Omneity® Pellets
All-In-One Vitamin & Mineral Pellet
Topic:Cell Culture
Cell culture in horses involves the in vitro cultivation of equine cells under controlled conditions. This technique is employed to study various cellular processes, including growth, differentiation, and response to external stimuli, in an isolated environment. Equine cell cultures can be derived from various tissues, such as skin, muscle, or bone, and are used in a range of research applications, including genetic studies, drug testing, and disease modeling. These cultures provide a valuable platform for understanding cellular mechanisms and developing therapeutic strategies. This page compiles peer-reviewed research studies and scholarly articles that explore the methodologies, applications, and findings related to cell culture in equine research.
Culture of 5-day horse embryos in microdroplets for 10 to 20 days. Embryos were recovered from the uteri of mares 5 d after ovulation. Six embryos, all morulae, were placed singly in 200-ul droplets of Ham's F-12 with 10% fetal calf serum and cultured at 37 degrees C in a 5% CO(2) atmosphere. The embryos expanded to form blastocysts by the third day of culture. The blastocysts hatched from their zona pellucida, rather than the zona thinning and flaking off, as occurs in vivo. Hatching from the zona pellucida began on the third day of culture and was complete in five of six embryos by the sixth day. The embryonic capsule, normally present in equine embryos aft...
Modulation of an adhesion-related surface antigen on equine neutrophils by bacterial lipopolysaccharide and antiinflammatory drugs. The essential role of the CD11/CD18 family of leukocyte adhesion molecules (LeuCams) in neutrophil-substrate adhesion is well documented. We have found that a monoclonal antibody designated 60.3 (MoAb 60.3) that recognizes the common beta-subunit (CD18) on human neutrophils (PMN) also recognizes a surface antigen on equine PMN. Antigen expression as assessed by immunofluorescence flow cytometry was enhanced by zymosan-activated serum (ZAS) or phorbol 12-myristate 13-acetate (PMA) stimulation. Pretreatment of equine PMN with MoAb 60.3 inhibited ZAS-stimulated aggregation, indicating that the mo...
An indirect sandwich ELISA utilising F(ab’)2 fragments for the detection of African horsesickness virus. African horsesickness virus (AHSV), an important disease of equines is caused by an orbivirus. Because of the need to contain the spread of the disease, it is often essential to make a rapid diagnosis. For this purpose, an ELISA capable of detecting viral antigen in animal tissue and in cell culture fluid was developed. Immobilised F(ab')2 fragments prepared by digestion of AHSV-specific IgG with pepsin were used to trap virus from tissue homogenates or cell culture supernatant. After addition of intact IgG as detecting antibody, Staphylococcus aureus protein A labelled with horseradish peroxi...
Cloning and characterization of cDNAs encoding equine infectious anemia virus tat and putative Rev proteins. We isolated and characterized six cDNA clones from an equine infectious anemia virus-infected cell line that displays a Rev-defective phenotype. With the exception of one splice site in one of the clones, all six cDNAs exhibited the same splicing pattern and consisted of four exons. Exon 1 contained the 5' end of the genome; exon 2 contained the tat gene from mid-genome; exon 3 consisted of a small section of env, near the 5' end of the env gene; and exon 4 contained the putative rev open reading frame from the 3' end of the genome. The structures of the cDNAs predict a bicistronic message in ...
Separation of equine bronchopulmonary lavage cells by density gradient centrifugation and expression of procoagulant activity in unpurified cells and cell subpopulations. Bronchopulmonary lavage was performed in 10 healthy horses and in 39 horses with chronic pulmonary disease. The predominant cell types were macrophages in healthy horses and neutrophils in severely diseased horses. Procoagulant activity (PCA) was detected in all 32 cell-free supernatants examined and in all 49 unpurified cell suspensions. Cells were separated by centrifugation on discontinuous gradients prepared either with Percoll or with Metrizamide. Macrophages were enriched in subpopulations of low density. Neutrophils could not be purified by density gradient centrifugation using either g...
Characterisation of Chlamydia psittaci isolated from a horse. This paper describes the isolation and characterisation of a strain of Chlamydia psittaci obtained from a nasal swab taken from a horse with serous nasal discharge. Initial isolation was achieved in cycloheximide-treated McCoy cell monolayers. Chlamydial inclusions stained by immunofluorescence either with a rabbit antiserum raised against C. psittaci or with a monoclonal antibody directed against the genus-specific lipopolysaccharide antigen were single and compact. They did not stain with iodine or with a monoclonal antibody reactive against Chlamydia trachomatis. The agent was re-isolated i...
