Acid peptidase activity released from in vitro produced porcine embryos: A candidate marker to predict developmental competence

Abstract

SUMMARYThe ability to efficiently create high quality embryos, competent to produce normal viable offspring in vitro, facilitates diverse technological advancements in animal agriculture and assisted reproduction. Current methods for evaluation of embryos are predominantly based on morphological characteristics which are prone to potential bias of the scorer. Metabolic and genetic markers have also been explored for quality assessment, but they are cost prohibitive or require longer periods of time for evaluation. We hypothesized that secreted enzymes could provide another means of embryo quality assessment. In this report, we provide evidence that medium conditioned by porcine embryos often has proteolytic activity that operates in acidic conditions (acid peptidase activity or APA). The APA could be inhibited by pepstatin A, suggesting that the activity is derived from one or more aspartic peptidases. We also provide evidence that single embryos, incubated for as few as 24 hr, released enough APA that it was possible to measure it accurately at day 5 of culture. We also observed that such activity on day 6 could be positively correlated with advanced developmental stage and embryo quality. In addition, those embryos that were graded identically by morphological evaluations often differed in the amount of APA-with some being significantly higher than the experimental threshold value. Therefore, the APA of embryos might serve as an additional marker for evaluation of embryos.

Activation method does not alter abnormal placental gene expression and development in cloned pigs

“…The function of PAG2 has not been elucidated. Normally, PAG2 has very high expression levels (174.4‐fold higher than the reference cDNA in this study) and also has measurable acid peptidase activity (Telugu and Green, 2008; Telugu et al, 2009), but in this study NT extraembryonic membranes showed a decrease in PAG2 transcript and protein expression and acid peptidase activity. The NT extraembryonic membranes in this study showed PAG2 protein accumulating at the microvillar junction by Day 35 and in one case PAG2 immunoreactivity was also present in the LE.…”

“…The optimum pH for proteolytic activity of porcine PAG2 is 3.5 (Telugu and Green, 2008), therefore acid peptidase activity of extraembryonic membrane extracts was measured at this pH as described previously (Telugu et al, 2009) with minor modifications. Briefly, APA was measured in 40 µg protein lysates (the same lysates that were used for Western blotting analysis), each of the three biological replicates was measured in quadruplicate, resulting in 12 measurements/treatment.…”

Cattle adapted to tropical and subtropical environments: genetic and reproductive considerations

“…Subspecies genotype also affects plasma pregnancyassociated glycoprotein (PAG) concentrations during pregnancy. These proteins are part of a large family of aspartic proteases detected in the maternal circulation throughout most of pregnancy (Xie et al, 1997;Telugu et al, 2009;Pohler et al, 2016).…”

Somatic cell nuclear transfer efficiency: How can it be improved through nuclear remodeling and reprogramming?

“…The function of PAG2 has not been elucidated. Normally PAG2 has very high expression levels in the placenta and also has measurable acid peptidase activity (Telugu and Green, 2008; Telugu et al, 2009). PAG2 message and protein were down‐regulated in SCNT placentas compared to in vivo placentas by more than four‐ and two‐fold, respectively (Fig.…”