143. DIFFERENCES IN GENE EXPRESSION BETWEEN APICAL AND BASAL CELLS OF THE MEMBRANA GRANULOSA

2010 ◽  
Vol 22 (9) ◽  
pp. 61
Author(s):  
H. F. Irving-Rodgers ◽  
S. T. Lee ◽  
N. Hatzirodos ◽  
K. Hummitzsch ◽  
T. R. Sullivan ◽  
...  
Keyword(s):  
Basal Lamina ◽  
Granulosa Cells ◽  
Single Layer ◽  
Cumulus Cells ◽  
Endocrine Function ◽  
Follicular Epithelium ◽  
Basal Cells ◽  
Expression Of Genes ◽  
Apical Cells ◽  
Follicle Maturation

Granulosa cells constitute the ovarian follicular epithelium which at the beginning of folliculogenesis forms a single layer of flattened cells. As the follicle matures the cells acquire a cuboidal morphology, proliferate and differentiate into the cumulus cells surrounding the oocyte, and the mural granulosa cells forming the inner layer of the follicle (the membrana granulosa). Mural granulosa cells may further differ in their functionality depending on whether they are situated apically or basally within the stratified membrana granulosa. Late in folliculogenesis granulosa cells develop the ability to produce oestradiol, and also a specialised extracellular matrix (focimatrix) which is more abundant between apical cells. In order to investigate possible differences between granulosa cells, the expression of genes for oestradiol synthesis (CYP11A1, CYP19A1), focimatrix components (LAMB2, COL4A1, HSPG2), FSH and LH receptors, and cell cycle genes (CCND2, CCNE1, CCNE2, CDKN1B, CDKN2D) were examined in apical and basal granulosa cells from large healthy bovine follicles [n = 18, 14.3 ± 0.3 mm (mean + SEM)] using quantitative RT-PCR. Apical granulosa cells were collected by flushing the follicle with balanced salt solution. The remaining cells were detached from the follicular basal lamina by gently scraping; these are the basal granulosa cells. This collection method resulted in equivalent cell yields of apical and basal cells. Expression for all genes was significantly higher in basal cells in comparison to apical cells (P < 0.05), except for the cycle genes CCND2 and CDKN2D, which did not differ between cell populations. These results suggest that functional heterogeneity exists within the membrana granulosa. How differences between apical and basal cells are established is unknown but may be due to the proximity of the basal cells to the follicular basal lamina. The relevance of this aspect of follicle maturation to the endocrine function of granulosa cells has yet to be determined.

Ultrastructure of the basal lamina of bovine ovarian follicles and its relationship to the membrana granulosa

Reproduction ◽  
2000 ◽  
pp. 221-228 ◽  
Author(s):  
HF Irving-Rodgers ◽  
RJ Rodgers
Keyword(s):  
Basal Lamina ◽  
Granulosa Cells ◽  
Light And Electron Microscopy ◽  
Single Layer ◽  
Antral Follicle ◽  
Basal Cells ◽  
Preantral Follicles ◽  
Antral Follicles ◽  
Cellular Processes ◽  
Innermost Layer

Different morphological phenotypes of follicular basal lamina and of membrana granulosa have been observed. Ten preantral follicles (< 0. 1 mm), and 17 healthy and six atretic antral follicles (0.5-12 mm in diameter) were processed for light and electron microscopy to investigate the relationship the between follicular basal lamina and membrana granulosa. Within each antral follicle, the shape of the basal cells of the membrana granulosa was uniform, and either rounded or columnar. There were equal proportions of follicles </= 4 mm in diameter with columnar basal cells and with rounded basal cells. Larger follicles had only rounded basal cells. Conventional basal laminae of a single layer adjacent to the basal granulosa cells were observed in healthy follicles at the preantral and antral stages. However, at the preantral stage, the conventional types of basal lamina were enlarged or even partially laminated. A second type of basal lamina, described as 'loopy', occurred in about half the preantral follicles and in half the antral follicles </= 4 mm diameter. 'Loopy' basal laminae were not observed in larger follicles. 'Loopy' basal laminae were composed of basal laminae aligning the basal surface of basal granulosa cells, but with additional layers or loops often branching from the innermost layer. Each loop was usually < 1 microm long and had vesicles (20-30 nm) attached to the inner aspect. Basal cellular processes were also common, and vesicles could be seen budding off from these processes. In antral follicles, conventional basal laminae occurred in follicles with rounded basal granulosa cells. Other follicles with columnar cells, and atretic follicles, had the 'loopy' basal lamina phenotype. Thus, follicles have different basal laminae that relate to the morphology of the membrana granulosa.