Cryopreservation of equine mononuclear cells for immunological studies. A rapid and simple technique for the cryopreservation and recovery of equine mononuclear cells was developed. Buffy-coat leukocytes were frozen in autologous plasma containing 10% DMSO and mononuclear cells were recovered by gradient sedimentation using a standard Ficoll-Hypaque purification procedure. The total numbers of mononuclear cells recovered from cryopreserved samples were 94%-82% of those recovered from fresh blood samples. The functional capabilities of the mononuclear cells from cryopreserved buffy coat preparations were compared with those of mononuclear cells from fresh samples b...
Polysulfated glycosaminoglycan accelerates net synthesis of collagen and glycosaminoglycans by arthritic equine cartilage tissues and chondrocytes. Low molecular weight polysulfated glycosaminoglycan (PSGAG) stimulated net collagen and glycosaminoglycan synthesis by normal and arthritic equine fetlock cartilage tissues in organ culture. Arthritic tissues were more sensitive to PSGAG stimulation. The rates of cartilage-specific type-II collagen and chondroitin sulfate-rich glycosaminoglycan synthesis by confluent chondrocyte cell cultures obtained from normal and arthritic equine cartilage tissues were increased by 25 and 50 mg of PSGAG/ml. Cells from arthritic cartilage were also more sensitive to the presence of PSGAG. In addition, conce...
Endotoxin-induced tumor necrosis factor activity production by equine peritoneal macrophages. A study was performed to determine whether equine macrophages produce tumor necrosis factor (TNF) activity in vitro in response to endotoxin and to study the effects of endotoxin concentration and incubation time on the amount of TNF produced. Equine peritoneal macrophages were isolated and cultured in vitro for 2, 6, 12, or 24 hr in tissue culture media containing 1) no additive (nonstimulated control), 2) endotoxin (0.5 ng/ml, 5 ng/ml, or 5 micrograms/ml), or 3) the calcium ionophore A23187 (0.95 microM). The supernatant media concentrations of TNF activity were determined by an in vitro cyt...
In vitro isolation of a neutralization escape mutant of equine infectious anemia virus (EIAV). A neutralization escape mutant (A/1 E) of equine infectious anemia virus was isolated after 13 passages in cell culture in the presence of serum containing antibodies to type- and group-specific determinants of EIAV envelope glycoproteins. Loss of neutralization by the selecting serum correlated with loss of two epitopes in the major envelope glycoprotein gp90 of A/1 E which were present in a parallel variant isolated from a persistently infected pony.
Secretion of prostaglandins and progesterone by cells from corpora lutea of mares. Corpora lutea (CL) were collected from mares during early (Day 4-5), mid- (Day 8-9), and late (Day 12-13) dioestrus. Dispersed cell suspensions were obtained by enzymic digestion of tissue. Two distinct luteal cell populations (large and small) were observed. The proportion of small luteal cells significantly increased as age of CL advanced. Cells (2 x 10(6)) from CL which were incubated for 24 h secreted prostaglandin (PG) F, PGE-2 and 6-keto-PGF-1 alpha (the stable metabolite of prostacyclin). Higher concentrations of all PGs were produced by cells from CL at early dioestrus than from those ...
Influence of technical parameters on the in vitro motility of equine neutrophils in the presence of streptococcal culture supernatant. To identify the influence of technical factors on the in vitro motility of equine neutrophils towards streptococcus culture supernatant in an under-agarose assay, we studied the changes in eight cell migration parameters. The distances the phagocytes travelled by directed, random and spontaneous migration increased with incubation time, cell concentration and the gelatin and serum contents of the migration plates. The contribution of chemotaxis to the phagocyte migrations, however, decreased simultaneously. The directed and random, though not the spontaneous, migrations of the phagocytes incre...
Isolation and characterization of equine microvascular endothelial cells in vitro. The use of cultured tissue has not yet become widespread in research involving equine disease, and this may be attributable in part to the scarcity of published reports concerning tissue culture methods for this species. We report here the isolation of equine microvascular endothelium (EMVE) from fresh omental tissue of horses and ponies. Fresh donor tissue was minced, subjected to collagenase digestion, and filtered. Cells were layered on 5% bovine serum albumin for gravity sedimentation, the bottom layer was collected, and the cells were plated onto fibronectin-coated flasks. Medium consiste...
Culture of equine trophoblastic vesicles in vitro. Trophoblastic vesicles have been used to study early embryonic development and maternal recognition of pregnancy in domestic animals. The purpose of this study was to characterize the formation of trophoblastic vesicles from Day-12 to Day-16 equine conceptuses. Conceptuses (n = 19) were collected nonsurgically from mares, the capsule was removed, and the conceptus (trophoblast and inner cell mass) was dissected into 2- to 4-mm fragments. Conceptus fragments were cultured in either Ham's F10 (HF10) or Minimum Essential Media (MEM) with 10% fetal bovine serum (FBS) in 24-well plates. Plates were...
