Quantitative MRI evaluation of bone marrow in sickle cell disease: relationship with haemolysis and clinical severity.

AIM: To evaluate bone marrow fat fraction using the Dixon technique (FFDix) of magnetic resonance imaging (MRI) as a potential biomarker of haemolysis and clinical severity in the overall assessment and follow-up of sickle cell disease (SCD) patients. MATERIAL AND METHODS: The present study was a cross-sectional study in which healthy individuals and SCD patients (matched for age, sex, and weight) were subjected to MRI of the lumbar spine and pelvis to quantify FFDix in the bone marrow using the Dixon technique. SCD severity was analysed by clinical and laboratory data, and an online calculator. A high degree of haemolysis was defined using the cut-off values haemoglobin (Hb) ≤10 g/dl, lactate dehydrogenase (LDH) ≥325 U/l, reticulocytes ≥3% and total bilirubin (TB) ≥1.2 mg/dl. Pearson's correlation, receiver operating characteristic (ROC) curve and binary logistic regression analysis were performed. RESULTS: Forty-eight SCD patients (26 homozygous: HbSS and 22 compound heterozygous: HbSC) and 48 healthy individuals participated in the study. FFDix was lower in SCD patients than in the control group, showing even lower values in the HbSS subtype and patients with a higher degree of haemolysis. HbSC patients with a higher degree of haemolysis using hydroxyurea (medium dosage 9.8 mg/kg/day) had lower FFDix. ROC curves and odds ratios for detecting patients with a higher degree of haemolysis at the different FFDix measurement sites demonstrated excellent performance: iliac bones (cut-off ≤16.75%, AUC = 0.824, p<0.001), femoral heads (cut-off ≤46.7%, AUC = 0.775, p=0.001), lumbar vertebrae (cut-off ≤7.8%, AUC = 0.755, p=0.002). CONCLUSION: Decreased FFDix is indicative of higher degree of haemolysis and SCD severity with great potential as a non-invasive biomarker contributing to the overall assessment and follow-up of SCD patients.

Mesh excision secondary to spondylodiscitis after colposacropexy graft rejection: a step by step video.

INTRODUCTION: Spondylodiscitis secondary to colposacropexy is an extremely rare entity. Infection and mesh rejection are the main causes. Removal of the mesh is essential for patient's recovery and it can be a very challenging surgical procedure. CASE: A 72-year-old woman presented with severe low back pain in the context of a recent colposacropexy. Magnetic resonance imaging was performed and spondylodiscitis secondary to prolapse correction surgery with mesh was suspected. In order to ensure an adequate recovery, removal of the mesh was required. CONCLUSIONS: Spondylodiscitis secondary to colposacropexy should be suspected when the patient starts with moderate lumbar pain and is not correctly controlled with first-level analgesia. Infection or mesh rejection should be considered. Mesh rejection should be suspected when the patient does not improve after antibiotics. Complete removal of the mesh is needed in order to ensure the patient's recovery.

Echoes of shame: a comparison of the characteristics and psychological sequelae of recalled shame experiences across the voice hearing continuum.

BACKGROUND: Voice hearing occurs across a number of psychiatric diagnoses and appears to be present on a continuum within the general population. Previous research has highlighted the potential role of past experiences of shame in proneness to voice hearing in the general population. AIMS: This study aimed to extend this past research and compare people with distressing voices, people with voices but no distress, and a non-voice hearing control group, on various dimensions of shame and shame memory characteristics. METHOD: In a cross-sectional, online study 39 distressed voice hearers, 31 non-distressed voice hearers and 50 non-voice hearers undertook a shame memory priming task in which they were prompted to recall a memory of a shaming experience from their past. They then completed questionnaires assessing the characteristics of the recalled shame event and the psychological sequalae of this event (i.e. intrusions, hyperarousal, avoidance, the centrality of shame memories, external shame, and self-criticism). RESULTS: The majority of recalled shame memories involved experiences such as interpersonal criticism or experiences of being devalued. Univariate analyses found no significant differences between the three groups with regard to the shame events that were recalled, but the distressed voice hearer group reported significantly more hyperarousal, intrusions, self-criticism, and external shame in relation to their experience. CONCLUSIONS: The findings suggest that voice hearers recall similar types of shame experiences to non-voice hearers, but that problematic psychological sequelae of these shame experiences (in the form of intrusive memories, hyperarousal, external shame, and self-criticism) may specifically contribute to distressing voice hearing.

