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Efeitos do α-pineno livre ou encapsulado em lipossomas durante o cultivo de folículos pré-antrais e maturação oocitária em bovinos

2025-12-16T20:30:39Z

Título: Efeitos do α-pineno livre ou encapsulado em lipossomas durante o cultivo de folículos pré-antrais e maturação oocitária em bovinos Autor(es): Azevedo, Venância Antonia Nunes Abstract: Growing evidence has demonstrated that oxidative stress compromises in vitro culture (IVC) of ovarian follicles and in vitro maturation (IVM) of oocytes. Thus, strategies to reduce oxidative stress are essential to improve the quality of cultured follicles and oocytes. The objectives of this study were: 1) to investigate the effects of α-pinene during IVC of pre-antral follicles in bovine ovarian tissue; 2) to develop and characterize liposomes loaded with α-pinene and analyze their cytotoxicity in bovine cumulus cells (CCs) and evaluate their effects during IVM of bovine oocytes and subsequent parthenogenetic embryonic development. In Phase 1, ovarian tissue fragments were cultured for 6 days in α-MEM+ (control) or supplemented with 1.25, 2.5, 5, 10, or 20 µg/mL of α-pinene. In Phase 2, liposomes loaded with α-pinene (Lip-α-pinene) were produced using the lipid film hydration technique, characterized, and evaluated Regarding cytotoxicity in bovine cumulus oocyte complexes (COCs), cumulus oocyte complexes (COCs) were subsequently matured for 22-24 hours in maturation medium (control) or supplemented with Lip-white (empty liposome) or Lip-α-pinene at concentrations of 0.01, 1, or 100 µg/mL. After IVM, only mature oocytes proceeded to parthenogenetic activation and embryo culture. Data were analyzed using chi-square, Student's t-test, Kruskal-Wallis test, and ANOVA followed by Tukey's test (GraphPad Prism 9.0), with a significance level set at P < 0.05. In Phase 1, α-pinene at concentrations of 2.5, 5, or 10 µg/mL increased the percentage of normal follicles (P > 0.05) compared to the control, without influencing follicular growth (P < 0.05). α-pinene (10 µg/mL) maintained stromal cell density and collagen levels in cultured ovarian tissue similar to non-cultured tissues (P > 0.05) and increased mRNA levels for NRF2 and PRDX6 (P < 0.05) compared to the control. In Phase 2, Lip-α-pinene presented a size of 75.86 ± 0.95 nm, low polydispersity (0.26 ± 0.00), zeta potential of -31.55 ± 2.23 mV, encapsulation efficiency of 85 ± 2.51%, and Lip-α-pinene was not cytotoxic to CCs (P > 0.05). Lip-α-pinene did not affect the nuclear maturation rate (P > 0.05); however, 1 µg/mL of Lip-α-pinene preserved the ultrastructure of CCs and the oocyte, reduced ROS and lipid accumulation compared to the control, Lip-blank, and 100 µg/mL of Lip-α-pinene (P < 0.05). This result was accompanied by an increase in NRF2, SOD, and PRDX6 levels (P < 0.05). Furthermore, 1 and 100 µg/mL Lip-α-pinene increased cleavage rates and the number of cells per blastocyst (P < 0.05), but did not affect blastocyst formation and lipid content (P > 0.05). In conclusion, α-pinene preserves follicular structure, tissue integrity, and modulates antioxidant genes during ovarian tissue culture. When encapsulated, α-pinene increases the antioxidant capacity of oocytes by reducing ROS and increasing NRF2, SOD, and PRDX6 levels, improving the quality of oocytes and parthenogenetic embryos. Tipo: Tese

Efeitos protetores in vitro e in vivo da aloe vera contra danos ovarianos induzidos por Doxorrubicina em camundongos fêmeas