An ongoing in vivo immune response affects the abundancy and differentiation of lymphokine-activated killer cell precursors, but does not influence their broad spectrum target reactivity. Using a model of local lymph node (LN) immunization, we investigated the effect of in vivo Ir on the generation of lymphokine-activated killer (LAK) cells or their precursors. Ag used for immunization were SRBC, horse RBC, OVA, keyhole limpet hemocyanin, or CFA. Ag-draining LN, in the acute phase of the Ir, did not contain detectable LAK effector activity, nor an enhanced NK activity. After culture for 3 to 5 days in the absence of exogenously added IL-2, immunized LN cells developed a spontaneous LAK-like cytotoxicity. This activity represented a substantial fraction of the IL-2-generated LAK...
Equine peritoneal macrophage production of thromboxane and prostacyclin in response to platelet activating factor and its receptor antagonist SRI 63-441. The formation of eicosanoids may be a primary route through which platelet activating factor (PAF) exerts its effects during endotoxemia. Since endotoxemia is a common cause of death in horses, a study was conducted to determine whether PAF could stimulate equine macrophage release of thromboxane A2 (TxA2) and prostacyclin (PGI2) and whether a PAF-receptor antagonist would alter macrophage eicosanoid synthesis. Equine peritoneal macrophages were cultured from clinically normal horses and exposed to various concentrations of PAF, the PAF-receptor antagonist SRI 63-441, endotoxin, or a combinati...
Change in host cell tropism associated with in vitro replication of equine infectious anemia virus. Similar to other human and animal lentiviruses, equine infectious anemia virus (EIAV) is detectable in vivo in cells of the monocyte-macrophage lineage. Owing to their short-lived nature, horse peripheral blood macrophage cultures (HMC) are rarely used for in vitro propagation of EIAV, and equine dermal (ED) or kidney cell cultures, which can be repeatedly passed in vitro, are used in most studies. However, wild-type isolates of EIAV will not grow in these cell types without extensive adaptation, a process which may attenuate viral virulence. To better define the effect of host cell tropism on...
Membrane viscosity of lymphocytes and influence of phytohemagglutinin. The membrane viscosity of peripheral blood lymphocytes (PBLs) of equine, bovine and canine was measured by the use of time-resolved fluorescence depolarization technique with 1, 6-diphenyl-1,3,5-hexatriene (DPH). The viscosity values were 0.55, 0.59 and 0.50 poise for equine, bovine and canine PBLs, respectively. These values were compared with steady-state anisotropies and order parameters measured from electron spin resonance (ESR) of 5-doxyl stearic acid. Both values were increased with increase of viscosity. The fluid property of the membranes stimulated with phytohemagglutinin-P (PHA) was...
Vitamins and other metabolites in various sera commonly used for cell culturing. Many cell culture media use different sera to enhance growth. We assayed vitamins and some related metabolites in different sera and identified the concentration of: thiamin, biotin, folates, riboflavin, pantothenates, nicotinates, vitamins B6, B12, A, E, C, and carotenes and some related metabolites: biopterins, free inositol, free and total choline, total carnitines in chicken, horse, rabbit, goat, pig, calf, newborn calf, fetal calf and human sera. Results indicate that vitamin and metabolite content of different sera vary. Such variations could produce fluctuant effects on cell culturings ...
Use of a monoclonal antibody to evaluate integrity of the plasma membrane of stallion sperm. Transmission electron microscopy was used to confirm that a monoclonal antibody (F79.3E2; class IgG1 kappa) was specifically localized to an antigen in the acrosomal ground substance of stallion sperm. This antibody was used to develop and validate an indirect immunofluorescent procedure to evaluate integrity of the plasma-acrosomal membranes of stallion sperm. The concept was that primary monoclonal antibody would be "shielded" from its acrosomal antigen by an intact plasma membrane. Conversely, sperm with damaged plasma-acrosomal membranes would exhibit green acrosomal fluorescence when view...
Detection of eastern equine encephalomyelitis virus antigen in equine brain tissue by enzyme-linked immunosorbent assay. Sensitivity and specificity of an antigen-capture ELISA vs virus isolation in cell culture were evaluated for the detection of eastern equine encephalomyelitis (EEE) virus in the brain tissue of naturally infected equids. Brain specimens from 16 equids with neurologic disease were examined by ELISA and by inoculation onto baby hamster kidney cell cultures. Of 10 brain samples from which virus was isolated in the cell culture bioassay, all were correctly identified as containing EEE virus antigen by ELISA. None of the remaining 6 specimens, without detectable infectious EEE virus, contained det...
