Analysis of Atresia in Bovine Follicles Using Different Methods: Flow Cytometry, Enzyme-Linked Immunosorbent Assay, and Classic Histology1

Blondin, Patrick; Dufour, Maurice; Sirard, Marc‐André

doi:10.1095/biolreprod54.3.631

Cited by 64 publications

(50 citation statements)

References 22 publications

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“…19,20) Gaultier et al 21) reported that heparin and over sulfated chondroitin sulfate had the capability to inhibit MMP-1 protein and mRNA expression. Wang et al 22) reported that intracellular MMP-1 was down-regulated by polysulphated polysaccharide which is consistent with our results.…”

Section: Discussionmentioning

confidence: 99%

Fucoidan Inhibits UVB-Induced MMP-1 Expression in Human Skin Fibroblasts

Moon

Lee

Shim

et al. 2008

Biological & Pharmaceutical Bulletin

125

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Skin can age in two ways: (i) intrinsic or chronologic aging, which is the process of senescence that affects all body organs; and (ii) extrinsic aging which occurs as a consequence of exposure to environmental factors including sunlight and cigarette smoke. 1)The most important of these factors is sunlight, particularly exposure to ultraviolet (UV)B irradiation, which causes photoaging. The photoaging process increases skin fragility, laxity, blister formation, leathery appearance and formation of wrinkles. 2)UVB induces the production of matrix metalloproteinases (MMPs) by activating cellular signaling transduction pathways 3) ; MMPs are responsible for the degradation or synthesis inhibition of collagenous extracellular matrix in connective tissues.3) Collagen represents the main component of the extracellular matrix of dermal connective tissue, and its concentration decreases in chronoaging and photoaging.All matrix macromolecules in the dermis can be digested by MMPs acting alone or in combination. The MMPs form a family of structurally and functionally related zinc endopeptidases that exhibit various substrate specificities. 4) Once collagen is initially cleaved by MMP-1, MMP-3 and other MMPs, collagen breakdown is further promoted. The enzyme mainly responsible for collagen breakdown in skin is, MMP-1 (fibroblast collagenase), which cleaves type I, III, VII, VIII and X collagen.Human fibroblast collagenase was the first vertebrate collagenase both purified to homogeneity as a protein, and cloned as a cDNA. This enzyme has been designated as matrix metalloproteinase-1 (MMP-1) and has served as the prototype for all the interstitial collagenases. MMP-1 is also known as collagenase-1. Starting from the N terminus the following features of domain organization are observed. 5)The pre-domain specifies a signal rich in hydrophobic amino acids that destines the synthesized polypeptide to the endoplasmic reticulum where it is removed during the transportation of the molecule from the cell to the outside. The propeptide domain indicates a sequence that is responsible of keeping the pro-form inactive. It presents a cysteine residue 73 that is located in a conserved sequence PRCGVPD opposite to a zinc atom at the active-form site and coordinated to it through an -SH group. The enzymatic activity of the proform enzyme is turned on by the displacement of this cysteine residue ("cysteine switch") and occurs by proteolytic cleavage, or by chemical disruption as in the case of oxidation or treatment with mercurial compounds. 6)Recent studies have shown that hairless mice exposed to UVB developed wrinkled skin and significantly enhanced MMP-1 mRNA expression. However, they have shown that the inihibition of MMP-1 activities by a specific MMP inhibitor suppresses UVB-induced wrinkle formation.7) This evidence suggests that MMP-1 plays a major role in the process of photoaging.Brown seaweed has been a staple of both Korean and Japanese diets and has also been documented as being used in traditional Chinese medicine for over ...

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Section: Discussionmentioning

confidence: 99%

Fucoidan Inhibits UVB-Induced MMP-1 Expression in Human Skin Fibroblasts

Moon

Lee

Shim

et al. 2008

Biological & Pharmaceutical Bulletin

125

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“…Thus, it is recommended that, if it is needed, a histological approach be undertaken, either by examination of cell morphology, or in combination with markers of cell death, or direct measurement of apoptosis by such methods of DNA end labelling (Jolly et al 1994) or FACS sorting of nuclei (Blondin et al 1996, Hendriksen et al 2003.…”

Section: Atresia Of Antral Follicles O5 MM In Diametermentioning

confidence: 99%

Morphological classification of bovine ovarian follicles

Rodgers¹,

Irving-Rodgers²

2010

Reproduction

129

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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 O5 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 O5 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.