scholarly journals The Differential Metabolomes in Cumulus and Mural Granulosa Cells from Human Preovulatory Follicles

2021 ◽  
Author(s):  
Er-Meng Gao ◽  
Bongkoch Turathum ◽  
Ling Wang ◽  
Di Zhang ◽  
Yu-Bing Liu ◽  
...  
Keyword(s):  
Oocyte Maturation ◽  
Granulosa Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Cumulus Cells ◽  
Cholesterol Transport ◽  
Vitro Fertilization ◽  
Expression Of Genes ◽  
Preovulatory Follicles ◽  
Morphological Differences

AbstractThis study evaluated the differences in metabolites between cumulus cells (CCs) and mural granulosa cells (MGCs) from human preovulatory follicles to understand the mechanism of oocyte maturation involving CCs and MGCs. CCs and MGCs were collected from women who were undergoing in vitro fertilization (IVF)/intracytoplasmic sperm injection (ICSI) treatment. The differences in morphology were determined by immunofluorescence. The metabolomics of CCs and MGCs was measured by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) followed by quantitative polymerase chain reaction (qPCR) and western blot analysis to further confirm the genes and proteins involved in oocyte maturation. CCs and MGCs were cultured for 48 h in vitro, and the medium was collected for detection of hormone levels. There were minor morphological differences between CCs and MGCs. LC-MS/MS analysis showed that there were differences in 101 metabolites between CCs and MGCs: 7 metabolites were upregulated in CCs, and 94 metabolites were upregulated in MGCs. The metabolites related to cholesterol transport and estradiol production were enriched in CCs, while metabolites related to antiapoptosis were enriched in MGCs. The expression of genes and proteins involved in cholesterol transport (ABCA1, LDLR, and SCARB1) and estradiol production (SULT2B1 and CYP19A1) was significantly higher in CCs, and the expression of genes and proteins involved in antiapoptosis (CRLS1, LPCAT3, and PLA2G4A) was significantly higher in MGCs. The level of estrogen in CCs was significantly higher than that in MGCs, while the progesterone level showed no significant differences. There are differences between the metabolomes of CCs and MGCs. These differences may be involved in the regulation of oocyte maturation.

scholarly journals Morphological classification of bovine ovarian follicles

Reproduction ◽  
2010 ◽  
Vol 139 (2) ◽  
pp. 309-318 ◽  
Author(s):  
R J Rodgers ◽  
H F Irving-Rodgers
Keyword(s):  
Basal Lamina ◽  
Granulosa Cells ◽  
Follicular Development ◽  
Basal Cells ◽  
Morphological Classification ◽  
Primordial Follicles ◽  
Antral Follicles ◽  
Different Types ◽  
Biological Differences

Follicle classification is an important aid to the understanding of follicular development and atresia. Some bovine primordial follicles have the classical primordial shape, but ellipsoidal shaped follicles with some cuboidal granulosa cells at the poles are far more common. Preantral follicles have one of two basal lamina phenotypes, either a single aligned layer or one with additional layers. In antral follicles <5 mm diameter, half of the healthy follicles have columnar shaped basal granulosa cells and additional layers of basal lamina, which appear as loops in cross section (‘loopy’). The remainder have aligned single-layered follicular basal laminas with rounded basal cells, and contain better quality oocytes than the loopy/columnar follicles. In sizes >5 mm, only aligned/rounded phenotypes are present. Dominant and subordinate follicles can be identified by ultrasound and/or histological examination of pairs of ovaries. Atretic follicles <5 mm are either basal atretic or antral atretic, named on the basis of the location in the membrana granulosa where cells die first. Basal atretic follicles have considerable biological differences to antral atretic follicles. In follicles >5 mm, only antral atresia is observed. The concentrations of follicular fluid steroid hormones can be used to classify atresia and distinguish some of the different types of atresia; however, this method is unlikely to identify follicles early in atresia, and hence misclassify them as healthy. Other biochemical and histological methods can be used, but since cell death is a part of normal homoeostatis, deciding when a follicle has entered atresia remains somewhat subjective.

scholarly journals Composition and morphology of the follicular basal lamina during atresia of bovine antral follicles

