Implantation, Pregnancy and Parturition
(T581) LOW BLOOD ISG15 mRNA AND PROGESTERONE LEVELS ARE PREDICTIVE OF NON-PREGNANT DAIRY COWS.
Austin, Kathleen1, Han, Hyungchul1, Rempel, Lea 1, Hansen, Thomas1, 1 University of Wyoming, Laramie, WY
ABSTRACT- Rectal palpation (day 45) and ultrasound (day 27+) are used to determine pregnancy status in dairy cows and are performed relatively late following establishment of pregnancy (days 14-19). Because most estrous cycles average 21 days, it is also possible to sample cows at a time when progesterone concentration is low, making non-pregnant cows distinguishable from pregnant cows with elevated progesterone concentration. ISG15 is induced by conceptus derived interferon and is expressed in very high levels in the endometrium on days 15-45 of pregnancy. It was hypothesized that low blood ISG15 mRNA levels might provide an indication of non-pregnant cows on day 18. Whole blood was collected from Holstein dairy cows (n = 21) on days 15, 16, 17, 18, 19, 20, 21, 25 and 32 following artificial insemination (AI). Whole blood was mixed with TRI Reagent BD (Molecular Research, Inc.) and snap-frozen until total RNA was extracted. Plasma was collected for determination of progesterone using radioimmunoassay. Total RNA was treated with DNAse I, converted to single stranded cDNA and subjected to Real Time PCR amplification using SYBR Green (BioRad, Hercules, CA). GAPDH was used as an internal standard. Pregnancy was determined on day 35 using ultrasound (7/21 pregnant). ISG15 mRNA expression in the blood increased after day 16, peaked at day 20 and then declined to day 16 levels by 32 days following AI. Mean ISG15 mRNA expression in the blood was 4.6-fold greater (P<0.05) in pregnant when compared to non-pregnant cows. Low ISG15 mRNA and low progesterone were both 100% accurate in predicting non-pregnant cows when compared with ultrasound results. However, high ISG15 mRNA resulted in detecting fewer (P<0.05) false positive pregnant cows (14.3%; 2 cows with high ISG15 mRNA/ 14 cows that were not pregnant) when compared to plasma progesterone (35.7%; 5/14 cows). These false positive rates may be due to a viable embryo on days 17-21, that dies by day 35 when ultrasound was performed. It is concluded that detection of low ISG15 mRNA levels in blood on days 17-25 provides an accurate indicator of cows that are not pregnant. This might be used as a management tool to help identify nonpregnant cows 18 days following AI so that they can immediately be re-synchronized and inseminated. Supported by a grant from Aspen Bio Inc, CO.
KEY WORDS: Pregnancy, Blood, Bovine, Embryo