scholarly journals Antibodies to the major insoluble milk fat globule membrane-associated protein: specific location in apical regions of lactating epithelial cells.

1981 ◽  
Vol 89 (3) ◽  
pp. 485-494 ◽  
Author(s):  
W W Franke ◽  
H W Heid ◽  
C Grund ◽  
S Winter ◽  
C Freudenstein ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Milk Fat ◽  
Lipid Production ◽  
Apical Cell ◽  
Cell Types ◽  
Specific Marker ◽  
Milk Lipid ◽  
Apical Surface ◽  
Milk Fat Globule ◽  
A Cell

Milk lipid globules of various species are surrounded by a membrane structure that is separated from the triglyceride core of the globule by a densely staining fuzzy coat layer of 10- to 50-nm thickness. This internal coat structure remains attached to the membrane during isolation and extraction with low- and high-salt buffers, is insoluble in nondenaturing detergents, and is enriched in an acidic glycoprotein (butyrophilin) with an apparent Mr of 67,000. Guinea pig antibodies against this protein, which show cross-reaction with the corresponding protein in some (goat) but not other (human, rat) species, have been used for localization of butyrophilin on frozen sections of various tissues from cow by immunofluorescence and electron microscopy. Significant reaction is found only in milk-secreting epithelial cells and not in other cell types of mammary gland and various epithelial tissues. In milk-secreting cells, the staining is restricted to the apical cell surface, including budding milk lipid globules, and to the periphery of the milk lipid globules contained in the alveolar lumina. These findings indicate that butyrophilin, which is constitutively secreted by surface budding in coordination with milk lipid production, is located at the apical surface and is not detected at basolateral surfaces, in endoplasmic reticulum, and in Golgi apparatus. This protein structure represents an example of a cell type-specific cytoskeletal component in a cell apex. It is suggested that this antigen provides a specific marker for the apical surface of milk-secreting cells and that butyrophilin is involved in the vectorial discharge of milk lipid globules.

scholarly journals Characterization of an apically derived epithelial membrane glycoprotein from bovine milk, which is expressed in capillary endothelia in diverse tissues.

1985 ◽  
Vol 100 (2) ◽  
pp. 397-408 ◽  
Author(s):  
D E Greenwalt ◽  
I H Mather
Keyword(s):  
Endothelial Cells ◽  
Epithelial Cells ◽  
Milk Fat ◽  
Bovine Milk ◽  
Mammary Tissue ◽  
Specific Marker ◽  
Total Carbohydrate ◽  
Milk Fat Globule Membrane ◽  
Milk Fat Globule ◽  
Fat Globule

A glycoprotein (PAS IV) of apparent Mr 76,000 was purified from bovine milk-fat-globule membrane and partially characterized. PAS IV contained mannose, galactose, and sialic acid as principal sugars (approximately 5.3% total carbohydrate [wt/wt]) and existed in milk in at least four isoelectric variants. The glycoprotein appeared to be an integral membrane protein by several criteria. PAS IV was recovered in the detergent phase of Triton X-114 extracts of milk-fat-globule membrane at room temperature. When bound to membrane, PAS IV was resistant to digestion by a number of proteinases, although after solubilization with non-ionic detergents, the protein was readily degraded. Amino acid analysis of the purified protein revealed a high percentage of amino acids with nonpolar residues. The location of PAS IV was determined in bovine tissues by using immunofluorescence techniques. In mammary tissue, PAS IV was located on both the apical surfaces of secretory epithelial cells and endothelial cells of capillaries. This glycoprotein was also detected in endothelial cells of heart, liver, spleen, pancreas, salivary gland, and small intestine. In addition to mammary epithelial cells, PAS IV was also located in certain other epithelial cells, most notably the bronchiolar epithelial cells of lung. The potential usefulness of this protein as a specific marker of capillary endothelial cells in certain tissues is discussed.

Multiple forms of lactadherin (breast antigen BA46) and butyrophilin are secreted into human milk as major components of milk fat globule membrane

1999 ◽  
Vol 66 (2) ◽  
pp. 295-301 ◽  
Author(s):  
MARIA CAVALETTO ◽  
MARIA G. GIUFFRIDA ◽  
CARLO GIUNTA ◽  
CAMILLO VELLANO ◽  
CLAUDIO FABRIS ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Molecular Mass ◽  
Milk Fat ◽  
Cell Surface Receptor ◽  
Sequencing Analysis ◽  
Apical Surface ◽  
Multiple Forms ◽  
Milk Fat Globule Membrane ◽  
Milk Fat Globule ◽  
Fat Globule

