Mammary Membranes Research Articles

MICROBIOTA OF VAGINA AND MAMMARY GLANDS SKIN IN THE PREGNANT WOMEN WITH PREECLAMPSIA

Background. Timely prediction, diagnosis and prevention of complications during the gestational period leading to perinatal loss and maternal mortality are the main tasks of contemporary obstetrics. About 50,000 women die from preeclampsia and eclampsia every year and perinatal mortality ranges from 15 to 25%.
 Objective. The aim of the research was to study and analyze the microecology of vagina and mammary glands skin in the pregnant women with underlying preeclampsia.
 Methods. The pregnancy examination was conducted at the Ternopil Regional Clinical Perinatal Center “Mother and Child”. The research involved 25 pregnant women with preeclampsia (the main group) and 15 healthy women with a physiological course of pregnancy (the control group). Material from the pregnant women was taken out of the skin of mammary glands and mucous membrane of vagina, then it was plated out in the nutrient medium for the cultivation and the microorganisms were defined.
 Results. The quantitative composition of a normal microflora of vagina and mammary glands skin in the control group was within the normal range. The quantitative composition of a normal microflora of vagina and mammary glands skin in the pregnant women of the main group decreased, the representatives of opportunistic and pathogenic flora were found.
 Conclusions. In the pregnant women with preeclampsia, abnormal microbiocenosis of vagina and breast skin was revealed, the degree of abnormality correlated with the severity of the disease. Our results may provide useful clinical knowledge to a broader understanding of microbiota role in pregnancy complications.

ECOLOGICAL FEATURES OF MICROBIOCENOSIS OF THE SKIN OF MAMMARY GLANDS AND VAGINA IN PREGNANT WOMEN WITH THREAT OF PRETERM LABOR

Background. The threat of preterm birth is one of the most topical issues in the world medicine. According to the statistics, from 12-13 to 25-35 % of all pregnancies end prematurely. One of the causes of preterm labour is chronic inflammatory processes of female genital organs and disorder of microbiocenosis. Timely diagnosis and adequate treatment will reduce the risk of premature labour and avoid perinatal loss.Objective. We examined and analysed the microflora of the skin of mammary glands and mucous membrane of vagina in healthy pregnant women and patients with threat of preterm labour.Materials and methods. The examination of the pregnant was conducted at the TRMPC “Mother and Child” in several stages. First of all, we rinsed the skin of mammary glands and smeared from mucous membrane the posterior vault of vagina with sterile swabs pre-moistened in physiological solution. After that, the tampons were placed in sterile tubes and delivered to laboratory. Sowing was carried out on Petri dishes with sterile medium: ZHSA, bloods MPA, Endo, Saburo, thioglycolic medium.Conclusions. As a result of the research we found saprophytic Gram-positive and Gram-negative microorganisms (in women with a physiological course of pregnancy). In pregnant women with preterm labour, there was the increase in the number of St. haemolyticus from 13% to 87%, appearance of representatives of pathogenic flora – St. aureus (in 20%).

Visualizing movement in the tumor microenvironment landscape

The local microenvironment influences tumor progression in several important ways. A recent study by Nguyen-Ngoc and colleagues used explants of primary human and mouse mammary tumors to examine how the composition of the extracellular matrix modulates tumor cell invasion. Culture in the presence of a three-dimensional laminin-rich basement membrane, similar to the mammary basement membrane in vivo, resulted in minimal invasion. In contrast, identical tumor fragments in a collagen I matrix which resembles the interstitial breast stroma exhibited a pronounced protrusive migration and local dissemination. These data emphasize the importance of the laminin-rich basement membrane in constraining tumor cell invasion.

Proteomic Approach Analysis of Mammary Membrane Proteins Expression Profiles in Holstein Cows

To investigate host defense mechanisms for protecting the mammary gland from mastitis infection, the membrane fraction of mammary tissues from Holstein cows was purified by differential velocity centrifugation, and then the sodium dodecyl sulfate-polyacrylamid gel electrophoresis (SDS-PAGE) separated proteins were identified by ion trap mass spectrometer equipped with a Surveyor high performance liquid chromatography (HPLC) system. A total of 183 proteins were identified. Bioinformatics software was applied to analyse physicochemical characteristics of the identified proteins and to predict biochemical function. These data may provide valuable information to investigate the mechanisms of mammary gland milk secretion and infectious disease, and enable a clear identification of proteins and potential protein targets for therapies.