One stage procedure for developmental dysplasia of the hip in patients aged eight years or older. Systematic review and meta-analysis.

The objective of the present study is to perform a systematic review with meta-analysis of the literature on treatment of developmental dysplasia of the hip (DDH) in patients over eight years of age to provide better understanding of therapeutic strategies and results. Authors carried out a systematic review and metanalysis of the literature on DDH treated in patients aged eight years of age or older. A literature search was carefully performed from June 2019 to June 2020. The articles had to report one stage reconstructive surgical treatment of DDH for patients eight years or older, presenting clinical and radiographic evaluation according to the Tonnis and Severin, and McKay systems. Meta-analysis was carried out using the software metanalyst to perform the pooled effect size Nine studies met the inclusion criteria. They assessed a total of 234 patients and 266 hips. Female patients accounted for 75.7% (eight unknown) and the follow-up ranged from 1 to 17.4 years. The majority of the procedures included an acetabular surgery (93.9%) while femoral shortening was performed in 78%. Acceptable outcomes were found in between 67% (Mckay system) and 91% (Severin system) of the cases. Femoral varus and derrotation shortening associated with redirectional osteotomy of the acetabulum (for those with closed triradiate cartilage) or acetabular redirection/reshaping were the most prevalent combined procedures; this strategy can lead to 60% of acceptable clinical results and 90% of radiographically acceptable results. Therefore, our findings give credence to the recommendation for the treatment of DDH in patients over eight years old.

Fibroblast growth factor-2 promotes in vitro activation of cat primordial follicles.

This study evaluated the effect of fibroblast growth factor-2 (FGF-2) on the morphology, primordial follicle activation and growth after in vitro culture of domestic cat ovarian tissue. Ovaries (n = 12) from prepubertal domestic cats were collected and fragmented. One fragment was fixed for histological analysis (fresh control). The remaining fragments were incubated in control medium alone or with 10, 50 or 100 ng/ml FGF-2 for 7 days. After in vitro culture, the following endpoints were analyzed: morphology, activation by counting primordial and developing follicles, and growth (follicle and oocyte diameters). Treatment with 100 ng/ml FGF-2 maintained (P > 0.05) the percentage of normal follicles similar to fresh control. Follicle survival was greater (P < 0.05) after culture in 100 ng/ml FGF-2 than in 50 ng/ml FGF-2. The percentage of primordial follicles decreased (P < 0.05) and the percentage of developing follicles increased (P < 0.05) in all treatments compared with fresh tissue. The proportion of developing follicles increased (P < 0.05) in tissues incubated with 100 ng/ml FGF-2 compared with control medium and other FGF-2 concentrations. Furthermore, culture in 10 or 100 ng/ml FGF-2 resulted in increased (P < 0.05) follicle and oocyte diameters compared with fresh tissues and MEM+. In conclusion, FGF-2 at 100 ng/ml maintains follicle survival and promotes the in vitro activation and growth of cat primordial follicles.

Leptin decreases apoptosis and promotes the activation of primordial follicles through the phosphatidylinositol-3-kinase/protein kinase B pathway in cultured ovine ovarian tissue.

