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Pregnancy course depends on the appropriate connection between the mother and the developing foetus. Pregnancy is completed when the placenta is timely expelled. Placental retention is one of the possible pregnancy complications. Extracellular matrix, including adhesive proteins and enzymes that can break down collagens, seems to be responsible for it. The aim of the present study was to examine the impact of one of the adhesive proteins - glycodelin (Gd) - on selected metalloproteinases degrading collagens (MMP2, MMP3, MMP7). Placental tissues from healthy pregnant cows collected during early-mid pregnancy (2nd month n = 7, 3rd month n = 8, 4th month n = 6) and in cows that properly released placenta (NR; n = 6) and cows with retained foetal membranes (R; n = 6) were experimental material. The concentrations of glycodelin and protein content of selected metalloproteinases were measured by ELISA in the maternal and foetal placental homogenates as well as in the culture of epithelial cells derived from the maternal part of the placenta. The presence of these protein molecules was confirmed by Western Blotting. In the bovine placenta, the concentrations of examined proteins exhibit significant changes during placental formation. Gd, MMP3 and MMP7 concentrations decrease with pregnancy progress (between the 2nd and 4th month), while MMP2 concentrations were on the same level in this period. During parturition, concentrations of Gd and MMP3 were significantly higher in the R group compared to the NR group. In parallel, MMP2 concentrations did not show significant differences between the groups (NR vs R), and MMP7 concentrations decreased significantly in the maternal part of the placenta in cows with retained foetal membranes (R). Obtained results show correlations between the gestational age and proteins' (Gd, MMP3, MMP7) concentration, both in the maternal and foetal part of the placenta., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Placental retention in cows may be the result of altered protein pattern in comparison to physiologically released fetal membranes. Aim of study was to separate and identify proteins from maternal and fetal part of placenta and to compare them between released and retained fetal membranes. Six not retained and 6 retained tissues were obtained from healthy cows during routinely performed caesarian section. Cows were allocated to appropriate groups retrospectively. Samples were homogenized in phosphate buffer and subjected to 2D electrophoresis. After analysis of gels selected spots were excised and proteins were identified by MS. Two-dimensional electrophoresis detected and identified 886 spots in examined tissues. Significant differences (p < .05) were noticed between appropriate parts of retained and released placenta. In maternal part of retained placenta 40 spots showed lower abundance and 47 higher abundance in comparison to healthy samples. While in fetal part of retained placenta respective values were 60 and 125 proteins. Out of 73 identified proteins, 26 were significantly different between respective maternal (19) and fetal (7) part of retained and released placenta. In summary, protein profile of released and retained placenta express the presence and abundance of different proteins. It may suggest that selected proteins could be target molecules in searching for reasons for placental retention. Further identification of spots obtained here may provide with more detailed explanation of mechanisms of placental retention., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Introduction: Regardless intensive research, the etiology and mechanisms of retention of fetal membranes in cows, still require elucidation. In our research approach, difference in gel electrophoresis (DIGE) and matrix-assisted laser desorption/ionization (MALDI) identification were used to obtain first results on protein profile of bovine placental membranes which were properly released or retained for more than 12 h after parturition., Methods: Placentomes from 6 cows that released placenta and from 6 cows that retained fetal membranes were homogenized, fluorescence labeled and subjected to DIGE., Results: Selected spots that significantly differed between retained and released placenta as well as spots with constant appearance were identified by MALDI. This allowed identification of the following proteins with high statistical reliability: Transforming growth factor beta 2 - high expression in maternal and fetal part of retained fetal membranes, Short transient receptor potential channel 5 -high expression in maternal part of retained and not retained fetal membranes, Rab GDP dissociation inhibitor beta - high expression in fetal part of retained and not retained fetal membranes, Proline dehydrogenase 2 - similar expression in all examined samples, Ras-related protein Rab-7b -high expression only in maternal part of not retained fetal membranes., Discussion: Up to now, these proteins have not been considered as possibly important molecules for the separation/retention of fetal membranes, but their biological roles may suggest it. Further studies are necessary to establish a full profile of bovine placental proteins and define target molecules that may be involved in separation/retention of fetal membranes., (Copyright © 2014 Elsevier Ltd. All rights reserved.)