In the lactating mammary gland, epithelial cells secrete triacylglycerols in the form of droplets enveloped by an apical surface membrane. This membrane is known as the milk fat globule membrane (MFGM; Mather & Keenan, 1983). MFGM-associated proteins have been studied and employed in developing antibodies against surface antigens of breast epithelial cells, which are used in breast cancer immunodiagnosis and histopathology (Salinas et al. 1987; Larocca et al. 1991; Peterson et al. 1995). So far, only a small number of proteins have been analysed and their sequence identified in bovine, murine and human MFGM; among them are butyrophilin (Jack & Mather, 1990; Ishii et al. 1995; Taylor et al. 1996), MFG-E8 (Stubbs et al. 1990), PAS 6/7 (Hvarregaard et al. 1996) and lactadherin or breast antigen BA46 (Couto et al. 1996; Taylor et al. 1997). Several minor proteins have yet to be characterized, since it is not easy to isolate them in large quantities from the membrane. SDS gel patterns give useful information about MFGM proteins, such as concentration, relative molecular mass and presence of carbohydrate. Over forty membrane components have been separated by electrophoretic techniques from bovine MFGM (Mather et al. 1980).The research reported here combined SDS-PAGE with sequencing analysis and describes the composition of human MFGM, with the exception of high molecular mass mucin, which only penetrates an acrylamide gel of 40 g/l. Mucins have been extensively studied and the sequence predicted from cDNA (Gendler et al. 1990). Surprisingly, identification of the protein bands in the present study revealed that three proteins alone constituted the major components of human MFGM: xanthine oxidase (EC 1.1.3.22), butyrophilin and lactadherin. Lactadherin belongs to a family of proteins possessing epidermal growth factor-like and factor V/VIII C1/C2-like domains, including bovine PAS 6/7, guinea pig GP55 (Hvarregaard et al. 1996) and murine MFG-E8 (Stubbs et al. 1990). In a previous investigation, we characterized lactadherin (formerly breast antigen BA46) and its truncated 30 kDa form as components of healthy human MFGM (Giuffrida et al. 1998). Human butyrophilin has recently been cloned and sequenced (Taylor et al. 1996); the presence of two extracellular immunoglobulin superfamily domains suggested a potential cell surface receptor function. This study was aimed at identifying and characterizing the multiple forms of the major proteins of MFGM.

scholarly journals Milk Fat Globule Membrane Proteome and Micronutrients in the Milk Lipid Fraction: Insights into Milk Bioactive Compounds

Dairy ◽  
2021 ◽  
Vol 2 (2) ◽  
pp. 202-217
Author(s):  
Michele Manoni ◽  
Donata Cattaneo ◽  
Sharon Mazzoleni ◽  
Carlotta Giromini ◽  
Antonella Baldi ◽  
...  
Keyword(s):  
Milk Fat ◽  
Lipid Fraction ◽  
Value Added ◽  
Milk Lipid ◽  
Milk Fat Globule Membrane ◽  
Fat Globules ◽  
Milk Fat Globules ◽  
Milk Fat Globule ◽  
Fat Globule ◽  
Value Added Products

Milk lipids are composed of milk fat globules (MFGs) surrounded by the milk fat globule membrane (MFGM). MFGM protects MFGs from coalescence and enzymatic degradation. The milk lipid fraction is a “natural solvent” for macronutrients such as phospholipids, proteins and cholesterol, and micronutrients such as minerals and vitamins. The research focused largely on the polar lipids of MFGM, given their wide bioactive properties. In this review we discussed (i) the composition of MFGM proteome and its variations among species and phases of lactation and (ii) the micronutrient content of human and cow’s milk lipid fraction. The major MFGM proteins are shared among species, but the molecular function and protein expression of MFGM proteins vary among species and phases of lactation. The main minerals in the milk lipid fraction are iron, zinc, copper and calcium, whereas the major vitamins are vitamin A, β-carotene, riboflavin and α-tocopherol. The update and the combination of this knowledge could lead to the exploitation of the MFGM proteome and the milk lipid fraction at nutritional, biological or technological levels. An example is the design of innovative and value-added products, such as MFGM-supplemented infant formulas.

