We developed a mathematical model based on reported data to identify variables of importance and simulate the transition between states of proliferation, death and senescence through time in a senescent culture induced through doxorubicin. We observed that doxorubicin-induced senescence in endothelial cells stimulates apoptosis but yields an acceptable percentage of induction. After transferring these experiments to the GelMA-based hydrogel, neither induction nor assessment seemed to present a problem. We observed that endothelial cells in GelMA tend to adjust to a specific percentage, giving insight into the importance of the pore size in the matrix. Finally, the mathematical model shed light on the importance of time after treatment in the transition to senescent cells but also the turnover to dead population.
- The role of HDL and phospholipids in endothelial cellular senescence(Instituto Tecnológico y de Estudios Superiores de Monterrey, 2021-05-26) De la Cruz Gutiérrez, Luis Fernando; DE LA CRUZ GUTIERREZ, LUIS FERNANDO; 672038; González Meljem, José Mario; puemcuervo; Magaña Aguirre, Jonathan Javier; Chávez Santos, Rocío Alejandra; School of Engineering and Sciences; Campus Monterrey; Pérez Méndez, Oscar ArmandoHigh-density lipoproteins (HDLs) are commonly recognized as antiatherogenic molecules; these possess anti-inflammatory properties and participate in the production of Nitric Oxide (NO) by endothelial cells. In addition, critical patients characterized by a dropdown of HDL-cholesterol plasma levels during the acute phase, have an increased risk of intrahospital death. The molecular mechanisms underlying the beneficial effects of HDL are not well understood. However, current clinical evidence suggests a relationship between HDL and cellular senescence, a stress response characterized by a mostly irreversible loss in the replicative potential of the cell. This permanent cell cycle arrest can be induced through a variety of stimuli including telomere shortening, oxidative stress, DNA damage and oncogenic signals. At the organismal level, senescence contributes to aging and has been associated with numerous age-related pathologies. Now, few studies have documented that cellular senescence exhibits an alteration in lipid composition, leading to morphological changes in membrane remodeling. Additionally, a relationship between the altered expression of regulatory genes of lipid metabolism and senescent cells has been recently uncovered. Therefore, here we aimed to analyze the role of HDLs in endothelial cell senescence induced by doxorubicin and hydrogen peroxide. The results reveal that there is a possible impact of HDL on the onset of cellular senescence, although this may be more associated with membrane homeostasis, as reflected in a decrease in the number of cells positive for the senescence marker SA-β-galactosidase, as well as a decrease in apoptosis associated with the stress generated by the senescence inducer. However, further study of these lipoproteins in senescent cells is needed. Likewise, this study opens the door to consider hydrogen peroxide as a strong candidate to induce senescence in HMEC-1 endothelial cells derived from the oxidative stress that these cells can encounter physiologically.