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Background: Polycystic ovarian syndrome (PCOS) with anovulation, hyperandrogenism, ovarian and uterine histological changes, menstrual irregularities, etc. signs is an infertility type. It seems that melatonin and metformin can improve these abnormalities. Objective: To evaluate the effects of melatonin and metformin on the ovary and uterus in PCOS-induced mice using stereological methods. Materials and Methods: Seventy-two adult female BALB/c mice (8-wk-old, 20-30 gr) were randomly divided into control (distilled water, gavage), PCOS (90 μg/kg letrozole, gavage), PCOS+metformin (500 mg/kg, gavage), PCOS+melatonin (10 mg/kg, intraperitoneal injection), and PCOS+melatonin control (0.5% ethanol saline) groups (n = 12/each). Another PCOS group was kept for a month to ensure that PCOS features remained. Finally, a stereological evaluation of the uterus and ovary was carried out, and vaginal cytology and serum testosterone levels were assessed. Results: PCOS mice treated with metformin and melatonin had lower testosterone levels, body weight, and more regular estrus cycles than the PCOS group (p ≤ 0.001). A significant decrease in conglomerate and daughter gland numbers, and primary, secondary, atretic, and cystic follicles numbers with a significant increase in primordial and Graafian follicles, and corpus luteum numbers (p ≤ 0.001) was seen in these treated mice. Also, endometrial vessels’ volume and length significantly increased, but ovarian, endometrial, myometrial, stromal, and glands volume, and endometrial and myometrial thickness dramatically declined (p ≤ 0.001). Conclusion: It appears that metformin and melatonin could restore the PCOS phenotype including estrus cycle irregularity, high testosterone level, and ovarian and uterine micromorphology to the control levels. However, the 2 treatments had similar effects on the examined parameters. Key words: Polycystic ovarian syndrome, Melatonin, Metformin, Ovary, Uterus, Mice, Stereology.

This book provides theoretical and empirical knowledge to researchers using stereological techniques. Most of the commonly used techniques are given in this book. Each of the techniques gives an excellent opportunity to researchers who would like to work with tissues in three dimensions. Accurate and valuable quantitative data on the tissues and organs can be obtained by employing those techniques. This book contains 32 chapters in which the authors present rich data that is analyzed, discussed and interpreted, including morpho-quantitative, histomorphometry and qualitative values.

In order to evaluate the homogeneity of asphalt mixture quantitatively, the distribution characteristic of internal phases of asphalt mixture were identified based on digital image processing technique and stereology theory, and the homogeneity coefficient (i.e., K) was proposed. At the same time, the trend of variation and reliability of homogeneity of asphalt mixture were analyzed by changing the nominal maximum aggregate size, aggregate gradation and asphalt content. The results suggest that the homogeneity of asphalt mixture could be comprehensively described using DIP technique combined with stereology theory. The smaller the K, the better the distribution homogeneity of the asphalt mixture. An improvement in the homogeneity of an asphalt mixture is achieved with the decrease of the nominal maximum aggregate size and a finer aggregate gradation. The asphalt content corresponding to the optimal homogeneity of the internal structure of asphalt mixture specimen is the optimum asphalt content. According to the experimental study, the suggested values of the homogeneity coefficient were given, which provides theoretical support to control the construction quality of the hot mixture asphalt.

Cavalieri estimation is a widely used technique in stereology (applied geometric sampling) for approximating the volume of a solid by sampling cross-sectional areas along a fixed axis. Classical theory shows that, under systematic equidistant sampling (the well-known Cavalieri estimator), the variance decay depends on the smoothness of the area function, which is essentially measured by the number of continuous derivatives. This paper focuses on the natural assumptions under which the theory holds. We first obtain sharp and explicit variance rates: when the Fourier decay is of order s>1/2, the variance of the Cavalieri estimator decays as t2s with a constant independent of t. Building on this, we show that the smoothness condition expressed in terms of the algebraic Fourier decay subsumes both integer- and fractional-order frameworks used to date. Finally, we establish a matching converse showing that, under general assumptions, no broader smoothness framework extends the theory; that is, any algebraic variance decay implies the corresponding Fourier decay.

