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.
Darlington RW, James C.Darlington, R. W. (St. Jude Children's Research Hospital, Memphis, Tenn.), and C. James. Biological and morphological aspects of the growth of equine abortion virus. J. Bacteriol. 92:250-257. 1966.-The growth of equine abortion virus (EAV) was studied by bioassay and electron microscopy in L-cell monolayer and suspension cultures, and in HeLa and BHK 21/13 cell monolayers. Results of virus assay (plaque-forming units) indicated that production of cell-associated virus (CAV) began at 6 to 9 hr after infection in all of the cell strains used. Virus release occurred 1 to 2 hr later. By 15 to 20 h...
Channing CP.OUR knowledge of the pathways of steroid biosynthesis in the ovary has been gained mainly by incubations of ovaries in vitro1,2. The tissues incubated have contained numerous cell types: granulosa cells, theca interna cells, stromal cells, interstitial cells, and sometimes luteal cells. Possibly such mixtures of two or more different cell types are able to secrete hormones that one cell type cannot secrete by itself3–9. Furthermore, during such incubations in vitro an exchange of precursors and products between different cell types may be facilitated because of breakdown of naturally occurri...
RANDALL CC, WALKER BM.Randall, Charles C. (University of Mississippi School of Medicine, Jackson) and Barbara M. Walker. Degradation of deoxyribonucleic acid and alteration of nucleic acid metabolism in suspension cultures of L-M cells infected with equine abortion virus. J. Bacteriol. 86:138-146. 1963.-Metabolic alterations in log-phase suspension cultures infected with equine abortion virus (EAV) were determined in L-M cells simultaneously labeled or prelabeled with H(3)- or C(14)-thymidine. Although infection produced an early stimulation of the uptake of labeled thymidine (TdR) into the acid-soluble fraction of...
RANDALL CC, LAWSON LA.The research article discusses the successful adaptation of the Equine Abortion Virus (EAV) to L-M 929 cells, the impact on infected cultures, and possible reasons for earlier unsuccessful attempts. It […]
Science (New York, N.Y.)August 25, 1961
Volume 134, Issue 3478 565-566 doi: 10.1126/science.134.3478.565
LOWENTHAL JP, BERMAN S, GROGAN EW.Protection tests in guinea pigs indicate that vaccines prepared from virus propagated in chick embryo cell cultures are as effective as the purified whole chick embryo vaccines which are currently used for human immunization against eastern equine encephalomyelitis.
RANDALL CC, ORR MF, SCHELL FG.Intracellular organisms resembling Histoplasma capsulatum have been observed in apparently normal amnio-allantoic membrane and adult horse spleen maintained in tissue culture.
The significance of this finding and comparison with another fungus Cryptococcus jarciminosus is discussed.
The objective of this study was to graft autologous mesenchymal stem cells (MSCs) at the site of surgical repair of a soft palate defect in an adult horse in an attempt to improve wound healing and to investigate whether the transplanted MSCs would integrate into the soft palate structure and participate in regeneration. Bone marrow was collected from an adult horse with a full-thickness soft palate defect. The MSCs were isolated, cultured in monolayers, and labeled with 5-bromo-2-desoxymidine (BrdU) and chloromethylbenzamido-DiI-derived (cm-DiI) before transplantation. The soft palate defect ...
Urayama S, Muko R, Muranaka M, Mita H, Ohta M, Matsuda H, Tanaka A.Systemic inflammatory response syndrome (SIRS) induced by endotoxemia is usually secondary to colitis and is a major cause of high morbidity and mortality in horses. Non-steroidal anti-inflammatory drugs (NSAIDs), such as flunixin meglumine (FM) and meloxicam (MX), are used to improve clinical outcomes in SIRS/endotoxemia. These NSAIDs suppress tumor necrosis factor-alpha (TNF-α) levels; however, the underlying mechanisms remain unclear. The aim of this study was to investigate the inhibitory effects of FM and MX on TNF-α in lipopolysaccharide (LPS)-stimulated peripheral blood mononuclear ce...
Broothaers K, Angel-Velez D, Molto FLG, Hedia M, Coster T, Govaere J, Soom AV, Menten B, Smits K.In 2022, a repeatable protocol for in vitro fertilization (IVF) using fresh semen was established in horses. This facilitated successful capacitation of equine semen allowing to explore novel applications. Objective: We aimed to extend this technique to IVF with frozen-thawed semen and intracytoplasmic sperm injection (ICSI), and determine the outcome parameters such as blastocyst production and euploidy rates. Methods: A total of 221 oocytes were subjected to either IVF with frozen-thawed semen, ICSI with frozen-thawed semen incubated under capacitating conditions (ICSI cap) or control ICSI w...