Reproduction ◽  
2002 ◽  
pp. 97-106 ◽  
Author(s):  
HF Irving-Rodgers ◽  
ML Mussard ◽  
JE Kinder ◽  
RJ Rodgers
Keyword(s):  
Basal Lamina ◽  
Granulosa Cells ◽  
Cell Shape ◽  
Type Iv Collagen ◽  
Basal Cells ◽  
Follicle Growth ◽  
Electron Microscopic ◽  
Antral Follicles ◽  
Type Iv ◽  
Follicle Diameter

The fate of the follicular basal lamina during atresia was investigated using bovine follicles, in which different follicle phenotypes have been observed. These phenotypes include: healthy follicles with rounded basal granulosa cells with an aligned basal lamina or follicles with columnar basal granulosa cells with a basal lamina of many loops (loopy), and atretic follicles in which either the antral granulosa cells (antral atresia) or the basal cells (basal atresia) die first. Loopy lamina and basal atresia occur only in small antral follicles < 5 mm in diameter. Follicles were collected from cattle of unknown reproductive history and processed for immunohistochemistry and electron microscopy, and from animals in which follicle growth had been monitored by daily measurements of follicle diameter by ultrasonography. Electron microscopic observations of dominant follicles during the growth phase, plateau and regression showed that the basal lamina was still visible and intact upon atresia. These follicles had a conventional aligned basal lamina, which they retained, except for some degree of folding, as they progressed into antral atresia. In small follicles (2-5 mm in diameter), the basal cell shape (rounded or columnar) and appearance of the basal lamina (aligned or of many loops) did not appear to be related to the type of atresia. On atresia the follicular basal laminae retained immunoreactive laminin alpha1 and beta2, type IV collagen alpha1 and nidogen. Laminin alpha2, which may come from the theca, was present in the follicular basal lamina of only 22% of healthy follicles, but was expressed very commonly in 71% of the atretic follicles. Laminin alpha2 expression was found in both phenotypes of healthy follicles, antral and basal atretic follicles, and follicles with aligned or loopy basal laminae. It is concluded that the basal lamina is not degraded upon atresia, but does undergo a variety of other changes.

Insulin-like growth factor binding proteins in follicular fluid from morphologically distinct healthy and atretic bovine antral follicles

2003 ◽  
Vol 15 (4) ◽  
pp. 241 ◽  
Author(s):  
H. F. Irving-Rodgers ◽  
K. D. Catanzariti ◽  
M. Master ◽  
P. A. Grant ◽  
P. C. Owens ◽  
...  
Keyword(s):  
Growth Factor ◽  
Basal Lamina ◽  
Granulosa Cells ◽  
Follicular Fluid ◽  
Binding Proteins ◽  
Basal Layer ◽  
Insulin Like Growth Factor ◽  
Basal Cells ◽  
Factor Binding ◽  
Different Types

In bovine follicles 2–5 mm in diameter, two morphologically distinct types of healthy follicles and two types of atretic follicles have been described recently. Healthy follicles either have columnar basal granulosa cells with follicular basal lamina composed of many layers or ‘loops’ or they have rounded basal cells with a conventional single-layered, aligned follicular basal lamina. In atretic follicles, cell death either commences at the basal layer and progresses to the antrum (basal atresia) with macrophage penetration of the membrana granulosa or death progresses from the antrum in a basal direction (antral atresia). Little is known about how these different phenotypes develop. To determine whether insulin-like growth factor binding protein (IGFBP) levels in follicular fluid differ between these different types of follicles, we measured IGFBP levels in fluids from these follicles. A total of 61 follicles were assessed by light microscopy and characterized by morphological analysis as either healthy, with columnar or rounded basal granulosa cells, or as undergoing antral or basal atresia. The IGFBP concentration in the follicular fluid of individual follicles from the four groups (n = 12–20 per group) was identified by Western ligand blots using 125I-insulin-like growth factor (IGF)-II as a probe. Insulin-like growth factor binding proteins 2, 3 (44 and 40 kDa), 4 (glycosylated and non-glycosylated) and 5 were observed. The levels (per volume of fluid) of IGFBPs 2, 4 and 5 were greater in atretic follicles than in healthy follicles. However, there were no statistical differences in levels of each IGFBP between either the two types of healthy follicle or between the two types of atretic follicles. Thus, IGFBP levels are not related to the different types of healthy or atretic follicles.

scholarly journals Atresia revisited: two basic patterns of atresia of bovine antral follicles