This study evaluated the effects of leptin on primordial follicle survival and activation after in vitro culture of ovine ovarian tissue and if leptin acts through the phosphatidylinositol-3-kinase/protein kinase B (PI3K/Akt) pathway. Ovarian fragments were fixed for histology (fresh control) or cultured for 7 days in control medium (α-MEM+) alone or supplemented with leptin (1, 5, 10, 25 or 50 ng/ml). Follicle morphology, activation and apoptosis were analyzed. Next, the fragments were cultured in the medium that showed the best results in the absence or the presence of the PI3K inhibitor (LY294002), and immunohistostaining of p-Akt protein was assessed. After culture, the percentage of normal follicles decreased (P < 0.05) in all treatments compared with the fresh control. Moreover, control medium and 1 ng/ml leptin had similar (P > 0.05) percentages of normal follicles, which were significantly higher than those in other treatments. However, culture with 1 ng/ml leptin maintained apoptosis similarly (P > 0.05) to that of the fresh control and lower (P < 0.05) than that in α-MEM+. Leptin did not influence follicle activation (P > 0.05) compared with the control medium (α-MEM+). Culture in 1 ng/ml leptin with LY294002 decreased the normal follicles and increased apoptosis, inhibited follicle activation (P < 0.05), and reduced p-Akt immunostaining, compared with the medium containing 1 ng/ml leptin without PI3K inhibitor. In conclusion, leptin at 1 ng/ml reduces apoptosis and promotes the activation of primordial follicles compared with the fresh control after in vitro culture of ovine ovarian tissue possibly through the PI3K/Akt pathway.

Angiotensin II receptor agonist antibodies are associated with microvascular damage in lupus nephritis.

Angiotensin II type 1 receptor agonist antibodies (AT1R-AAs) have been associated with hypertension, atherosclerosis and vascular inflammation in human diseases. The aim of the study was to evaluate the prevalence of AT1R-AAs in active lupus nephritis (LN) patients and their association with vascular damage. One hundred and seven active LN patients underwent a complete clinical examination, measurement of AT1R-AAs, ambulatory blood pressure monitoring, carotid intima-media thickness measurement and morphometric analysis of subintimal fibrosis and medial hyperplasia of the vessels in the kidney tissue. Plasma AT1R-AAs were positive in 58 (54.2%) patients. The Systemic Lupus Erythematosus Disease Activity Index 2000 (SLEDAI-2K) score, complement C3 and C4 levels and titers of anti-dsDNA antibodies were higher in the group with positive AT1R-AAs compared with those with negative AT1R-AAs. The AT1R-AA titers correlated with anti-dsDNA antibody titers and with complement C3 and C4 serum levels. In the kidney biopsy, the percentage of subintimal fibrosis and the area of medial hyperplasia were greater in the AT1R-AA-positive patients. No differences in arterial pressure, carotid intima-media thickness and response to therapy were detected. In conclusion, AT1R-AAs are prevalent in active LN patients and are associated with histologic features of microvascular damage.

MRI findings of the cervical spine in patients with mucopolysaccharidosis type VI: relationship with neurological physical examination.

AIM: To describe the magnetic resonance imaging (MRI) findings of the cervical spine of patients with mucopolysaccharidosis type VI (MPS VI) and correlate them with clinical manifestations. MATERIALS AND METHODS: This is a cross-sectional study involving 12 patients with MPS VI. A limited neurological examination was undertaken in each patient including Tinel's test, assessment of muscle tone, and the evaluation of deep tendon reflexes. Additionally, each patient underwent cervical spine MRI to evaluate platybasia, odontoid dysplasia, periodontoid soft-tissue thickening, spinal canal stenosis, myelopathy, basilar invagination, platyspondyly, and reduction of nasopharyngeal airway. RESULTS: Nine patients were male (75%). The average age was 12.5 (±3.5 years). Tinel's test was negative in all patients. No muscle tone abnormalities were observed. Approximately 48% of the tested reflexes were considered abnormal, 10 of which (8.3%) were pathological occurring in five different patients (41.6%). At MRI, all patients showed periodontoid soft-tissue thickening and cervical spinal stenosis; six showed spinal cord compression and two showed myelopathy. Odontoid hypoplasia and basilar invagination were observed in nine patients. All patients with cervical stenosis on MRI had abnormal reflexes; however, only two of the six patients with evidence of cord compression on MRI had abnormal reflexes on clinical examination. CONCLUSIONS: The present study of 12 patients with MPS VI demonstrated that a normal neurological examination cannot confidently exclude potential cord compression in patients with this condition. MRI may aid in the timely identification of cervical spine abnormalities, and potentially play a role in lessening morbidity and mortality in patients with MPS.