Adhesion of uric acid crystals to the surface of renal epithelial cells

2000 ◽  
Vol 278 (6) ◽  
pp. F989-F998 ◽  
Author(s):  
Rima M. Koka ◽  
Erick Huang ◽  
John C. Lieske
Keyword(s):  
Uric Acid ◽  
Epithelial Cells ◽  
Cell Surface ◽  
Hydrophobic Interactions ◽  
Calcium Phosphates ◽  
Apical Cell ◽  
Apical Surface ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Renal Tubular Cells

Adhesion of microcrystals that nucleate in tubular fluid to the apical surface of renal tubular cells could be a critical step in the formation of kidney stones, 12% of which contain uric acid (UA) either alone or admixed with calcium oxalates or calcium phosphates. UA crystals bind rapidly to monolayer cultures of monkey kidney epithelial cells (BSC-1 line), used to model the surface of the nephron, in a concentration-dependent manner. The urinary glycoproteins osteopontin, nephrocalcin, and Tamm-Horsfall glycoprotein had no effect on binding of UA crystals to the cell surface, whereas other polyanions including specific glycosaminoglycans blocked UA crystal adhesion. Specific polycations also inhibited adhesion of UA crystals and appeared to exert their inhibitory effect by coating cells. However, removal of anionic cell surface molecules with neuraminidase, heparitinase I, or chondroitinase ABC each increased UA crystal binding, and sialic acid-binding lectins had no effect. These observations suggest that hydrogen bonding and hydrophobic interactions play a major role in adhesion of electrostatically neutral UA crystals to renal cells, unlike the interaction of calcium-containing crystals with negatively charged molecules on the apical cell surface via ionic forces. After adhesion to the plasma membrane, subsequent cellular events could contribute to UA crystal retention in the kidney and the development of UA or mixed calcium and UA calculi.

Epithelial sorting of a glycosylphosphatidylinositol-anchored bacterial protein expressed in polarized renal MDCK and intestinal Caco-2 cells

1995 ◽  
Vol 108 (1) ◽  
pp. 369-377 ◽  
Author(s):  
K.L. Soole ◽  
M.A. Jepson ◽  
G.P. Hazlewood ◽  
H.J. Gilbert ◽  
B.H. Hirst
Keyword(s):  
Epithelial Cells ◽  
Cell Surface ◽  
Intestinal Epithelial Cells ◽  
Mdck Cells ◽  
Basolateral Membrane ◽  
Apical Cell ◽  
Apical Surface ◽  
Intestinal Epithelial ◽  
Gpi Anchor ◽  
Sorting Signal

To evaluate whether a glycosylphosphatidylinositol (GPI) anchor can function as a protein sorting signal in polarized intestinal epithelial cells, the GPI-attachment sequence from Thy-1 was fused to bacterial endoglucanase E' (EGE') from Clostridium thermocellum and polarity of secretion of the chimeric EGE'-GPI protein was evaluated. The chimeric EGE'-GPI protein was shown to be associated with a GPI anchor by TX-114 phase-partitioning and susceptibility to phosphoinositol-specific phospholipase C. In polarized MDCK cells, EGE' was localized almost exclusively to the apical cell surface, while in polarized intestinal Caco-2 cells, although 80% of the extracellular form of the enzyme was routed through the apical membrane over a 24 hour period, EGE' was also detected at the basolateral membrane. Rates of delivery of EGE'-GPI to the two membrane domains in Caco-2 cells, as determined with a biotinylation protocol, revealed apical delivery was approximately 2.5 times that of basolateral. EGE' delivered to the basolateral cell surface was transcytosed to the apical surface. These data indicate that a GPI anchor does represent a dominant apical sorting signal in intestinal epithelial cells. However, the mis-sorting of a proportion of EGE'GPI to the basolateral surface of Caco-2 cells provides an explanation for additional sorting signals in the ectodomain of some endogenous GPI-anchored proteins.

Regeneration of mammary glands in vivo from isolated mammary ducts

Development ◽  
1986 ◽  
Vol 96 (1) ◽  
pp. 229-243
Author(s):  
E. Jane Ormerod ◽  
Philip S. Rudland
Keyword(s):  
Epithelial Cells ◽  
Milk Fat ◽  
Cell Types ◽  
Mammary Glands ◽  
Myoepithelial Cells ◽  
Basal Cells ◽  
Alveolar Cells ◽  
Terminal End Buds ◽  
Mammary Cell