Reproduction ◽  
2001 ◽  
pp. 761-775 ◽  
Author(s):  
HF Irving-Rodgers ◽  
IL van Wezel ◽  
ML Mussard ◽  
JE Kinder ◽  
RJ Rodgers
Keyword(s):  
Basal Lamina ◽  
Granulosa Cells ◽  
Immunohistochemical Study ◽  
Steroid Hormone ◽  
Basal Layer ◽  
Follicle Development ◽  
Basal Cells ◽  
Apoptotic Bodies ◽  
Antral Follicles ◽  
Follicular Fluids

Our observations of bovine follicles indicated that the original histological classifications of atresia were inaccurate. A detailed histological, ultrastructural and immunohistochemical study of antral follicles from bovine ovaries collected from an abattoir and from animals whose large follicles had been monitored by ultrasonography was conducted to investigate this further. Nidogen and CD68 were immunolocalized to observe the follicular basal lamina and macrophages, respectively. In randomly collected ovaries, approximately one quarter of all antral follicles were undergoing antral atresia, as designated in this study. Antral atresia was characterized by early destruction of the layers of the membrana granulosa closest to the antrum, whereas the most basal cells remained intact. Numerous pyknotic nuclei were observed in the most antral layers and in the antrum close to the membrana granulosa. This is the classic description of atretic follicles and was observed at all sizes of follicle development and almost universally in large follicles (> 5 mm in diameter), including dominant follicles. Basal atretic follicles, as designated in this study, were almost as prevalent as the antral atretic follicles, and were characterized by initial destruction of the most basal layer of granulosa cells, whereas the cells in the most antral layers remained associated with each other and were predominantly healthy. Pyknotic nuclei and the nuclei of dying basal cells budded into apoptotic bodies were observed rarely. The basal lamina of basal atretic follicles was often breached by macrophages, which were phagocytosing dying basal granulosa cells. The theca was characterized by an increased deposition of collagen, and the cells were orientated randomly, rather than lying parallel to the membrana granulosa as in healthy follicles. Basal atresia occurred in small (< 5 mm in diameter) follicles only. Importantly, these basal atretic follicles were originally identified incorrectly in the literature. Thus, on the basis of the results of this study and another on the expression of steroidogenic enzymes in atretic follicles, it is suggested that the standard biochemical methods for measuring steroid hormone concentrations in follicular fluids to assess atresia should be re-evaluated.

Development of the membrana granulosa of bovine antral follicles: structure, location of mitosis and pyknosis, and immunolocalization of involucrin and vimentin

1999 ◽  
Vol 11 (1) ◽  
pp. 37 ◽  
Author(s):  
I. L. van Wezel ◽  
R. J. Rodgers ◽  
M. Krupa
Keyword(s):  
Granulosa Cells ◽  
Ovarian Follicle ◽  
Follicular Development ◽  
Staining Intensity ◽  
Keratinocyte Differentiation ◽  
Follicular Epithelium ◽  
Intermediate Filament Protein ◽  
Basal Cells ◽  
Filament Protein ◽  
Dying Cells

The membrana granulosa of the ovarian follicle is termed the ‘follicular epithelium’, yet there have been no studies considering its epithelial nature and how it changes during follicular development. Therefore, these issues were investigated using histology (n = 45 ovaries), considering its structure and the location of proliferating and dying cells, and drawing analogies with other epithelia. Additionally, differences between the layers of granulosa cells were demonstrated by immunohistochemistry (n =7 ovaries). The structure of the membrana granulosa differed between follicles. Six arbitrary classifications were designed based on these structures, 80 follicles were allocated (n = 13 ovaries) to these classes and the follicular diameters were then measured. For the first time, differences in membrana granulosa structure were shown to correspond to follicle size. Follicles in classes 1–3, where basal granulosa cells were columnar with nuclei positioned basally in the cell, were all ≤3 mm in diameter. All follicles larger than 3 mm had either columnar basal cells with nuclei positioned centrally (class 4), or had rounded basal cells (class 5), and all follicles >5 mm had only rounded basal cells. In all these classes, cells in the middle zone were rounded; cells aligning the antrum were often flattened. Irrespective of follicle class, cell proliferation and cell death were shown to be predominantly in the middle portions, rather than the most antral or most basal portions, of the membrana granulosa of healthy and atretic follicles. Involucrin, a marker of keratinocyte differentiation, was localized to the suprabasal region of the membrana granulosa of healthy follicles, particularly in the second and third cellular layers in from the follicular basal lamina. Conversely, the staining intensity for the intermediate filament protein vimentin was lowest in this region, and greatest in the more antral and basal regions. In atretic follicles, there was widespread staining for involucrin and vimentin throughout the membrana granulosa. In conclusion, the membrana granulosa is highly structured, and alters with follicular development. Layers in the membrana granulosa can differ in terms of cell shape, and differ in proliferation and gene expression. In the light of the current work, and an associated study, it is proposed that proliferation occurs in the middle layers, and that granulosa cells then progress basally or antrally, the latter undergoing terminal differentiation.