Kaempferol promotes primordial follicle activation through the phosphatidylinositol 3-kinase/protein kinase B signaling pathway and reduces DNA fragmentation of sheep preantral follicles cultured in vitro.

The aim of this study was to evaluate the effects of kaempferol on the morphology, follicular activation, growth, and DNA fragmentation of ovine preantral follicles cultured in situ, and the effects of a phosphatidylinositol 3 kinase (PI3K) inhibitor and the expression of phosphorylated protein kinase B (pAKT) after culture. Ovine ovarian fragments were fixed for histological and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) analyses (fresh control) or cultured in α-MEM+ alone (control) or with different concentrations of kaempferol (0.1, 1, 10, or 100 µM) for 7 days. Follicles were classified as normal or atretic, primordial or growing, and the oocyte and follicle diameters were measured. Proliferating cells were analyzed and DNA fragmentation was evaluated by the TUNEL assay. Inhibition of PI3K activity was performed through pretreatment in media added with 50 µM LY294002 for 1 hr and pAKT immunohistochemistry was performed after culture in the absence or presence of LY294002. After culture, the percentage of normal follicles was similar among the treatments (p > 0.05), except for 100 µM kaempferol, which had less normal follicles (p < 0.05). Moreover, kaempferol at 10 µM showed a higher percentage of follicular activation and cell proliferation than the other treatments (p < 0.05) and a percentage of TUNEL-positive cells similar to that in the fresh control and lower than other treatments (p < 0.05). LY294002 significantly inhibited primordial follicle activation stimulated by α-MEM+ and 10 µM kaempferol and reduced pAKT expression in those follicles. In conclusion, 10 µM kaempferol promotes primordial follicle activation and cell proliferation through the PI3K/AKT pathway and reduces DNA fragmentation of ovine preantral follicles cultured in vitro.

Self-rated health and health-related quality of life are related with adolescents' healthy lifestyle.

OBJECTIVES: Self-rated health (SRH) and health-related quality of life (HRQoL) are related with health outcomes. It is expected that these constructs are also associated with healthy behaviours. Thus, the present study sought to analyse the associations between a healthy lifestyle composite score and SRH and HRQoL among adolescents. STUDY DESIGN: This was an observational, cross-sectional study. METHODS: Data were obtained from the cross-sectional health behaviour in school-aged children (HBSC) 2014 Portuguese survey. Participants were 5024 adolescents (2373 boys), aged 10-17 years. A composite score of healthy lifestyle was obtained using the combination of the following factors: daily physical activity, sleep at least 8-9 h/night, daily consumption of fruit and vegetables, spend <2 h daily in screen-based behaviours, never drink and never smoke. RESULTS: Adolescents with a healthy lifestyle had significantly better SRH and higher HRQoL than those who were not engaged in all healthy behaviours. CONCLUSIONS: Healthy lifestyle (composed of physical activity, sleep at least 8-9 h/night, daily consumption of fruit and vegetables, spend <2 h daily in screen-based behaviours, never drink and never smoke) is related to SRH and HRQoL. Promoting a comprehensive approach to these behaviours is a public health strategy to be considered when aiming to improve health, in general, and SRH and HRQoL, in particular.

Quercetin influences in vitro maturation, apoptosis and metabolically active mitochondria of goat oocytes.