Bovine lactoferrin binds to insulin-like growth factor-binding protein-3

Insulin-like growth factor (IGF)-binding protein-3 (IGFBP-3) has been shown to have IGF independent actions that appear to be mediated by specific IGFBP-3 binding proteins located on cell membranes. We show here using Western ligand blotting, a number of mammary membrane proteins that bind 125 I -labeled rhIGFBP-3. Immunoprecipitation studies demonstrated that the >70 kDa protein was identified from bovine mammary microsomes as bovine lactoferrin (bLf). In addition to being a secretory protein, Lf is tightly associated with cellular membranes. Labeled rhIGFBP-3 was shown to bind to commercially purchased and processed apo- or holo-human or bLf, but not bovine transferrin (bTf). Binding of [ 125 I ]rhIGFBP-3 to other positively charged proteins was not detected nor was binding to rhIGFBP-5 or other mammary-secreted IGFBPs observed. Reciprocal specific binding of [ 125 I ]bLf to rhIGFBP-3 was shown, but [ 125 I ]bTf did not show binding to rhIGFBP-3. While [ 125 I ]rhIGF-II does not bind to bLf, unlabeled rhIGF-II was shown to compete with [ 125 I ]bLf for rhIGFBP-3 binding. More detailed analysis by dot blot showed that Lf competes (ED 50=3 μg/ml) or displaces (ED 50=1 mg/ml) bound [ 125 I ]rhIGF-II from dot blotted rhIGFBP-3. In vitro studies with a bovine primary mammary epithelial cell culture showed that all- trans-retinoic acid stimulates the appearance of bovine IGFBP-3 and bLf in the conditioned media and that [ 125 I ]rhIGFBP-3 could be utilized to detect conditioned media bLf. These findings reveal a novel role for bLf, binding to IGFBP-3 and perhaps disassociating IGFBP-3:IGF when in high concentration.

Cloning of cDNA Encoding PAS-4 Glycoprotein, an Integral Glycoprotein of Bovine Mammary Epithelial Cell Membrane

Bovine PAS-4 is an integral membrane glycoprotein expressed in mammary epithelial cells. Complementary DNA (cDNA) cloning of PAS-4 was performed by reverse-transcriptase polymerase chain reaction (RT-PCR) with oligonucleotide probes based on it's amino terminal and internal tryptic-peptides. The cloned PAS-4 cDNA was 1,852 nucleotides (nt) long and its open reading frame (ORF) was encoded 1,413 base long. The deduced amino acid sequence indicated that PAS-4 consisted of 471 amino acid residues with molecular weight of 52,796, bearing 8 potential N-glycosylation sites and 9 cysteine residues. Partial bovine CD36 cDNA from liver also was sequenced and the homology of both nucleotide sequence was 94%. Most of the identical amino acid residues were in the luminal/extracellular domains. Contrary to PAS-4, bovine liver CD36 displays 6 potential N-glycosylation sites, which were located, except for those at positions 101 and 171, at same positions as PAS-4 cDNA. Cysteine residues of PAS-4 and CD36 were same at position and in numbers. Northern blot analysis showed that PAS-4 was widely expressed, although its mRNA steady-state levels vary considerably among the analyzed cell types. PAS-4 possessed hydrophobic amino acid segments near the amino- and carboxyl-termini. Two short cytoplasmic tails of the amino- and carboxyl-terminal ends constituted of a 5-7 and 8-11 amino acid residues, respectively.

Differentiation-associated expression of beta-N-acetylgalactosaminylated N-linked oligosaccharides in mammary epithelial cells.

Not only mammalian pituitary glycoprotein hormones but also many glycoproteins from a variety of animal species have been shown to contain N-linked oligosacchardies with the GalNAcbeta1 --> 4GlcNAc structure. Two types of beta-1,4-GalNAcT were found; one transfers N-acetylgalactosamine to acceptor oligosaccharides, which is stimulated by the hormone peptide and the other simply transfers sugar without such activation. In the case of bovine mammary membrane glycoproteins, the expression of beta-N-acetylgalactosaminylated N-linked oligosaccharides was associated with the functional differentiation of the epithelial cells. In contrast, the expression level of such oligosaccharides was much reduced in glycoprotein samples from human breast tumors compared with those from the unaffected regions. These results strongly suggest that the beta-N-acetylgalactosaminylation is one which is regulated under cellular differentiation and dedifferentiation of the mammary gland. Whether or not beta-N-acetylgalactosaminylated N-linked oligosaccharides have unique functions in addition to clearance of the hormone from the circulation remains to be elucidated.