Classification of small type B/C follicles as primordial follicles in mature rats

Reproduction ◽  
2000 ◽  
pp. 43-48 ◽  
Author(s):  
S Meredith ◽  
G Dudenhoeffer ◽  
K Jackson
Keyword(s):  
Dna Synthesis ◽  
Granulosa Cells ◽  
Single Layer ◽  
Reliable Indicator ◽  
Type B ◽  
Primordial Follicles ◽  
Type C

In the present study, follicles were classified according to the morphology of their granulosa cells. Type B follicles contained only flattened granulosa cells; type B/C follicles had a mixture of flattened and cuboidal granulosa cells in a single layer, and type C follicles had a single layer of cuboidal granulosa cells. The primary objectives of the study were to determine whether 5-bromo-2-deoxyuridine incorporation into type B/C follicles was a marker for initiation of growth and how long type B/C follicles could remain at the same stage before transformation to type C follicles. Female Holtzman rats received bromo-deoxyuridine for 7 days. After the infusion (day minipumps were removed = day 0), rats were ovariectomized on days 0 (n = 9), 30 (n = 8), 90 (n = 8) and 150 (n = 9). The numbers of type B, B/C and C follicles within one ovary were determined using modified fractionator counting. Analysis over all times demonstrated that there were more (P < 0.0001) type B/C (941 +/- 61 per ovary) than type C (140 +/- 18 per ovary) or type B (159 +/- 19 per ovary) follicles. The numbers of type B and type C follicles did not differ from each other at any time. Only one of 34 rats evaluated had bromo-deoxyuridine-labelled type B follicles. On day 150, 57% of the bromo-deoxyuridine-labelled type B/C follicles remained from day 0. It is concluded that (1) DNA synthesis in granulosa cells of type B/C follicles is not a reliable indicator of impending growth; and (2) type B and type B/C follicles are both components of the pool of primordial follicles.

Cytological and Histological Study of Adult and Neonate Epidermis in Thick and Thin Skin of Various Anatomical Sites

2018 ◽  
Vol 9 (2) ◽  
Author(s):  
Hydar Muhsin Khalfa ◽  
Adnan Albideri ◽  
Haider Salih Jaffat
Keyword(s):  
Histological Study ◽  
Single Layer ◽  
Sweat Glands ◽  
Anatomical Site ◽  
Basal Cells ◽  
Histological Changes ◽  
Mature Adult ◽  
Skin Development ◽  
Stage Of Development ◽  
Thin Skin

The integumentary system covers the surface of the embryo (skin) and its specialized skin structures including hair, nails, sweat glands, mammary glands and teeth. During fetal skin development, the epidermis changes from a single layer of ectodermal cells at 7–8 days of gestation into a more apparent stratified, keratinized epithelium at 22–24 weeks. The aim of the study is to identify the histological and cytological changes that take place during neonatal and adult epidermis development. Human neonatal and adult samples were obtained from fully informed, consenting parent or releatives from Al-hilla mortary / Iraq. Neonatal samples were obtained from neonates after sudden deaths from maternity wards. Anatomical Sites included abdomen, forehead, back, shoulder and feet sole. A totoal of 15 neonates and 10 mature adults were used for this study. Fresh tissues were sectioned using a freezing cryostat. Tissues were sectioned at 5µm in -24°C and collected on microscopic slides. Slides were allowed to air dry for 30 min prior to hematoxyline and eosin staining. Tissues were also photographed using scanning electron microscopy SEM. Cytological measurements were taken using image j software and data was analysed using graph prism. Various cytological and histological changes takes place during neonatal and adult and epidermis development. Our study shows the stages of fair follicule formation as well as number of nucleated layers present at each stage of development and at different anatomical sites. Major histological changes takes places during the transition frm a neonate to a mature adult including the number of basal cells and epidermal thickness depending on the anatomical site.

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