Rat mammary ducts, free of buds, can alone regenerate complete mammary trees when transplanted into the interscapular fat pads of syngeneic host rats. All the main mammary cell types are identified within such outgrowths. Epithelial cells, which show the presence of milk fat globule membrane antigens and microvilli on their luminal surfaces, line the ducts. Basal cells surrounding the ducts show characteristic features of myoepithelial cells: immunoreactive actin and keratin within the cytoplasm, myofilaments, pinocytotic vesicles and hemidesmosomal attachments to the basement membrane. Cells within the end buds and lateral buds, however, show few if any cytoplasmic myofilaments and are relatively undifferentiated in appearance. Intermediate morphologies between these cells and myoepithelial cells are seen nearer the ducts. In this respect they exactly resemble the cap cells found in terminal end buds (TEBs) of normal mammary glands. Occasional epithelial cells within alveolar buds show the presence of immunoreactive casein, which is a product of secretory alveolar cells in the normal rat mammary gland. Dissected terminal end buds can regenerate similar ductal outgrowths. Thus, ductal tissue alone can generate all the major mammary cell types seen in the normal gland, including the cap cells.

scholarly journals Comparative effect of bovine buttermilk, whey, and lactoferrin on the innate immunity receptors and oxidative status of intestinal epithelial cells.

2020 ◽  
Author(s):  
Berta Buey ◽  
Andrea Bellés ◽  
Eva Latorre ◽  
Inés Abad ◽  
María Dolores Pérez ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Lipid Peroxidation ◽  
Innate Immunity ◽  
Epithelial Cells ◽  
Milk Fat ◽  
Functional Foods ◽  
Intestinal Epithelial Cells ◽  
Protein Carbonyl ◽  
Intestinal Epithelial ◽  
Milk Fat Globule

Milk contains active molecules with important functional properties as the defensive proteins; among them are the whey protein lactoferrin and proteins of the milk fat globule membrane (MFGM) present in buttermilk. The aim of this study has been to investigate the effect of lactoferrin, whey and buttermilk as modulators of intestinal innate immunity and oxidative stress on intestinal epithelial cells, to evaluate its potential use for the development of functional foods. Innate immune Toll-like receptors (TLR2, TLR4, and TLR9) mRNA expression, lipid peroxidation (MDA+4-HDA) and protein carbonyl levels were analyzed in enterocyte-like Caco-2/TC7 cells treated for 24 hours with different concentrations of lactoferrin, whey or buttermilk. None of the substances analyzed caused oxidative damage; however, whey significantly decreased the levels of lipid peroxidation. Furthermore, both lactoferrin and whey were able to reduce the oxidative stress induced by lipopolysaccharide. Respect to TLR receptors, lactoferrin, whey and buttermilk specifically altered the expression of TLR2, TLR4 and TLR9 receptors, with a strong decrease in TLR4 expression. These results suggest that lactoferrin, whey and buttermilk could be interesting potential ingredients for functional foods as they seem to modulate oxidative stress and inflammatory response induced by TLRs activation.

scholarly journals Characterization of epithelial membrane antigen expression in human mammary epithelium by ultrastructural immunoperoxidase cytochemistry.

1986 ◽  
Vol 34 (6) ◽  
pp. 801-809 ◽  
Author(s):  
O W Petersen ◽  
B van Deurs
Keyword(s):  
Epithelial Cells ◽  
Cell Surface ◽  
Milk Fat ◽  
Breast Biopsy ◽  
Intracellular Localization ◽  
Surface Membrane ◽  
Carcinoma Cells ◽  
Annulate Lamellae ◽  
Apical Surface ◽  
Human Breast Carcinomas

Ultrastructural immunocytochemistry was used to analyze cell surface distribution and intracellular localization of milk fat globule membrane antigen (MFGM-A) in cryosections from human breast carcinomas and benign breast biopsy specimens. The specimens were fixed in formaldehyde and frozen. Cryostat sections were cut at 15 micron, incubated with mouse monoclonal antibody to MFGM-A, and then with a peroxidase-conjugated goat anti-mouse antibody. After glutaraldehyde fixation, the sections were incubated with diaminobenzidine-H2O2 and further processed for electron microscopy. MFGM-A was specific for epithelial cells. MFGM-A staining was strictly confined to the apical surface membrane of normal ductal epithelium, never involving basolateral membranes below the tight junctions. In normal epithelial cells, MFGM-A was readily detected in cisternae of the endoplasmic reticulum (ER), but only to a lesser extent in Golgi complexes and presumptive secretory vesicles. In carcinoma cells, surface staining for MFGM-A was either distributed in a non-polarized manner on the entire cell surface or else was totally absent. In some carcinoma cells without surface-associated MFGM-A, very pronounced intracellular MFGM-A staining was seen in the ER, in the nuclear envelope, and in annulate lamellae. The observations on MFGM-A expression were supported by studies on a cell culture model system.