Effect of Feeding Level and Nitrogen Source of the Diet on GH, IGF-T, and Prolactin Receptors in Mammary and Liver Tissue from Pre-pubertal Heifers

Pre-pubertal (86-220 kg BW) Finnish Ayrshire heifers were used to study the effect of feeding level and nitrogen source on GH, IGF-I, and prolactin (PRL) receptors in mammary and liver parenchymal tissue. The heifers were fed a hay-barley diet supplemented with either urea or rapeseed meal. The treatments were: (1) LU, low feeding level+urea; (2) LR, low feeding level+rapeseed meal; (3) HU, high feeding+urea; (4) HR, high feeding level+rapeseed meal. The average daily gains of the heifers on LU, LR, HU and HR diets were 692, 655, 805 and 890 g. At slaughter, mammary and liver parenchymal tissue was collected for preparation of crude membranes, and ligand-binding assays were performed. Specific binding of bGH to mammary membranes could not be detected. Linear Scatchard plots were obtained showing the presence of a single class of high-affinity binding sites for IGF-I in mammary tissue. Specific binding of IGF-I to mammary membranes was unaffected by feeding level and nitrogen source. Furthermore, Scatchard analysis showed unchanged binding capacities and binding affinities of the IGF-I receptors in mammary tissue from low and high fed heifers. Specific binding of prolactin to mammary membranes was increased by high feeding level (P<0.02) and by rapeseed meal (P<0.01). No interaction between feeding level and nitrogen source was identified. Hepatic binding of bGH, IGF-I and prolactin was unaffected by feeding level and nitrogen source. However, specific binding of prolactin tended to be lower (P<0.07) in liver tissue from heifers fed rapeseed meal as diet nitrogen source. The present study shows that the previously reported negative effect of high feeding level on pubertal mammary development (Mäntysaari et al., 1995), cannot be explained by changes in binding to GH and IGF-I receptors. The importance of the increase in prolactin binding to mammary membranes from heifers at high feeding level and with rapeseed meal needs further investigation.

Mammary gland membrane transport systems.

The secretion of milk depends on the activity of a large number of membrane transport systems located on the apical and basolateral membranes of mammary secretory cells. It follows that a thorough knowledge of individual mammary tissue membrane transport systems is required if we are to fully understand the process of milk secretion. The distribution of the transporters between the apical and basolateral poles of the mammary epithelium must be asymmetrical given that the mammary gland is capable of vectorial transport. This is particularly evident in the case of glucose and amino acid transport systems: the transport mechanisms for these compounds are predominantly situated in the blood-facing aspect of the secretory cells. In addition. it is apparent that there is a polarized distribution of transport systems (carriers and channels) which accept sodium, potassium, chloride, phosphate, and calcium as substrates.

Rapidly reversible binding of rabbit prolactin to the rabbit prolactin receptor accounts for the differences between homologous and heterologous binding.

The binding of radioiodinated rabbit (rb) prolactin (PRL) to rabbit mammary membranes is low and its affinity constant, 0.02 nM-1, calculated from heterologous inhibition assays, is about 300 times lower than that of ovine (o) PRL. Although the differences between homologous and heterologous binding are well documented in different species, the reasons for such differences are still unknown. Here we show that the low affinity of rbPRL for the native receptor does not affect its in vitro bioactivity compared with that of oPRL. We also show that rbPRL displays high specific binding to the baculovirus-expressed recombinant receptor and further establish that its lower affinity for binding to the homologous receptor is due to its faster and more complete dissociation compared with that of oPRL. Hormone binding affinity for full-length and carboxy-terminal truncated rbPRL receptor mutants expressed in mammalian or in baculovirus-infected cells was not affected by partial truncation of the cytoplasmic domain of the receptor, whereas the affinity for oPRL increased and that for rbPRL decreased upon truncation of both the cytoplasmic and membrane domains. The affinity of rbPRL for the native receptor is two orders of magnitude lower than that for the recombinant receptor. Affinity cross-linking and binding experiments showed that this difference in affinities is not related to selective cleavage of the native microsomal receptor during the binding reaction; however, this difference may be related to cell context-dependent differences in the oligomerization state of the receptor. Thus, obviously, the cloned receptor is alone sufficient for binding to rbPRL without requiring any receptor-associated protein. The lower affinity for rbPRL binding to its homologous receptor in comparison with higher affinity binding of oPRL to the same receptor is attributable to differences in their dissociation kinetics and in the conformational requirements of the receptor-hormone interaction site for binding to the two hormones.