Salinas-Varas C, Espinosa G, Muñoz-Caro T, Conejeros I, Gärtner U, Fey K, Arnhold S, Taubert A, Hermosilla C.Neutrophil extracellular trap (NET) are thin and long web-like structures composed of DNA and antimicrobial proteins released by activated polymorphonuclear neutrophils (PMN) as part of the innate immune response. Adipose-derived stem cells (ADSCs) represent an accessible, abundant and minimal invasive source of mesenchymal stem cells (MSCs), with high regenerative potential, immunomodulatory and anti-inflammatory properties. Although recognized immunomodulatory properties of ADSCs, their interaction with PMN and their role on NET formation remains poorly characterized. The present study aimed...
Broothaers K, Jouneau A, Angel-Velez D, De Coster T, Festuccia N, Archilla C, Calderari S, Jouneau L, Van den Branden E, Peere S, Polfliet E....In vitro production of equine embryos has been performed using intracytoplasmic sperm injection (ICSI) for the last two decades. Since 2022, a repeatable protocol for conventional in vitro fertilization (IVF) provides a successful alternative. However, little is known about the influence of the fertilization method on embryo quality and the transcriptomic profile. In this study, we aimed to examine differentially expressed genes (DEGs) between ICSI and IVF embryos in the horse. Therefore, ten equine sibling blastocysts, produced in vitro by either ICSI or IVF from three different mares, were s...
Sabzpoosh M, Hoveizi E, Gooraninejad S.This study aimed to extract and isolate endometrial stromal cells from Arabian mares and investigate their growth and differentiation potential. Endometrial biopsies were obtained from three healthy 6-year-old Arabian mares using a standardized, minimally invasive protocol. The isolated cells were characterized using flow cytometry and differentiation analysis. Flow cytometry revealed mesenchymal markers CD90 (95.2%) and CD105 (97.4%) and hematopoietic markers CD34 (1.17%) and CD45 (0.339%). The cells exhibited differentiation potential into adipocytes, osteoblasts, and chondrocytes. The findi...
Song D, Wang G, Baterin T, Weng Y, Dugarjaviin M, Li B.Normal spermatogenesis in Mongolian horses depends on the mitotic division of spermatogonia, two successive meiotic divisions, and the morphological transformation of spermatids into mature spermatozoa. The gene is involved in the meiosis cycle and is required for normal chromosome association during meiosis. Previous studies have shown that alternative splicing of may promote spermatogenesis in Mongolian horses. In this paper, the regulatory effects of different alternative splicing events on Mongolian horse spermatogenesis are investigated. Results: In this study, two overexpressed lentiv...
Cooper BL, Hobbs KJ, Bayless R, Stinson-Miller A, Gruber E, Hepworth-Warren K, Lavoie JP, Sheats MK.Severe equine asthma (sEA) is characterized by increased lower airway neutrophils that contribute to dysregulated inflammation through the release of cytokines, reactive oxygen species and neutrophil extracellular traps (NETs). NETs are composed of cell-free DNA (cfDNA) intercalated with enzymatic proteins and are known to be increased in the lower airway of asthmatic horses. The objectives of this study were two-fold: 1. Determine if cfDNA can be accurately measured in equine bronchoalveolar lavage fluid (BALF) and tracheal wash (TW) with a Qubit 4 fluorometer. 2. Determine whether Qubit-meas...
Graide H, Duysens J, Frank T, Mouithys-Mickalad A, Niesten A, Sandersen C, Ceusters J, Serteyn D.Musculoskeletal pathologies present challenges in athletic horses, often leading to functional impairment. The slow or limited regenerative capacity of bone, joint, and tendon/ligament injuries, coupled with the limitations of conventional treatments, highlights the need for innovative therapies such as ortho-biologics and mesenchymal stem/stroma cells. Traditional 2D cell culture systems with fetal bovine serum (FBS) fail to replicate the complexity of the in vivo environment, whereas 3D cultures more accurately mimic native tissue architecture and cell-cell interactions. This study describes...
Meeremans M, Devriendt B, Bairiot S, Van Poucke M, Peelman L, Demeyere K, Meyer E, Van Vlierberghe S, De Schauwer C.Macrophages play key roles in tissue homeostasis and regeneration-associated inflammation. Unlike humans, a reliable protocol to obtain and polarise equine monocyte-derived macrophages is lacking. In this study the polarisation of equine macrophages, derived from CD172a peripheral blood monocytes is described. After differentiation, IFN-γ/LPS or IL-4 were used to induce pro- and anti-inflammatory phenotypes, respectively. Evaluation criteria included morphology, mRNA (RT-qPCR) and protein expression (flow cytometry, immunofluorescence), nitric oxide and arginase production, cytokine secretion...