SummaryThe present study aimed to investigate the effect of quercetin as an alternative antioxidant to cysteamine on in vitro maturation. Oocytes were collected from goat ovaries, destined for in vitro maturation and distributed into three groups: CIS group, oocytes were immersed in MIV base medium; in Groups Q4 and Q8, oocytes were immersed in the medium of the CIS group, adding 4 µM or 8 µM of quercetin, respectively, and cultured for 24 h at 38.5°C with 5% CO2. The CIS and Q4 groups presented the same percentage of expanded cumulus cells, but the per cent in the Q8 group was significantly lower than that of the other groups (P<0.05). The oocyte retraction rate in the Q8 group was higher (P<0.05) than in the CIS and Q4 groups. Treatment with 8 µM of quercetin presented a lower proportion of expanded oocytes than the CIS group and 4 µM of quercetin (P<0.05). The percentage of MII oocytes was higher in the Q4 group than in the CIS group (P<0.05), but the percentages in the CIS and Q8 groups were similar. The rate of apoptosis was higher in the CIS group than in the other groups (P<0.05). In addition, oocytes matured with 4 µM quercetin showed higher mitochondrial activity than matured oocytes in the CIS and Q8 groups (P<0.05). In conclusion, 4 µM of quercetin can be used as an alternative to cysteamine in the in vitro maturation of goat oocytes.

Non-invasive joint decompression: An important factor in the regeneration of the bone marrow and disc recapture in temporomandibular arthropathies.

BACKGROUND: This article aims to demonstrate the importance of the TMJ (Temporomandibular Joint) decompression in the treatment of degenerative processes and disc displacements, reporting two clinical cases treated with orthopedic and decompressive correction of TMJ. MATERIAL AND METHODS: The studies reported in this article show patients with muscle and joint pain who were evaluated pre and post-treatment through MRI (Magnetic Resonance Irradiation) to follow-up bone marrow regeneration and TMJ disc placement. Transcutaneous electrical stimulation (TENS), measurement equipment and IO (Intraoral Orthotic) were used to evaluate and treat the patients. A critical review of literature has also been conducted to confront clinical outcomes. RESULTS: Marrow bone regeneration and disc placement were observed in both patients. CONCLUSIONS: The use of measurement equipment associated with TENS to find the correct rest position of the Jaw an the use of IO to decompress the TMJ was an effective way to promote bone marrow regeneration and disc placement, consequently improving function and quality of life.

Experimental and Computational Thermochemical Study of Maleic Anhydride and Vinylene Carbonate.

The standard molar enthalpies of formation of maleic anhydride and vinylene carbonate in gaseous phase, at T = 298.15 K, were derived from the standard molar enthalpies of formation of the compounds in condensed phase combined with the phase transition enthalpies. The standard molar enthalpies of formation in condensed phase were obtained from the enthalpies of combustion measured using static bomb combustion calorimetry and mini-bomb combustion calorimetry for vinylene carbonate and maleic anhydride, respectively. Phase transition enthalpies were obtained by Calvet microcalorimetry. High level quantum calculations were performed at the composite G3 level of theory in order to estimate the standard molar enthalpies of formation of both compounds in gaseous phase. Good agreement was obtained between experimental and computational results. In addition, analysis of the factors affecting the relative stability of both systems has been carried out in the framework of the ab initio valence bond (VB) theory in order to clarify the aromaticity/antiaromaticity issues involving these molecular systems.

Resveratrol has dose-dependent effects on DNA fragmentation and mitochondrial activity of ovine secondary follicles cultured in vitro.

The worldwide consumption of red wine, nuts and grapes has resulted in increased human exposure to resveratrol, which could affect reproductive function. However, the effect of resveratrol on in vitro culture of early-stage ovarian follicles has never been investigated. The aims of the present study were to evaluate the effect of resveratrol on sheep secondary follicle morphology, growth, DNA fragmentation, intracellular levels of glutathione (GSH) and active mitochondria. Secondary follicles were isolated from the ovaries and cultured for 18 days in supplemented α-MEM+ (control medium) or in control medium containing resveratrol (2, 10 or 30 µM). The parameters analyzed were morphology, antrum formation, follicle diameter, DNA fragmentation, GSH levels and mitochondrial activity. After 18 days, all resveratrol groups significantly decreased the percentages of morphologically normal follicles compared with the control group (α-MEM+). Antrum formation was higher in both α-MEM+ and 2 µM resveratrol groups than in the 10 µM resveratrol group. In addition, 30 µM resveratrol increased the percentage of oocytes with DNA damage compared with the control. Oocytes from follicles treated with 10 or 30 µM resveratrol significantly decreased intracellular GSH levels compared with the 2 µM resveratrol group. Moreover, follicles in α-MEM+ (control) showed more active mitochondria than those in 10 or 30 µM resveratrol. In conclusion, ovine isolated secondary follicles are able to grow to the antral stage after in vitro culture in medium containing 2 µM resveratrol, maintaining the same rates of DNA damage, GSH levels and mitochondrial function as the control medium. However, the addition of 30 µM resveratrol increased DNA fragmentation and oxidative stress through decreasing mitochondrial activity.