Remarkably little is known about the postnatal cellular development of the human amygdala. It plays a central role in mediating emotional behavior and has an unusually protracted development well into adulthood, increasing in size by 40% from youth to adulthood. Variation from this typical neurodevelopmental trajectory could have profound implications on normal emotional development. We report the results of a stereological analysis of the number of neurons in amygdala nuclei of 52 human brains ranging from 2 to 48 years of age [24 neurotypical and 28 autism spectrum disorder (ASD)]. In neurotypical development, the number of mature neurons in the basal and accessory basal nuclei increases from childhood to adulthood, coinciding with a decrease of immature neurons within the paralaminar nucleus. Individuals with ASD, in contrast, show an initial excess of amygdala neurons during childhood, followed by a reduction in adulthood across nuclei. We propose that there is a long-term contribution of mature neurons from the paralaminar nucleus to other nuclei of the neurotypical human amygdala and that this growth trajectory may be altered in ASD, potentially underlying the volumetric changes detected in ASD and other neurodevelopmental or neuropsychiatric disorders.

Background Bronchopulmonary dysplasia (BPD) is a chronic lung condition of premature neonates, yet without an established pharmacological treatment. The BPD rabbit model exposed to 95% oxygen has been used in recent years for drug testing. However, the toxicity of the strong hyperoxic hit precludes a longer-term follow-up due to high mortality after the first week of life. This study aimed to extend the preterm rabbit model to postnatal day (PND) 14 to mimic the evolving phase of BPD and enable the investigation of therapeutic interventions at later and more relevant time points. Methods Preterm rabbit pups delivered on the 28th day of gestation were either exposed to room air or different degrees of hyperoxia (50% and 70% O 2 ) for 14 days. Single (immediately after birth) or double (at birth and at PND5) intratracheal lipopolysaccharide (LPS) administrations were also tested in combination with 50% O 2 . Age-matched rabbits delivered vaginally at term were used as controls. Survival, weight gain, lung function, pulmonary artery micro-ultrasound Doppler analysis, lung histology (alveolarization, lung injury score, and design-based stereology), and longitudinal micro-CT imaging were used to compare the outcomes at PND14. Results Premature birth itself, without any other BPD hit, was associated with lung function deficits, delayed lung development, and cardiovascular abnormalities. The BPD-like lung phenotype was enhanced by 70% O 2 but not by 50% O 2 hyperoxia. Intratracheal LPS delivered immediately after birth was associated with significantly higher lung injury scores at PND14 and increased tissue damping, a marker of parenchymal air resistance. Conclusion Several strategies are feasible to extend the preterm rabbit model of BPD to PND14. Preterm birth at the saccular phase itself, even in the absence of other postnatal BPD hits, was associated with lung function deficits, delayed lung development, and cardiovascular abnormalities compared with age-matched term rabbit pups. Enhanced BPD-like phenotypes can be further achieved by continued exposure to moderate hyperoxia (70% O 2 ) and the intratracheal administration of LPS.

Ethanolic extract of bitter fraction of Stevia rebaudiana ( Srbf ) was extracted to investigate its antihyperglycemic and protective effects on renal structural changes in STZ-induced diabetes. Thirty-five male mice were divided into five groups randomly; the first group as non-diabetic control, the second group as untreated diabetic, the third group treated with glibenclamide 0.5 mg/kg, and the fourth and fifth groups treated with Srbf by 200 and 400 μg/kg bw through gavage, respectively, for 15 days. Diabetes was induced in the second to fifth groups by administration of 60 mg/kg bw of streptozotocin intraperitoneally. Serum glucose level was monitored every day. At the 16th day, the subjects were sacrificed and their left kidneys were removed. Tissue sections were stained by periodic acid Schiff and used for stereological analysis. The means were compared by one-way ANOVA and Tukey’s post hoc test at the significance level of p  ≤ 0.05. The results showed that Srbf significantly restored the blood glucose level toward normal level faster than glibenclamide. High dose of Srbf could significantly decrease the length and volume of proximal and distal tubules and vessels and the volume of the interstitial tissue in the diabetic treated group. Both doses of Srbf could significantly prevent the glomerular hypertrophy and reduction of glomerular number in comparison with the untreated diabetic group. It can be concluded that the antihyperglycemic properties of a bitter fraction of S. rebaudiana are better than glibenclamide, and at high dose, it can ameliorate structural nephropathy in diabetic mice.