Feedback control of milk secretion from milk.

Extracellular storage allows biologically-active substances in milk to influence mammary function. Among these factors is one which regulates the rate of milk secretion acutely according to frequency or completeness of milk removal in each mammary gland. The active factor in goat's milk has been identified by screening milk constituents for their ability to inhibit milk constituent secretion in tissue and cell culture bioassays, and found to be a novel milk protein. The proteins identified by bioassy in vitro, also inhibited milk secretion in lactating goats in a reversible, concentration-dependent manner. This protein, termed FIL (feedback inhibitor of lactation), acts by reversible blockade of constitutive secretion in the mammary epithelial cell. As the inhibitor is synthesized in the same epithelial cells, feedback inhibition is, therefore, an autocrine mechanism. FIL's unusual mechanism of action also influences other aspects of mammary function. Acute disruption of mammary membrane trafficking is associated with downregulation of prolactin receptors and followed by a decrease in epithelial cell differentiation. Thus, in addition to acutely-regulating milk secretion, FIL may induce the adaptation in mammary cell differentiation which acts in vivo to sustain the secretory response to a sustained change in milk removal. In the long term, matching of milk output to demand is achieved by a change in mammary cell number. This developmental response is also local in nature. Whether it too is due to autocrine modulation by FIL of mechanisms influencing cell proliferation or survival, or elicited by another milk-borne factor, remains to be determined.

Characterization of Binding of Transforming Growth Factor-β1 to Bovine Mammary Membranes

This study developed a procedure to measure binding of transforming growth factor-β1 to bovine mammary membranes. Mammary membranes were incubated with 3 M MgCl2 to remove endogenously bound transforming growth factor-β1. Binding was optimized by incubation of 200μg of membrane protein with 125I-labeled transforming growth factor-β1 in 25mM Tris, 10mM CaCl2, and 1.0% BSA in a total volume of .5ml in the presence or absence of unlabeled transforming growth factor-β1. The reaction equilibrated in 2h at 4°C. Specific binding was linear from 142 to 1140μg of membrane protein. The reaction was specific for the beta transforming growth factors; transforming growth factor-β1, transforming growth factor-β2, and transforming growth factor-β3 could compete effectively with 125I-labeled transforming growth factor-β1 for the receptor. The growth factors, epidermal growth factor, IGF-I, or transforming growth factor-α did not compete effectively with 125I-labeled transforming growth factor-β1 for binding to bovine mammary membranes. Scatchard analysis showed that the number of receptors averaged 251 pmol/mg of membrane protein and the affinity was 8.7 × l0−11 M. Binding to mammary membranes was higher during the prepubertal and pubertal periods than during lactation. Binding to mammary membranes during early lactation averaged 24% of the binding observed during other physiological states. Therefore, transforming growth factor-β1 binds specifically to bovine mammary tissue and varies with stages of development and differentiation, indicating that the transforming growth factor-β1 receptors may play a role in regulating the activity of transforming growth factor-β1 in the mammary gland.

Effect of bovine GH and ovariectomy on mammary tissue sensitivity to IGF-I in prepubertal heifers

Sixteen prepubertal Holstein Friesian heifers were used to study the effect of long-term administration of bovine GH (bGH; 15 mg/day for 15 weeks) and ovariectomy (at 2.5 months of age) on GH and IGF-I receptors in mammary and liver parenchymal tissue. Heifers were slaughtered on the day after the last injection of bGH or excipient. Mammary and liver parenchymal tissue was collected for preparation of crude membranes for ligand-binding assays. Specific binding of [125I]IGF-I to mammary membranes increased linearly with increasing membrane protein concentration and was displaced by IGF-I, less effectively by IGF-II and least effectively by insulin. Linear Scatchard plots were obtained showing the presence of a single class of high-affinity binding sites for IGF-I in prepubertal mammary tissue. Specific binding of IGF-I was unaffected by bGH treatment but tended to be higher (P < 0.10) in membranes from ovariectomized heifers. Hepatic binding of GH and IGF-I was unaffected by bGH treatment and ovariectomy. However, specific binding of GH tended to be lower (P < 0.06) in liver tissue from bGH-treated intact heifers compared with control heifers. The present study confirms the presence of IGF-I receptors in prepubertal mammary tissue in agreement with the suggested involvement of IGF-I in the mechanism of action of GH on mammary glands in this period. The results, however, indicate that the stimulatory effect of GH on mammary growth is mediated via increased serum concentration of IGF-I and not by increased IGF-I binding to its mammary receptors.(ABSTRACT TRUNCATED AT 250 WORDS)