Factors affecting the well-being of adolescents living with unemployed parents in times of economic recession: findings from the Portuguese HBSC study.

OBJECTIVES: Using a national representative sample of adolescents with unemployed parents, this study examined which factors (sociodemographic and of satisfaction with family life and peers) are related to the negative effect of parents' unemployment on emotional well-being. STUDY DESIGN: Cross-sectional survey study. METHODS: Data on adolescents (14.1 ± 1.7 years old), with at least one parent unemployed (n = 1311, 53.2% girls), was provided by the Portuguese Health Behaviour in School-aged Children study. Descriptive statistics and logistic regression analyses were performed. RESULTS: A high proportion of adolescents reported being emotionally affected by father's unemployment but not by their mother's. Older boys and older girls were more likely to report that their same-gender parent's unemployment situation (sons-fathers and daughters-mothers) has had a negative effect on their well-being. Girls from low socio-economic status and with poor family satisfaction were more likely to report negative emotional well-being related to parental unemployment. CONCLUSIONS: This study presents evidence on factors that can shape adolescents' emotional well-being related to parents' unemployment situation. Given the recessionary context and high unemployment rates, these insights are valuable to assist the design of an action to improve the levels of well-being of Portuguese adolescents from unemployed families.

Evidence for Gamow-Teller Decay of

The ß-delayed neutron emission of ^{83,84}Ga isotopes was studied using the neutron time-of-flight technique. The measured neutron energy spectra showed emission from states at excitation energies high above the neutron separation energy and previously not observed in the ß decay of midmass nuclei. The large decay strength deduced from the observed intense neutron emission is a signature of Gamow-Teller transformation. This observation was interpreted as evidence for allowed ß decay to ^{78}Ni core-excited states in ^{83,84}Ge favored by shell effects. We developed shell model calculations in the proton fpg_{9/2} and neutron extended fpg_{9/2}+d_{5/2} valence space using realistic interactions that were used to understand measured ß-decay lifetimes. We conclude that enhanced, concentrated ß-decay strength for neutron-unbound states may be common for very neutron-rich nuclei. This leads to intense ß-delayed high-energy neutron and strong multineutron emission probabilities that in turn affect astrophysical nucleosynthesis models.

Tracking changes in chromosomal arrangements and their genetic content during adaptation.

There is considerable evidence for an adaptive role of inversions, but how their genetic content evolves and affects the subsequent evolution of chromosomal polymorphism remains controversial. Here, we track how life-history traits, chromosomal arrangements and 22 microsatellites, within and outside inversions, change in three replicated populations of Drosophila subobscura for 30 generations of laboratory evolution since founding from the wild. The dynamics of fitness-related traits indicated adaptation to the new environment concomitant with directional evolution of chromosomal polymorphism. Evidence of selective changes in frequency of inversions was obtained for seven of 23 chromosomal arrangements, corroborating a role for inversions in adaptation. The evolution of linkage disequilibrium between some microsatellites and chromosomes suggested that adaptive changes in arrangements involved changes in their genetic content. Several microsatellite alleles increased in frequency more than expected by drift in targeted inversions in all replicate populations. In particular, there were signs of selection in the O3+4 arrangement favouring a combination of alleles in two loci linked to the inversion and changing along with it, although the lack of linkage disequilibrium between these loci precludes epistatic selection. Seven other alleles increased in frequency within inversions more than expected by drift, but were not in linkage disequilibrium with them. Possibly these alleles were hitchhiking along with alleles under selection that were not specific to those inversions. Overall, the selection detected on the genetic content of inversions, despite limited coverage of the genome, suggests that genetic changes within inversions play an important role in adaptation.