Neurogenesis within the dentate gyrus is thought to play an important role in cognitive processes such as reversal learning and pattern separation. The α7 nicotinic acetylcholine receptor (α7 nAChR) is expressed early in newly formed granule cells of the dentate gyrus, though its role in neurogenesis and related cognitive function is not fully understood.IntroductionNeurogenesis within the dentate gyrus is thought to play an important role in cognitive processes such as reversal learning and pattern separation. The α7 nicotinic acetylcholine receptor (α7 nAChR) is expressed early in newly formed granule cells of the dentate gyrus, though its role in neurogenesis and related cognitive function is not fully understood.To better characterize relevant function of α7 nAChRs, we performed unbiased stereology to quantify hippocampal granule cells, pyramidal cells, and total volume and used a touchscreen operant spatial discrimination/reversal task to test pattern separation in a global α7 nAChR knockout mouse line.MethodsTo better characterize relevant function of α7 nAChRs, we performed unbiased stereology to quantify hippocampal granule cells, pyramidal cells, and total volume and used a touchscreen operant spatial discrimination/reversal task to test pattern separation in a global α7 nAChR knockout mouse line.The knockout resulted in an ≈22% reduction in granule cells and a ≈ 20% reduction in pyramidal cells in both sexes, with no change in total hippocampal volume. However, the knockout impaired performance in the touchscreen task for males only. The sex-dependent difference in behavioral, but not stereological, results suggest a divergence in the structure-function relationship in males versus females. Detailed analyses revealed males were more biased by the initial reversal contingency relative to females indicating a potential source of the sex-specific interaction with the loss of α7 nAChRs.ResultsThe knockout resulted in an ≈22% reduction in granule cells and a ≈ 20% reduction in pyramidal cells in both sexes, with no change in total hippocampal volume. However, the knockout impaired performance in the touchscreen task for males only. The sex-dependent difference in behavioral, but not stereological, results suggest a divergence in the structure-function relationship in males versus females. Detailed analyses revealed males were more biased by the initial reversal contingency relative to females indicating a potential source of the sex-specific interaction with the loss of α7 nAChRs.These findings argue that the α7 nAChR plays a critical role in hippocampal development, not just granule cell neurogenesis, and plays a sex-dependent role in cognitive function.DiscussionThese findings argue that the α7 nAChR plays a critical role in hippocampal development, not just granule cell neurogenesis, and plays a sex-dependent role in cognitive function.

Bronchopulmonary dysplasia (BPD), the most common sequela of preterm birth, is a severe disorder of the lung that is often associated with long-lasting morbidity. A hallmark of BPD is the disruption of alveolarization whose pathogenesis is incompletely understood. Here, we tested the vascular hypothesis that disordered vascular development precedes the decreased alveolarization associated with BPD. Neonatal mouse pups were exposed to 7, 14 or 21 days of normoxia (21% O2) or hyperoxia (85% O2) with n=8-11 for each group. The right lungs were fixed by vascular perfusion and investigated by design-based stereology or three-dimensional (3D) reconstruction of data sets obtained by serial block-face scanning electron microscopy. The alveolar capillary network of hyperoxia-exposed mice was characterized by rarefaction, partially altered geometry and widening of capillary segments as shown by 3D reconstruction. Stereology revealed that the development of alveolar epithelium and capillary endothelium was decreased in hyperoxia-exposed mice, however, the time course of these effects was different. That the surface area of the alveolar epithelium was smaller in hyperoxia-exposed mice first became evident at day 14.In contrast, the surface area of the endothelium was reduced in hyperoxia-exposed mouse pups at day 7. The thickness of the air-blood barrier decreased during postnatal development in normoxic mice whereas it increased in hyperoxic mice. The endothelium and the septal connective tissue made appreciable contributions to the thickened septa. In conclusion, the present study provides clear support for the idea that the stunted alveolarization follows the disordered microvascular development, thus supporting the vascular hypothesis of BPD.

The estimation of the number or density of neurons and types of glial cells and their relative proportions in different brain areas are at the core of rigorous quantitative neuroanatomical studies. Unfortunately, the lack of detailed, updated, systematic and well-illustrated descriptions of the cytology of neurons and glial cell types, especially in the primate brain, makes such studies especially demanding, often limiting their scope and broad use. Here, following an extensive analysis of histological materials and the review of current and classical literature, we compile a list of precise morphological criteria that can facilitate and standardize identification of cells in stained sections examined under the microscope. We describe systematically and in detail the cytological features of neurons and glial cell types in the cerebral cortex of the macaque monkey and the human using semithin and thick sections stained for Nissl. We used this classical staining technique because it labels all cells in the brain in distinct ways. In addition, we corroborate key distinguishing characteristics of different cell types in sections immunolabeled for specific markers counterstained for Nissl and in ultrathin sections processed for electron microscopy. Finally, we summarize the core features that distinguish each cell type in easy-to-use tables and sketches, and structure these key features in an algorithm that can be used to systematically distinguish cellular types in the cerebral cortex. Moreover, we report high inter-observer algorithm reliability, which is a crucial test for obtaining consistent and reproducible cell counts in unbiased stereological studies. This protocol establishes a consistent framework that can be used to reliably identify and quantify cells in the cerebral cortex of primates as well as other mammalian species in health and disease.