Tissue-specific protein profile is determined by its function, structure, intensity of metabolism and usefulness. This profile remains under hormonal control. Any disturbance in the general metabolism may be reflected in changes in both protein quantity and quality. These changes can be of low or high specificity, and some can be used as clinical markers of pathological conditions. The aim of this study was to describe and to compare the protein profile of caruncle and foetal villi of bovine placenta that was either properly released or retained. Placental tissues were collected from healthy cows, divided into releasing and retaining foetal membranes, homogenized and subjected to 1D and 2D electrophoresis. Computer-aided analysis of gel images showed essential qualitative and quantitative alterations in protein profile between tissues that were properly released and retained. Alterations concerned both the number of fractions and spots as well as the intensity of staining. This preliminary study provides a general overview of the differences in the protein profile between released and retained foetal membranes. It may allow for selecting the group of proteins or single molecules, which should be further analysed in detail as possible markers differentiating the retention of foetal membranes in cows from placentas that were released spontaneously. The continuation of the study for the identification of particular spots detected in 2D gels is necessary., (© 2013 Blackwell Verlag GmbH.)

The maintenance of antioxidative/oxidative balance is crucial for cellular and extracellular environment. That is why antioxidative enzymes express their activity in different isoforms in different cell compartments and extracellular space. The aim of study was to verify the results of previous experiment on activities of antioxidative enzymes by the determination of their enzymatic proteins in bovine placental tissues by Western blotting technique. Moreover, the presence of particular isoenzymes was detected and differentiated. Homogenates of maternal and foetal part of both properly released and retained bovine placenta were subjected to PAGE electrophoresis in non-reducing and reducing conditions and Western blotting with appropriate antibodies against superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px). Electrophoresis allowed for the detection of protein bands of molecular weight related to CuZn-SOD as well as cGSH-Px isoenzymes. The reaction with appropriate antibodies confirmed this. Densitometric analysis, although semi-quantitative, allowed for the observation of trends in differences in antioxidative enzyme proteins, which may partly confirm previously described results in cases of retained and released placenta. Local antioxidative enzymatic mechanisms in bovine placental tissues are represented by CuZn-SOD and cGSH-Px, which show the changes in their expression during improper placental release., (© 2013 Blackwell Verlag GmbH.)

Poly(ADP-ribose) glycohydrolase (PARG) is the enzyme which degrades poly(ADP-ribose) polymers synthesized by poly(ADP-ribose) polymerase (PARP). Both enzymes are activated in response to different stimuli like oxidative stress and are involved in DNA repair processes. The retention of bovine foetal membranes (RFM) is supposed to be connected with oxidative stress conditions. The aim of the study was to detect the presence of PARG protein in bovine placenta in order to find the relationship between the process of releasing, retaining placenta and DNA repair. Placentomes, collected alter spontaneous delivery or caesarian section were divided into maternal as well as foetal part of placenta, homogenized and subjected to electrophoresis. Animals were divided into six groups as follows: A--caesarian section before term with RFM; B--caesarian section before term without RFM; C--spontaneous delivery at term with RFM; D--spontaneous delivery at term without RFM; E--caesarian section at term with RFM; F--caesarian section at term without RFM. PARG protein was detected in nitrocellulose membranes using commercially available bovine anti-PARG antibody and Western blotting technique. Single bands referred to bovine PARG standard were observed in all examined tissues as well as in human placenta used as the control of procedure. In addition, the intensity of staining was stronger in retained than properly released term placenta and in foetal than in maternal part of the placenta. These results may suggest the differences in enzyme protein content and careful conclusions can be drawn that the activities of PARG may be altered between compared groups of animals. It may confirm the presence of oxidative stress conditions and their consequences on metabolic pathways, the content of biologically active substances and processes of proper releasing placenta. Further experiments on PARG activity in bovine foetal membranes with respect to proper and improper placental release are necessary.

Poly(ADP-ribose) polymerase (PARP) synthesizes poly(ADP-ribose) in response to DNA strand breaks using NAD+ as a substrate. It leads to consequences for metabolism not only on a cellular level, but also on a tissue level, among others: NAD+ and ATP depletion. Retention of foetal membranes (RF) in cows is supposed to be connected with the imbalance between production and neutralization of reactive oxygen species, leading to oxidative damage to DNA, lipids and proteins. The aim of this preliminary study was to detect the presence of PARP in bovine placenta and to describe the enzyme with respect to type of placental tissue, time and mode of delivery. Placentomes, collected after spontaneous delivery or caesarian section, were divided into maternal and foetal parts of placenta, homogenized, and subjected to electrophoresis. Cows were divided into six groups as follows: (A) caesarian section before term with RF, (B) caesarian section before term without RF, (C) spontaneous delivery at term with RF, (D) spontaneous delivery at term without RF, (E) caesarian section at term with RF, (F) caesarian section at term without RF. PARP was detected by Western blotting using commercially available bovine anti PARP antibody. Bands referred to as bovine PARP standard were present in all examined tissues as well as the products of its cleavage. However, the patterns of bands were different with respect to type of tissue, time, and mode of delivery. Further experiments on detailed relationship between PARP activity and the process of releasing and retaining of bovine placenta are necessary.

