EFFECT OF KISSPEPTIN ON IN VITRO MATURATION OF BUFFALO OOCYTES
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Date
27-11-2017
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PVNR TVU
Abstract
This study was aimed at investigating the effect of kisspeptin (Kp) on In Vitro
maturation of buffalo oocytes. The oocytes were aspirated from the ovaries collected from
the nearby slaughter house. From a total of 1371 ovaries, 2400 culture grade oocytes were
collected with a mean of 1.78 oocytes per ovary.
Two different experiments were conducted to investigate the effect of Kp alone at
different concentrations (experiment 1) or in combination with other hormones viz...
Follicle Stimulating Hormone (FSH), Luteinizing Hormone (LH) and Estradiol (E2) and
IVM media (experiment 2) on In Vitro maturation of buffalo oocytes. In the first
experiment, oocytes were allowed to mature in media supplemented with three different
concentrations (5, 10 and 15 μg/ml) of Kp, with the sole purpose of arriving at an
appropriate concentration of Kp to be used in experiment-2. Tissue Culture Medium 199
(TCM 199B), supplemented with Gentamicin was used as control. Culture grade oocytes
xxi
were randomly allotted to different In Vitro maturation media and cultured at 39oC with
5% CO2 under humidified atmosphere for 24 hours. At the end, oocytes were evaluated for
Cumulus Cell Expansion (CCE), extrusion of 1st Polar Body (PB). The proportion of
oocytes showing CCE and extrusion of 1st PB was highest in the medium supplemented
with 10 μg/ml of Kp.
Subsequently, experiment-2 was designed to evaluate the effect of Kp (10 μg/ml)
on In Vitro maturation of buffalo oocytes, in combination with various hormones. In this
experiment, oocytes were allowed to mature in 12 different maturation media viz. T1 to T12
[Control (TCM-199); T1, Kp; T2, FSH; T3, FSH+Kp; T4, LH; T5, LH+Kp; T6, E2; T7,
E2+Kp; T8, FSH+LH+E2; T9, FSH+LH+E2+Kp; T10, IVM; T11, IVM+Kp; T12]. Kp, FSH,
LH were supplemented at the concentration of 10 μg/ml each while E2 was used at 1 μg/ml
concentration in experiment-2. IVM medium consists of TCM-199 was supplemented with
FSH (10 μg/ml), LH (10 μg/ml), E2 (1μg/ml), Fetal Calf Serum (10%) and Bovine Serum
Albumin (10 μg/ml).
The proportion of CCE and extrusion of PB has also significantly increased when
Kp was supplemented with FSH (86.05±0.44 and 63.99±0.48), LH (81.34±0.54 and
59.94±0.46) and E2 (63.98±0.48 and 53.33±0.80). The maturation rates increased
significantly further when FSH, LH and E2 combination (T9) were additionally
supplemented with Kp (T10). However, they were significantly lower than T11 (IVM) and as
well T12 (IVM+Kp).
To conclude, the present study demonstrated that In Vitro addition of Kp (10 μg/ml)
to FSH, LH and Estradiol supplemented media enhances buffalo oocyte maturation In
Vitro.
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