Chemotactic and chemokinetic activity of Streptococcus faecalis culture supernatant for equine neutrophils. Although equine neutrophils did not respond towards formylated methionyl peptides, Streptococcus faecalis culture supernatant caused an in vitro stimulation of equine neutrophil motility when measured by an under-agarose assay. The migration of neutrophils towards the culture supernatant increased sigmoidally with the logarithmic concentration of the culture supernatant in the chemoattractant wells. The streptococcal culture supernatant was chemokinetic because it stimulated the random motility of the phagocytes. Because granulocytes migrated further towards the supernatant than could be expla...
Establishment of equine T-lymphocyte cultures dependent on recombinant human interleukin-2. Long-term equine lymphocyte cultures were initiated and maintained in continuous culture with medium containing recombinant human interleukin-2. Cultures were successfully maintained with lectin activation signals and recombinant human interleukin-2 or with recombinant human interleukin-2 alone. All cell cultures that were characterized had a T-lymphocyte phenotype and had lectin-dependent or -independent cytotoxicity directed to various cell types. These findings demonstrate that long-term equine T-lymphocytes cultures can be initiated and maintained easily.
Propagation and quantitation of animal herpesviruses in eight cell culture systems. A comparative study was carried out to determine the relative sensitivities of eight different cell culture systems to six different herpesviruses of animals. The cells used were: OFL (ovine fetal lung), ML (mink lung), FK (ferret kidney), PTK-2 (potoroo kidney), TEK (turkey embryo kidney), ED (equine dermal), BT (bovine turbinate), and PK15 (porcine kidney). The viruses tested were: PRV (pseudorabies) of swine, CPHV (caprine herpesvirus), IBRV (infectious bovine rhinotracheitis virus), DN-599 strain of bovine herpesvirus type 4, EHV-1 (equine herpesvirus), and CHV (canine herpesvirus). On the...
A monoclonal antibody identifying a T-cell marker in the horse. A cell surface molecule of equine T lymphocytes was identified and characterized using a mouse monoclonal antibody, HT23A. The molecule was detected on all T cells but not on other cells in peripheral blood, with the possible exception of a small subpopulation (about 5%) of B cells, as assessed by indirect immunofluorescence and flow cytometry. HT23A labelled T cell areas of horse lymph nodes and spleen when used in an indirect immunoperoxidase assay on frozen sections. Macrophages and neutrophils were not labelled by the antibody nor were frozen sections of horse liver, kidney, or brain. HT23...
Difference in growth behavior of human, swine, equine, and avian influenza viruses at a high temperature. Growth characteristics of a wide range of influenza A viruses from different mammals and bird species were examined in an established line of canine kidney (MDCK) cells at an ordinary (37 degrees C) and a high temperature (42 degrees C). Although all viruses employed in the present study possessed a capability of replicating at 37 degrees C, virus growth at 42 degrees C showed considerable variation and reflected differences in the natural hosts of the isolates. All reference strains and isolates from bird species grew well in the MDCK cells maintained at 42 degrees C, but human viruses did no...
Serum neutralizing antibody titers in dairy cattle administered an inactivated vesicular stomatitis virus vaccine. Two doses of a formalin-killed, cell culture-derived vesicular stomatitis virus (vsv)-New Jersey serotype vaccine were administered intramuscularly, 30 days apart, to all lactating and nonlactating cows in a 350-cow dairy herd. Serum specimens were obtained serially from 96 cows before vaccination and at 30, 52 and 80 days after vaccination and from 24 of these cows 175 days after vaccination. Serum neutralizing antibody titers to vsv-New Jersey serotype were determined from serum-dilution, plaque-reduction tests. Serum neutralizing antibody titers also were determined during the same period f...
Preservation of skin by refrigeration for autogenous grafting in the horse. Eighteen stored split thickness meshed skin grafts were applied to surgically created lesions on the metacarpal and metatarsal regions of six horses. Donor skin was harvested from the sternal region, meshed and stored at 4 degrees C in a cell culture medium containing 10% serum. Stored grafts were applied to the wounds at 1, 2, and 3 week intervals. Acceptance of the grafts stored for 1 week was generally poor (1 of 6 grafts), whereas that of the 2 and 3 week old grafts was generally excellent (10 of 12 grafts). Poor acceptance of the 1 week old grafts was attributed to streptococcal infection...
Ultrastructure of equine endothelial cells exposed to endotoxin and flunixin meglumine and equine neutrophils. An in vitro system of cultured equine endothelial cells was evaluated as a model for endotoxin (ET) exposure in the horse. Primary cell lines from pulmonary vessels and aortas were cultured from tissues of 6 horses. Effects of ET alone with and without serum and in combination with the cyclo-oxygenase inhibitor flunixin meglumine and isolated equine neutrophils were evaluated by transmission electron microscopy. Cells plus serum were incubated with 10, 25, 50, or 100 micrograms of ET/ml of incubation medium for 1, 3, 8, or 24 hours. Cells without serum were cultured for 1 and 3 hours. Flunixin...
