Effect of prostaglandin producing modulators on in vitro growth of buffalo uterine epithelial cellsTheriogenology

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Authors
S. Nandi, S. Mondal, I.J. Reddy
Year
2012
DOI
10.1016/j.theriogenology.2011.09.017
Subject
Equine / Food Animals / Small Animals / Animal Science and Zoology

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Lip linoleic 0, 0.01, 0.1, 1, 10 and 100 g/ml, d) oxytocin: 0, 10, 100, 1,000, 10,000 and 100,000 nM, e) tumor necrosis factor- (TNF-): 0, 0.05, 0.5, 1, 2.5 and 5 nM, f) progesterone: 0.1, 10, 25, 50, 75 and 100 nM, and g) estradiol: 0, 2.5, 5, 10, 20 and 50 nM. The con of sig and 0.0 to ©

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Available online at www.sciencedirect.com 2) 1014 009 doitrol medium consisted of RPMI-1640 plus 10% bovine fetal serum. The growth of uterine epithelial were measured in terms viability, cell number increment and monolayer formation. Results suggested that the growth of uterine epithelial cells were nificantly (P  0.05) higher in media containing 10 g/ml, 10 g/ml, 1 nM and 10 g/ml linoleic acid, linolenic acid, TNF-

LPS, respectively compared to control and lower doses used. Progesterone, estradiol and oxytocin did not significantly (P  5) increase the growth of uterine epithelial cells. In conclusion, the growth of uterine epithelial cells increased when exposed modulators in the order of linoleic acid  linolenic acid  LPS  TNF-  progesterone  estrogen  oxytocin. 2012 Elsevier Inc. All rights reserved. words: Modulators; Prostaglandin; Uterine growth; Buffalo

Introduction

Early embryonic mortality is regarded as one of the jor causes of reproductive failure resulting in reced pregnancy rate, slower genetic improvement and stantial economic loss to buffalo production [1]. staglandins (PG) play a crucial role in various reductive events such as ovulation, maternal recognin of pregnancy, implantation, and parturition [2]. e production of endometrial PGs is mainly governed the rate limiting enzymes—PGE synthase (PGES) d PGF synthase (PGFS) which catalyze the convern of PGH2 to PGE2 and PGF2, respectively. PGF2 acts as the luteolytic agent which brings the animal back into the cycle. In contrast, PGE2 protects the corpus luteum from spontaneous regression and helps in establishment of pregnancy [3]. Various modulators like tumor necrosis factor (TNF-), lipopolysaccharide (LPS), fatty acids (linoleic acid, linolenic acid), hormones (oxytocin, estrogen and progesterone), and interferons alter the prostaglandin biosynthesis during estrous cycle and early pregnancy [4–7]. We had standardized the techniques of isolation, culture and characterization of uterine endometrial epithelial and stromal cells of in buffalo [8,9]. Most of the earlier studies elucidated the role of modulators on PG production from uterine endometrial cells. The effects of modulators on growth pattern of uterine epithelial cells are notTechni

Effect of prostaglandin producin of buffalo uterin

S. Nandia,*, S. M a National Institute of Animal Nutrition and Physi stract

Studies were conducted to examine the effect of seven prost thelial cells in buffalo. The uterine epithelial cells isolated f opolysaccaride (LPS): 0, 0.01, 0.1, 1, 10 and 100 g/ml, b)

Theriogenology 77 (201ex pa

Corresponding author. Tel.: 91-080-25704146; fax: 91-08011420.

E-mail address: snandi71@yahoo.co.in(S. Nandi). 3-691X/$ – see front matter © 2012 Elsevier Inc. All rights reserved. :10.1016/j.theriogenology.2011.09.017ote odulators on in vitro growth pithelial cells la, I.J. Reddya

NIANP), Adugodi, Bangalore-560030, India in producing modulators on the in vitro growth of uterine aughtered buffaloes were cultured in media containing a) acid: 0, 0.01, 0.1, 1, 10 and 100 g/ml, c) linolenic acid: –1020 www.theriojournal.comamined till date. The importance of studying growth ttern of uterine cells lies in the fact that the uterine cel ide gro be eff in 2. 2.1 his ab on the ser the 2– en ear tim an ml ute ret cel sin cel

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Di 3. all to 1015S. Nandi et al. / Theriogenology 77 (2012) 1014–1020ls can be manipulated in a defined environment, for ntification of factors involved in regulation of wth and to have a better understanding the cell havior in culture. The present study investigated the ects of seven prostaglandin producing modulators on vitro growth of buffalo uterine epithelial cells.

Materials and methods . Isolation of endometrial epithelial cells

Uteri of non-descriptive healthy buffaloes with no tory of uterine infection were obtained from a local attoir within 1 h after slaughter and were transported ice to the laboratory within 1 to 1.5 h. The stages of estrous cycle were determined by macroscopic obvation of the ovary as described previously [10]. In present study, uteri of the early estrous cycle (Days 5) were used. The epithelial cells from the buffalo dometrium were isolated by the method as described lier [8]. Briefly, the uterine lumen was washed three es with sterile Ca2and Mg2 free Hank’s Balced Salt Solution (HBSS) supplemented with 100 IU 1 gentamycin and 0.1% BSA. The ends of the rine horn ipsilateral to corpus luteum were ligated to ain the trypsin solution for solubilizing the epithelial ls. Twenty ml of sterile HBSS containing 0.3% trypwas then infused into the uterine lumen. Epithelial ls were isolated by incubation at 37 °C for 60 min. e cell suspension obtained from the digestion was ered through plastic strainer (70 M) to remove unsociated tissue fragments. The filtrate was washed ee times with HBBS supplemented with gentamycin d 0.1% BSA by centrifugation at 600g for 10 min. ter the washes, the cells were counted with a hemotometer. The cell viability was higher than 95% as essed by 0.5% trypan blue dye exclusion. . Culture of endometrial epithelial cells

After cell counting and viability determination, the ithelial cells were seeded at the rate of 1 105 viable ls in RPMI 1640 medium at 38.5 °C in presence of

CO2 for two and seven days. The viability of ithelial cells at the time of plating was 92.4%. The ls were cultured in 100-l droplet covered by minl oil in 35 mm petri dish. The medium was changed ery 2 to 3 d until the confluency (80–90% of the plet) was reached. . Treatment doses