Prolactin receptor and signal transduction to milk protein genes.

After cloning of the mammary gland prolactin (PRL) receptor cDNA, a functional assay was established using co-transfection of PRL receptor cDNA together with a milk protein promoter/chloramphenicol acetyl transferase (CAT) construct in Chinese hamster ovary (CHO) cells. Different mutants of the PRL receptor were tested in this CAT assay to delimit the domains in the receptor necessary for signal transduction to milk protein genes. In CHO cells stably transfected with PRL receptor cDNA, high numbers of PRL receptor are expressed. By metabolic labeling and immunoprecipitation, expressed PRL receptor was identified as a single species of 100 kDa. Using these cells, we analyzed the effects of PRL on intracellular free Ca++ concentration. PRL stimulates Ca++ entry and induces secondary Ca++ mobilization. The entry of Ca++ is a result of an increase in K+ conductance that hyperpolarizes the membranes. We have also analyzed tyrosine phosphorylation induced by PRL. In CHO cells stably transfected with PRL receptor cDNA, PRL induced a very rapid and transient tyrosine phosphorylation of a 100-kDa protein which is most probably the PRL receptor. The same finding was obtained in mammary membranes after PRL injection to lactating rabbits. Whereas tyrosine kinase inhibitors genistein and lavendustin were without effect, PRL stimulation of milk protein gene promoters was partially inhibited by 2 microM herbimycin in CHO cells co-transfected with PRL receptor cDNA and the beta lactoglobulin CAT construct. Taken together these observations indicate that the cytoplasmic domain of the PRL receptor interacts with one or several tyrosine kinases, which may represent early postreceptor events necessary for PRL signal transduction to milk protein genes.

2-[125I]iodomelatonin binding sites in murine mammary tissue.

2-[125I]iodomelatonin binds in vitro to murine mammary gland membranes, in a specific, saturable, reversible and temperature-dependent and pH-dependent way, displaying an affinity within the low nM range. The order of affinity to displace the radioligand from mammary membranes was: 6-hydroxy-melatonin > 5-hydroxyindoleamines > noradrenaline > 6-chloromelatonin > 5-methoxyindoles (melatonin included). There was a diurnal rhythm in Kd and Bmax of 2-[125I]iodomelatonin binding, displaying the highest affinity and the lowest binding site concentration during the scotophase. The functional significance of melatonin binding sites was indicated by changes in binding parameters related to different rates of mammary growth during puberty, estrous cycle, pregnancy, lactation and the postlactation period.

Endogenous ligands for the epidermal growth factor receptor in human breast tumors do not interfere in an epidermal growth factor receptor radioligand binding assay.

Ligands for the epidermal growth factor receptor (EGFR) (e.g. EGF and transforming growth factor alpha, TGF alpha) may be included in membrane preparations from human breast tumors, either complexed or not to EGFR. Assessment of EGFR in human placental membranes (HPM) after preincubation with human EGF (hEGF) or human TGF alpha (hTGF alpha) indicated that the presence of these ligands in a membrane preparations did not affect the apparent number of binding sites, but only resulted in an increased apparent dissociation constant (Kd). To obviate the effect of endogenously bound EGFR ligands in a radioligand binding assay, an acid treatment procedure has been used recently. In the present study we show that acid treatment of mammary tumor membranes does not result in increased EGFR levels but does result in an EGFR enriched membrane preparation by elimination of contaminating cytosol proteins. This however also occurred by washing the membranes with assay buffer at neutral pH. From these experiments we conclude that endogenous ligands, if present in membrane fractions from human breast tumors and occupying EGFR, do not interfere in a multipoint radioligand binding assay.