2025-11-27T16:34:06Z

Título: Efeitos protetores in vitro e in vivo da aloe vera contra danos ovarianos induzidos por Doxorrubicina em camundongos fêmeas Autor(es): Assis, Ernando Igo Teixeira de Abstract: Doxorubicin (DOX) is used to treat several types of cancer, but it causes ovarian toxicity by increasing the formation of reactive oxygen species (ROS), causing oxidative stress and an exacerbated inflammatory response. Thus, the use of natural compounds with antioxidant and anti-inflammatory properties against the toxic side effects induced by antineoplastic drugs has been highlighted. This study aimed to investigate the action of Aloe vera extract against the deleterious effects of DOX on follicular survival and growth in female mice. For this purpose, female Swiss mice with a regular estrous cycle in phases 1 and 2 were used. For phase 1 the animals (n = 37) had their ovaries collected and cultured individually in a 24-well plate at 37.5° C, in 5% CO2, for 6 days. The ovaries were cultured in DMEM+ alone; in medium supplemented with DOX (0.3 μg/mL); or in this case in DMEM+ supplemented with DOX (0.3 μg/mL) combined with Aloe vera (5%, 10%, 25% or 50%). After culture, the ovaries were collected and fixed for analysis. In phase 2, the animals (n = 48) were divided into six groups (8 per group): positive control (NAC+DOX), which received pretreatment with N-acetylcysteine orally (PO) and, 1 hour later, a single intraperitoneal (IP) dose of DOX; negative control (SAL+DOX), with PO pretreatment with saline followed, 1 hour later, by DOX (IP); three groups pretreated with Aloe vera extract (0.1, 1.0 or 10.0 mg/kg, PO) and, 1 hour later, subjected to a dose of DOX (IP) (AV0.1+DOX, AV1.0+DOX and AV10.0+DOX); and control (SAL+SAL), which received saline solution by PO and IP. DOX was administered in a single dose and Aloe vera was administered on three consecutive days; 24 hours after the last application (on the fourth day), the ovaries were collected, thus, the experiment lasted four days. The ovaries collected in phases 1 and 2 were intended for histological analysis (morphology, growth, activation, extracellular matrix (ECM) configuration and stromal density), immunohistochemistry (TNF-α) and evaluation of mRNA levels for superoxide dismutase (SOD), catalase (CAT), nuclear factor-erythroid 2-related factor 2 (NRF2) and tumor necrosis factor-α (TNF-α) by real-time PCR. Statistical analysis was performed using GraphPad Prism and differences were considered significant when P < 0.05. Phase 1 results showed that ovaries cultured with DOX alone showed a reduction in the percentage of normal follicles and in the density of stromal cells. However, the addition of Aloe vera extract to the medium not only attenuated these structural damages but also increased collagen deposition. Furthermore, ovaries cultured with DOX and Aloe vera (10 and 25%) showed a reduction in TNF-α immunostaining and an increase in mRNA expression for antioxidant enzymes (SOD, CAT and NRF2). The results of phase 2 showed that 0.1 and 1.0 mg/kg of Aloe vera preserved both ovarian follicles and stromal density against DOX-induced damage. In addition, 0.1 and 1.0 mg/kg of Aloe vera reduced TNF-α immunostaining and increased the percentage of collagen fibers. Furthermore, the 1.0 mg/kg dose of Aloe vera increased the mRNA expression for antioxidant enzymes (SOD, CAT and NRF2), while 0.1 mg/kg of Aloe vera maintained these expression levels similar to the saline group. In conclusion, the results obtained indicate that Aloe vera extract, both in vitro and in vivo, demonstrated a protective effect on the ovaries of female mice against damage induced by DOX. This effect was evidenced by increased antioxidant response, reduced inflammation, preservation of the stroma and maintenance of follicular morphology. Tipo: Tese

Remodelação do córtex ovariano durante o cultivo in vitro e efeitos de bioandaimes descelularizados e nanopartículas poliméricas carregadas com resveratrol no crescimento e viabilidade de folículos secundários bovinos.

2025-10-13T20:53:15Z

Título: Remodelação do córtex ovariano durante o cultivo in vitro e efeitos de bioandaimes descelularizados e nanopartículas poliméricas carregadas com resveratrol no crescimento e viabilidade de folículos secundários bovinos. Autor(es): Costa, Francisco das Chagas Tipo: Tese

Estabelecimento de métodos moleculares para detecção de Mycobacterium leprae como ferramenta de diagnóstico para hanseníase

2025-04-15T14:25:23Z

Título: Estabelecimento de métodos moleculares para detecção de Mycobacterium leprae como ferramenta de diagnóstico para hanseníase Autor(es): Albuquerque, Andressa Almeida Abstract: Leprosy is a chronic infectious disease caused by the bacillus Mycobacterium leprae. As it is a pathology that presents a long time for incubation, children diagnosed with leprosy indicate active transmission in the community. Leprosy skin lesions, which can be easily confounded with other diseases, and the subjectivity of dermatoneurological evaluation, reveal the need for laboratory tests that could assist in the diagnosis, making it less subjective and, thus, minimizing risks and challenges related to late diagnosis. The objective of the present study was to establish methods for detecting M. leprae DNA by molecular biology to identify groups at risk for the development of leprosy contacts under 15 years of age and its association with results of dermatoneurological evaluation and with positivity of serum antibodies to the phenolic glycolipid antigen 1 (PGL1), specific to M. leprae. Two types of real-time PCR (qPCR) techniques were validated for detecting M. leprae DNA in blood samples, that is, Sybr green and Taqman®. The commercial NATHANS-Leprosy kit, recently approved for use with skin biopsy samples, was also used to analyze DNA samples extracted from whole blood. 56 index cases and 317 contacts under the age of 15 living in Santana do Ipanema (AL), São Gonçalo do Amarante (CE) and Canindé (CE) were selected. After dermatoneurological evaluation, whole blood samples were collected for detection of M. leprae DNA by qPCR and serum for anti-PGL1 measurement by indirect ELISA. The targets chosen for qPCR were the 16SrRNA and RLEP genes. An RLEP genetic sequence designed from the Primer Express 3.0 Program was also used. The tests for RLEP and 16S rRNA showed analytical sensitivities of 1.57 fg/µL and 97.66 fg/µL. While the tests for RLEP showed analytical specificity to M. leprae DNA, 16S rRNA also showed amplification to DNA from other mycobacteria. Nonetheless, 16S rRNA in Taqman® tests presented specificity to M. leprae DNA. All tests performed as expected for PCR amplification curves. When they were applied to DNA samples extracted from contacts and cases, there was no correlation between serology and molecular tests. Among the molecular tests, the NATHANS-Leprosy kit shows a slight correlation with the Taqman® and Sybr Green tests. In conclusion, in the present work it was possible to establish two molecular methods for Mycobacterium leprae DNA that proved to be sensitive, specific and reproducible using two RLEP sequences and one 16S rRNA sequence. However, they cannot yet be considered validated for use with whole blood samples, as the current study showed a complete discrepancy among the methods. Tipo: Tese