Poly(ADP-ribose) polymerase (PARP) is the enzyme which utilises NAD to synthesise poly(ADP-ribose) polymers. This process appears in response to DNA lesions. Oxidative stress, which might be involved in bovine placental retention, is the reason for oxidative DNA injury. In this mini-review, the relationship between PARP activity and bovine placental retention is discussed. The results of our experiments on PARP activity in placental tissues showed that the enzyme of 113 kDa and its cleavage products were present in retained as well as released fetal membranes. Western blotting technique showed different intensities in the staining of bands which might suggest different activities of the enzyme.

Vitamin C and glutathione (GSH) are water-soluble antioxidants which take part in defence mechanisms against reactive oxygen species (ROS). They may also be involved in processes of releasing/retaining bovine fetal membranes. Hence vitamin C, reduced (GSH) and oxidised (GSSG) glutathione levels were determined in retained and not-retained bovine fetal membranes in order to describe the non-enzymatic antioxidative status. Placental samples were collected immediately after spontaneous delivery or during caesarean section before term and at term, and 6 groups were formed as follows: (A) pre-term caesarean section without retained placenta; (B) pre-term caesarean section with retained placenta; (C) term caesarean section without retained placenta; (D) term caesarean section with retained placenta; (E) spontaneous delivery without retained placenta and (F) spontaneous delivery with retained placenta. Homogenates of maternal and fetal placental tissues were prepared, and vitamin C, GSH and GSSG were measured spectrophotometrically. Vitamin C levels were significantly higher in the maternal part than in the fetal part of the placenta in all groups examined. In retained placenta cases the levels were significantly lower than in control cows, except in pre-term groups. GSH concentrations were significantly higher in placentas without retention than with retention. GSSG levels showed the opposite relationship and were significantly higher in samples with retention of fetal membranes than in controls. Further experiments on antioxidative as well as oxidative status in bovine placenta are necessary.

Glutathione peroxidase (GSH-Px), glutathione transferase (GSH-Tr), catalase (CAT) and superoxide dismutase (SOD)-the members of enzymatic antioxidative defence mechanisms against reactive oxygen species-may play an important role in proper or improper release of bovine fetal membranes. The aim of the following study was the determination of GSH-Px, GSH-Tr, CAT and SOD activity in order to define antioxidative status of bovine placenta during retention of fetal membranes (RFM) in cows. Placental samples were collected immediately after spontaneous parturition or during caesarean section before term and at term and divided into six groups as follows: A: caesarean section before term without RFM; B: caesarean section before term with RFM; C: caesarean section at term without RFM; D: caesarean section at term with RFM; E: spontaneous delivery at term without RFM; F: spontaneous delivery at term with RFM. The enzyme activities in placental homogenates were measured spectrophotometrically. GSH-Px activity was statistically significantly higher in fetal than in maternal placenta in all examined groups, increased towards parturition and was higher in caesarean section groups than spontaneous delivery groups. Statistically significantly higher activities were noticed in retained than not-retained placentae. GSH-Tr activity was significantly lower in fetal than in maternal placenta. In preterm groups, the activity was statistically significantly higher in retained than not retained placenta. In term groups, the opposite relationship was observed, higher values in caesarean section groups than spontaneous delivery were noticed. CAT activity was statistically significantly higher in fetal than in maternal part of placenta in all groups examined. The highest values in C and D groups and the differences between retained and not-retained placenta were observed. SOD exhibited the highest values in preterm placenta and alterations between retained and not-retained fetal membranes. In conclusion, the activities of GSH-Px, GSH-Tr, CAT and SOD are altered in cases of retained fetal membranes which may suggest the activation of antioxidative mechanisms caused by the imbalance between production and neutralization of reactive oxygen species., (Copyright 2001 Harcourt Publishers Ltd.)