Milk fat globule is an alternative to mammary epithelial cells for gene expression analysis in buffalo

2016 ◽  
Vol 83 (2) ◽  
pp. 202-208 ◽  
Author(s):  
Qiuming Chen ◽  
Yanjun Wu ◽  
Mingyuan Zhang ◽  
Wenwen Xu ◽  
Xiaoping Guo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Mammary Gland ◽  
Epithelial Cells ◽  
Expression Analysis ◽  
Milk Fat ◽  
Gene Expression Analysis ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Milk Fat Globule ◽  
Fat Globule

Owing to the difficulty in obtaining mammary gland tissue from lactating animals, it is difficult to test the expression levels of genes in mammary gland. The aim of the current study was to identify if milk fat globule (MFG) in buffalo milk was an alternative to mammary gland (MG) and milk somatic cell (MSC) for gene expression analysis. Six buffalos in late lactation were selected to collect MFG and MSC, and then MG was obtained by surgery. MFG was stained with acridine orange to successfully visualise RNA and several cytoplasmic crescents in MFG. The total RNA in MFG was successfully isolated and the integrity was assessed by agarose gel electrophoresis. We analysed the cellular components in MFG, MG and MSC through testing the expression of cell-specific genes by qRT-PCR. The results showed that adipocyte-specific gene (AdipoQ) and leucocyte-specific genes (CD43, CSF1 and IL1α) in MFG were not detected, whereas epithelial cell marker genes (Keratin 8 and Keratin 18) in MFG were higher than in MSC and lower than in MG, fibroblast marker gene (vimentin) in MFG was significantly lower than in MG and MSC, milk protein genes (LALBA, BLG and CSN2) and milk fat synthesis-related genes (ACC, BTN1A1, FABP3 and FAS) in MFG were higher than in MG and MSC. In conclusion, the total RNA in MFG mainly derives from mammary epithelial cells and can be used to study the functional gene expression of mammary epithelial cells.

scholarly journals Differential Infection of Polarized Epithelial Cell Lines by Sialic Acid-Dependent and Sialic Acid-Independent Rotavirus Strains

2001 ◽  
Vol 75 (23) ◽  
pp. 11834-11850 ◽  
Author(s):  
Max Ciarlet ◽  
Sue E. Crawford ◽  
Mary K. Estes
Keyword(s):  
Sialic Acid ◽  
Epithelial Cell ◽  
Epithelial Cells ◽  
Cell Lines ◽  
Apical Cell ◽  
Paracellular Permeability ◽  
Apical Surface ◽  
Apical Cell Membrane ◽  
Basolateral Side ◽  
Epithelial Cell Lines

ABSTRACT Infection of epithelial cells by some animal rotaviruses, but not human or most animal rotaviruses, requires the presence ofN-acetylneuraminic (sialic) acid (SA) on the cell surface for efficient infectivity. To further understand how rotaviruses enter susceptible cells, six different polarized epithelial cell lines, grown on permeable filter membrane supports containing 0.4-μm pores, were infected apically or basolaterally with SA-independent or SA-dependent rotaviruses. SA-independent rotaviruses applied apically or basolaterally were capable of efficiently infecting both sides of the epithelium of all six polarized cell lines tested, while SA-dependent rotaviruses only infected efficiently through the apical surface of five of the polarized cell lines tested. Regardless of the route of virus entry, SA-dependent and SA-independent rotaviruses were released almost exclusively from the apical domain of the plasma membrane of polarized cells before monolayer disruption or cell lysis. The transepithelial electrical resistance (TER) of cells decreased at the same time, irrespective of whether infection with SA-independent rotaviruses occurred apically or basolaterally. The TER of cells infected apically with SA-dependent rotaviruses decreased earlier than that of cells infected basolaterally. Rotavirus infection decreased TER before the appearance of cytopathic effect and cell death and resulted in an increase in the paracellular permeability to [3H]inulin as a function of loss of TER. The presence of SA residues on either the apical or basolateral side was determined using a Texas Red-conjugated lectin, wheat germ agglutinin (WGA), which binds SA residues. WGA bound exclusively to SA residues on the apical surface of the cells, confirming the requirement for SA residues on the apical cell membrane for efficient infectivity of SA-dependent rotaviruses. These results indicate that the rotavirus SA-independent cellular receptor is present on both sides of the epithelium, but SA-dependent and SA-independent rotavirus strains infect polarized epithelial cells by different mechanisms, which may be relevant for pathogenesis and selection of vaccine strains. Finally, rotavirus-induced alterations of the epithelial barrier and paracellular permeability suggest that common mechanisms of pathogenesis may exist between viral and bacterial pathogens of the intestinal tract.

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