Quantitative dot blot analyses of blood‐group‐related antigens in paired normal and malignant human breast tissues

Membranes were prepared from 31 breast-cancer specimens and adjacent mammary tissues, dot-blotted to nitrocellulose paper, and reacted with monoclonal antibodies (MAbs) (A, B, Lewis a, Lewis b, sialylated Lewis a, Lewis x, and Lewis y) and lectins (Ulex europaeus, peanut agglutinin) having various blood-group specificities. The expression of epithelial membrane antigen was assayed with MAb MA5. The ratio of breast-cancer to normal mammary membrane preparations (C/N ratios) of these reagents was measured by densitometric scanning. We observed a decrease in the levels of A, B, Lewis a, Lewis b, sialylated Lewis a, and Lewis y antigens and an increase of Lewis x, T, and MA5-reactive determinants in breast cancers. The incidence of incompatible A, as well as A and B, antigens was demonstrated for 2 patients of blood group B and O respectively. When the receptor content was plotted against the C/N ratio of these various reagents, a significant inverse relationship between the C/N ratio of Lewis x antigen and estrogen (ER) and progesterone receptor (PR) content was observed in breast cancers. The mean C/N ratio of Lewis x antigen was significantly higher in the ER-negative/PR-negative (ER-/PR-; 2.33 +/- 1.17), as compared with the ER-positive/PR-positive (ER+/PR+; 0.97 +/- 0.80). According to these observations, Lewis x antigen expression may be influenced by hormonal stimuli such as estrogen and progesterone.

Dietary fat composition influences fatty acid composition of milk fat globule membrane in lactating cows

Milk fat globule membranes are derived directly from the apical plasma membrane of mammary epithelial cells. To evaluate the effect of dietary fat on mammary membranes, we determined the fatty acid composition of the milk fat globule membrane in lactating dairy cows fed diets supplemented with fats of different fatty acid composition, or infused intravenously with soy oil emulsion. A preliminary survey, using an abbreviated preparation procedure (membranes isolated at 48,000 x g-max for 15 min), yielded about 45% of the total membrane fatty acids that could be recovered by centrifuging at the same speed for 120 min, and showed that changes in fatty acid composition of membranes reflected dietary fatty acids to some extent. Dietary palmitic acid increased the content of 16:0 in the membranes. A high corn diet increased ruminal formation of t18:1, and its level increased to 12% of membrane fatty acids. Infusion of soy oil emulsion increased 18:2 membrane content, and decreased the levels of 18:1 and 20:4. All treatments decreased the ratio of unsaturated/saturated fatty acids as compared to controls, whereas the ratio of polyunsaturated/saturated fatty acids was increased by feeding a high corn diet or by infusing soy oil. The ratio of 18:2/c18:1 increased from 0.31 to 1.0 after infusing soy oil for 4 days. The fatty acids of membranes isolated upon 120-min centrifugation were slightly more saturated. The differences were not sufficiently large, however, to affect overall results significantly.

Insulin Promotes the Release from Rat Mammary Gland Plasma Membranes of a Factor Which Activates Mitochondrial Pyruvate Dehydrogenase

Insulin Promotes the Release from Rat Mammary Gland Plasma Membranes of a Factor Which Activates Mitochondrial Pyruvate Dehydrogenase

Binding and bioactivity of ovine and porcine prolactins in porcine mammary tissue

Differential binding of homologous and heterologous prolactin was investigated in porcine mammary tissue. Specific binding of ovine prolactin to porcine mammary membranes or tissue slices was significantly greater than specific binding of the homologous porcine prolactin. Ovine prolactin was also more potent than porcine prolactin in stimulating proliferation of Nb2 cells. In contrast, stimulation of glucose metabolism in porcine mammary explants by porcine prolactin was greater than that by ovine prolactin. Differences in specific binding were probably not due to damage during iodination, as low concentrations of iodinated prolactins were similar to unlabelled prolactins in their abilities to stimulate proliferation of Nb2 cells. Furthermore, electrophoretic analysis of medium from binding reactions suggested that differences in specific binding were not due to proteolytic cleavage of the homologous prolactin into large (greater than 10 kDa) fragments. These studies suggest that ovine prolactin either binds to sites in addition to the authentic lactogenic receptor in porcine mammary tissue or that a significantly higher affinity of ovine prolactin for the porcine lactogenic receptor has little effect on its biological activity.