Proteins as well as lipids and other biologically active substances may undergo peroxidative damage caused by reactive oxygen species (ROS). The aim of this study was the determination of this damage in cases of retained (RFM) and not-retained bovine foetal membranes. The levels of thiol groups were measured spectrophotometrically. Tryptophan, formylokinurenine and bityrosine bridges were determined using spectrofluorimetric methods. All parameters were determined in placental maternal and foetal homogenates collected from cows divided into six groups as follows: A, pre-term caesarian section without RFM; B, pre-term caesarian section with RFM; C, term caesarian section without RFM; D, term caesarian section with RFM; E, spontaneous delivery without RFM; F, spontaneous delivery with RFM. All parameters except tryptophan showed statistically significant differences between the maternal and foetal parts of the placenta. The concentrations of the thiol groups were lower in RFM than in not-RFM cases, whereas formylokinurenine and bityrosine concentrations showed the opposite relationship. Tryptophan levels were lower in cases of RFM rather than not-RFM samples but statistically significant differences occurred in groups C and D and the foetal part of groups E and F. Further experiments are necessary to determine the oxidative status of other biologically active substances in cases of retained bovine placenta.

Hyaluronidase, a proteoglycan-degrading enzyme, may have an influence on collagenolysis in bovine placenta and take part in the separation processes of the placenta in cows. There was, however, no evidence concerning the activity of this enzyme in those tissues. The experiment was performed on cows divided into two groups as follows: A, placenta not retained (n = 16); and B, placenta retained (n = 9). The activity of the enzyme was measured using a spectrophotometric method and zymography. The results showed statistically significant higher activity of hyaluronidase in the maternal and fetal parts of the placenta in cows with a retention of fetal membranes. Further experiments require substrate specificity studies and the presence of regulatory factors of this enzyme in bovine placental tissues.

This study compared the activity of matrix metalloproteinases (MMP-2 and MMP-9) in retained and non-retained bovine placenta. The activities of MMPs and their zymogens were measured in fetal and maternal placental tissues from control cows (group B) and animals affected with retention of fetal membranes (group A) using a zymography technique on 10 per cent SDS polyacrylamide gels. The activity of proMMP-9 detected only in the maternal part of the placenta was lower in group A than in group B. ProMMP-2 activity was higher in group A than in group B in both tissues. The active forms of MMP-2 were observed in the maternal and fetal part of placenta in group B, but only the 68-kDa form was detected in the placental tissues of group A. The differences in enzyme activity between the groups and the lack of 64- and 60-kDa active forms of MMP-2 in the maternal and fetal parts of the retained placenta may have influenced the hydrolysis of collagen and the proper release of fetal membranes.

The aim of this study was the determination and the comparison of activities of AST, ALT, GGT, 5'-NU and CK in the placental tissues of cows with and without retained fetal membranes. Placental samples were obtained from 16 cows immediately after parturition and divided into 2 groups: A-not retained (n = 10, NRF), B-retained (n = 6, RF). The activities of the examined enzymes were determined in supernatants of homogenized tissues using spectrophotometric methods and ready kits. The activity of AST was statistically significantly lower in the maternal part of the placenta in group B than in group A. There were no differences in ALT activity. The activity of GGT was statistically significantly higher in the maternal part of the placenta in group B than in group A. The activity of 5'-NU was statistically significantly higher in the maternal part than in the fetal part of placenta in both groups examined. The activity of CK did not differ, except for statistically significant lower activity in the fetal part of the placenta in group B. The results can suggest that the metabolism of amino acids is altered to some extend in cases of the retained placenta. Changes in GGT my indicate on imbalance in free radicals generation and neutralisation. Energetic status may not be influenced by the retention of the fetal membranes. Further experiments concerning more frequent sample collecting during whole periparturient period are necessary.

Changes in the levels of the activity of two enzymes that neutralize free radicals-glutathione peroxidase [(EC.1.11.1.9), GSH-Px] and superoxide dismutase--[(EC.1.15.1.1), SOD]--in cows with fetal membranes retention (RFM) were studied. The activities were measured in maternal and fetal placental tissues after spontaneous parturition without (group A) and with (group B) retained placenta. GSH-Px activities were higher in group B than in group A (P < 0.05). The activity of this enzyme was lower in the maternal than in the fetal part of the placenta in both groups. The activity of SOD was significantly (P < 0.05) lower in maternal part, and higher in fetal part, of placenta in group B compared with group A. Experiments on defence mechanisms against free radicals in periparturient cows in connection with more frequent sampling, prostaglandins levels, oxidative stress, nutrition and retained placenta are required to further elucidate the role of these enzymes.