Kit ligand promotes the transition from primordial to primary follicles after in vitro culture of ovine ovarian tissue.

This study evaluated the effects of kit ligand (KL) on the morphology and development of ovine preantral follicles (fresh control) and after 7 days of in vitro culture in α-Minimal Essential Medium (α-MEM; control medium) or the presence of KL (1, 10, 50, 100 or 200 ng/ml). There was an increase in the percentage of primary follicles at the concentration of 100 ng/ml KL, compared with the fresh control, control medium (α-MEM) and the other KL concentrations. Follicle diameter was significantly higher than the control medium only at concentrations of 50 and 100 ng/ml KL. In conclusion, 100 ng/ml KL promoted the transition from primordial to primary follicles (follicular activation) after in vitro culture of ovine ovarian tissue.

Expression of angiotensin II receptors in the caprine ovary and improvement of follicular viability in vitro.

This study aimed to evaluate mRNA levels of angiotensin II (ANG II) receptors (AGTR1 and AGTR2) in caprine follicles and to investigate the influence of ANG II on the viability and in vitro growth of preantral follicles. Real-time polymerase chain reaction (PCR) was used to quantify AGTR1 and AGTR2 mRNA levels in the different follicular stages. For culture, caprine ovaries were collected, cut into 13 fragments and then either directly fixed for histological and ultrastructural analysis (fresh control) or placed in culture for 1 or 7 days in α-minumum essential medium plus (α-MEM+) with 0, 1, 5, 10, 50 or 100 ng/ml ANG II. Then, the fragments were destined to morphological, viability and ultrastructural analysis. The results showed that primordial follicles had higher levels of AGTR1 and AGTR2 mRNA than secondary follicles. Granulosa/theca cells from antral follicles had higher levels of AGTR1 mRNA than their respective cumulus-oocyte complex (COCs). After 7 days of culture, ANG II (10 or 50 ng/ml) maintained the percentages of normal follicles compared with α-MEM+. Fluorescence and ultrastructural microscopy confirmed follicular integrity in ANG II (10 ng/ml). In conclusion, a high expression of AGTR1 and AGTR2 is observed in primordial follicles. Granulosa/theca cells from antral follicles had higher levels of AGTR1 mRNA. Finally, 10 ng/ml ANG II maintained the viability of caprine preantral follicles after in vitro culture.

Amburana cearensis leaf extract maintains survival and promotes in vitro development of ovine secondary follicles.

The antioxidant properties of Amburana cearensis extract may be a useful substitute for standard cell culture medium. Thus, the aim of this study was to evaluate the effect of this extract, with or without supplementation, on in vitro survival and development of sheep isolated secondary follicles. After collection of the ovaries, secondary follicles were isolated and cultured for 18 days in α-MEM+ supplemented with bovine serum albumin, insulin, transferrin, selenium, glutamine, hypoxanthine and ascorbic acid (control medium) or into medium composed of different concentrations of A. cearensis extract without supplements (Amb 0.1; 0.2 or 0.4 mg/ml) or A. cearensis extract supplemented with the same substances described above for α-MEM+ supplementation. The A. cearensis supplemented medium was named Amb 0.1+; 0.2+ or 0.4+ mg/ml. There were more morphologically normal follicles in Amb 0.1 or Amb 0.4 mg/ml than in the control medium (α-MEM+) after 18 days of culture. Moreover, the percentage of antrum formation was significantly higher in Amb 0.1 or Amb 0.2 mg/ml than in α-MEM+ and Amb 0.1+ mg/ml, and similar to the other treatments. All A. cearensis extract media induced a progressive and significant increase in follicular diameter throughout the culture period. In conclusion, this study showed that 0.1 mg/ml of this extract, without supplementation, maintains follicular survival and promotes the development of ovine isolated secondary follicles in vitro. This extract can be an alternative culture medium for preantral follicle development.