GnRH Agonist (Buserelin) Induced In Vitro Apoptosis in Bovine Endometrium.
Bibliographic record
Abstract
Apoptosis is a vital biological process that maintains tissue homeostasis by selective elimination of cells. The GnRH regulates gonadal physiology by acting as local modulator of apoptosis in ovaries, testes and prostate. Recently, we elucidated the presence of GnRH-R in the follicular and luteal phase bovine endometrium. The local modulatory role of GnRH, GnRH-R system in uterine biology is still not clear. Apoptosis acquires more significance in renewable structures like endometrium, undergoing cycles of cellular proliferation and elimination during the reproductive process. Therefore, we for the first time studied GnRH agonist (buserelin), induced apoptosis at the molecular (Bax, Bcl-2, Caspase, Fas mRNA), and the cellular level in the follicular and luteal phase bovine endometrium. 100 mg of endometrial explants (1-2 mm2) obtained from slaughter house reproductive tracts were cultured at 37°C in air with 5% CO2 in Dulbecco's modified eagle's medium containing 0.1% BSA, 100 IU/mL penicillin G and 100 μg/mL streptomycin for 20 h, then replaced media and treated the tissues with buserelin (0, 200, 500, 1000 ng/mL respectively), GnRH antagonist- antide (500 ng/mL) and antide + buserelin (500+200 ng/mL) for 6 h and stored the tissues at -80°C for RNA extraction. Two μg of total RNA was reverse transcribed using gene-specific primers and the PCR products were electrophoresed and visualized under UV illumination. The relative mRNA levels were measured using computerized software (Scion imaging beta) and normalized to the housekeeping gene G3PDH. To further study apoptosis at the cellular level, endometrial epithelial cells from the follicular and luteal phase endometrium were isolated, tested for cell viability using trypan blue exclusion method and seeded at a density of 100,000 cells/chamber in a collagen coated eight chamber glass slide at 37°C, 5% CO2 in DMEM/Ham F-12 medium (1:1), containing 10% heat-inactivated calf serum and 20 mg/ml gentamycin for 48 h. The cells were characterized by immunofluoroscence and using anticyokeratin as primary and FITC-labeled secondary antibody. After 48 h medium was replaced containing 1% calf serum and gentamycin and treated the cells with buserelin (200 ng/ml), antide (500 ng/ml) and a combination of antide + buserelin (500 + 200 ng/ml) for 6 h and allowed the cells to further grow for 24 h. The monolayers were then stained using acridine orange ethidium bromide and visualized under fluorescent microscope. Live and apoptotic cells were counted in three randomly selected fields in each chamber. The data were statistically analyzed using ANOVA and Student t and Dunett test (Caspase mRNA). This study revealed that buserelin (200 ng/ml) upregulates (P≤0.05) Bax and Caspase in follicular phase endometrium without affecting Bcl-2 and Fas mRNA levels. The cells in the present culture system were confirmed to be endometrial epithelial cells by observing clear labeling against cytokeratin. Buserelin (200 ng/mL) induced apoptosis (P≤0.05) in follicular endometrial epithelial cells. Luteal phase endometrium did not show apoptotic response to similar buserelin treatments at molecular or cellular level. This study for the first time demonstrate that GnRH agonist, buserelin induces receptor mediated apoptosis in follicular phase bovine endometrium at the molecular and cellular level and further indicates the involvement of mitochondrial cell death pathway in this phenomenon. (poster)
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.002 | 0.001 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".