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“…The healthy follicles were those with intact membrana granulosa and few pyknotic nuclei (<5% pyknotic nuclei) in this layer. Alternatively, atretic follicles showed attenuated membrana granulosa, disruption, loosely attached granulosa cells and increased number of pyknotic nuclei (>5% pyknotic nuclei) (15).…”

Section: Histological Evaluation Of Follicular Health and Atresiamentioning

confidence: 99%

Histological evaluation and in situ localization of apoptosis in fresh and cryopreserved ovarian tissue

Hussein

Jeremias

Sharma

et al. 2002

Fertility and Sterility

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Objective: To study the feasibility of using combined morphology and terminal deoxynucleotidyl transferase-mediated dUTP-digoxigenin nick-end labeling (TUNEL) for apoptosis detection and the impact of cryopreservation on this process. Materials and methods:We conducted this investigation using a porcine animal model. Bilateral oophorectomy was performed in eight sows. The ovarian tissues were divided into two parts; one part was immediately fixed while the other was cryopreserved. The cryopreserved specimens were subsequently thawed and then fixed. All the specimens were sectioned, fixed and stained with H&E. The ovarian follicles were counted, histologically categorized as healthy and atretic and evaluated for the presence of apoptosis. Then, in situ examination of apoptosis was performed using TUNEL assay. Results: In the cryopreserved tissues: 1) the count of the primordial follicle was significantly reduced as compared to freshly-fixed samples (4.9±5.3 vs. 7.2±5.4, p=0.03 respectively) and 2) the mean values of apoptosis were insignificantly higher when compared to the freshly-fixed group (p=0.74). Moreover: 1) apoptosis was found in the atretic, but not in the healthy follicles (primordial, primary and secondary); 2) the nuclei of the granulosa cells, but not those of theca or stromal cells, were TUNEL positive; 3) some cells with histological features of necrosis and apoptosis were TUNEL negative, and 6) The distribution of apoptosis was not different between cryopreserved tissue and freshly fixed tissue. Conclusions: the presence of apoptosis in the atretic follicles may suggest its involvement in follicular atresia; and 2) combined histology and TUNEL assay may be a useful method for detection of apoptosis. Key words: Apoptosis/ovarian cryopreservation/Morphology/TUNEL assay.In 1885, the first observation of apoptosis in the ovary was made following morphological analysis of granulosa cells in the rabbit ovary (1). This form of cell death is involved in the ovarian homeostasis (2) and its regulation depends on a balance between (3,4). Morphologically, apoptosis is characterized by specific changes, such as cytoplasmic vacuolization, chromatin condensation and formation of apoptotic bodies (5, 6). Biochemically, apoptosis is characterized by DNA fragmentation into oligonucleosome-sized fragments (6). This DNA fragmentation has been examined in the pig ovarian tissues; using DNA fluorescence flowcytometry (7) and autoradiography (8-10). However, these methods have the following limitations 1) they require large amounts of DNA and 2) they do not allow identification of apoptotic changes in specific tissue compartments or at the single cell level. 163Although morphology provided the original criteria for apoptosis and therefore remains as a reliable indicator, detection of early apoptotic changes may be beyond the scope of histological examination. Alternatively, it is well known that apoptosis may occur without evidence of DNA fragmentation (11) and therefore its detection becomes beyond the scope...

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Analysis of Atresia in Bovine Follicles Using Different Methods: Flow Cytometry, Enzyme-Linked Immunosorbent Assay, and Classic Histology1

Cited by 64 publications

References 22 publications

Fucoidan Inhibits UVB-Induced MMP-1 Expression in Human Skin Fibroblasts

Fucoidan Inhibits UVB-Induced MMP-1 Expression in Human Skin Fibroblasts

Morphological classification of bovine ovarian follicles

Histological evaluation and in situ localization of apoptosis in fresh and cryopreserved ovarian tissue

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