Ramos Serrano B, Ávalos Rodríguez A, Edward Kjelland M, Ernesto Hernández Pichardo J.Semen is a complex fluid that, in addition to spermatozoa, contains other cell populations, including immune cells, immature male germ cells, epithelial cells, and fibroblasts. These cells share the diploid condition, making them suitable candidates as nuclear donors for somatic cell nuclear transfer (SCNT) cloning. The generation of viable embryos and offspring has been demonstrated using these cells. Effective methods for isolating them from semen include centrifugation and osmotic gradient techniques; however, prolonged in vitro culture periods are necessary to establish primary cultures fr...
Rodrigues MF, Neves AP, Ferreira R, Pessoa GA, Araujo DN, de Souza GV.Melatonin has demonstrated beneficial effects in regulating inflammatory processes in various tissues, but its role in mare uterine physiology remains unclear. Objective: To investigate the effects of melatonin on the gene expression of cytokines (IL-1β, IL-6, IL-8, IL-10, and TNF-α) in equine uterine explants cultured for 6, 12, or 24 hours. Methods: The study was divided into two experiments. Part 1 aimed to evaluate the cytotoxicity of melatonin at concentrations of 0, 0.5, 1, 2, 4, and 8 mM on endometrial cells. Part 2 aimed to determine the expression of inflammatory cytokines in uterin...
Li Z, Yu T, Ge L, Lv S, Fu Q, Shi H.Equine herpesvirus type 1 (EHV-1) is a major veterinary pathogen causing significant economic losses in the livestock industry. Despite its impact, effective vaccines and targeted antiviral strategies remain limited, largely due to an incomplete understanding of host factors regulating viral replication and pathogenesis. Unassigned: To systematically identify host genes essential for EHV-1 infection, we established a BHK-21 cell line stably expressing Cas9 and performed a genome-wide CRISPR/Cas9 knockout screen using a pooled lentiviral single-guide RNA library. Significantly enriched candidat...
Zanacchi A, Fruscione F, Grasselli S, Razzuoli E, Pastorino L, Di Lisa D.Developing advanced preclinical in vitro cancer models is essential for understanding tumor biology, improving drug discovery, and enhancing the clinical translation of new therapies. Papillomaviruses (PVs) cause diverse lesions in humans and animals; notably, bovine papillomavirus (BPV) is the main cause of equine sarcoid, the most common skin tumor in horses. Its pathogenesis involves viral infection, fibroblast transformation, and extracellular matrix (ECM) remodeling. These processes highlight the limitations of conventional two-dimensional (2D) culture systems, which fail to replicate the...
Thieulent CJ, Sarkar S, Carossino M, Bhowmik M, Zhu H, Balasuriya UBR.Our laboratory identified the susceptible allelic variant of equine CXCL16 protein (EqCXCL16S) as an entry receptor for equine arteritis virus (EAV). However, EAV has a broad host cell tropism and infects cells that lack EqCXCL16S. Thus, we hypothesized that EAV interacts with other host cell protein(s) that facilitate EAV infection. A virus overlay protein-binding assay in combination with a Far-Western blot from EAV-susceptible equine pulmonary artery endothelial cells (EECs) and equine dermal fibroblasts (E. Derm) identified a 57 kDa protein, present in the membrane fraction of the protein ...
Beniwal N, Lal B, Mithina S, Verma CK, Kumar S, Phagna V, Jakhar K, Sonar S, Gupta V, Singh R, Kumar N, Tan CW, Thachamvally R, Singha H, Murzello K....Rabies is a neglected tropical zoonotic disease caused by rabies-virus (RV) infection and is responsible for almost 60,000 annual deaths globally, largely affecting the socio-economically disadvantaged population. Although fatality is preventable by immunization either before or after exposure with therapeutic antibodies, the high cost of prophylaxis or treatment limits their accessibility for the affected population. However, due to the almost 100% fatality rate in symptomatic individuals, almost 29 million annual vaccinations are performed, imposing high financial burden. Human transmission ...
Sadowska A, Molcan T, Słyszewska M, Skarzynski DJ, Ferreira-Dias G, Szóstek-Mioduchowska A.Equine endometrosis is a chronic degenerative condition of the endometrium. A hallmark feature of endometrosis is endometrial fibrosis accompanied by degenerative changes in the adjacent tissue structure. Tumor necrosis factor α (TNFα), a pleiotropic cytokine produced by various immune cells, plays a dual role in coordinating immune responses and regulating cell/tissue homeostasis, making it an important regulator of fibrotic-related disorders. However, the exact role of TNFα in the development of equine endometrial fibrosis remains to be discovered and explained. Therefore, the main aim of...
Parker JL, Berghaus LJ, Gordon J, Mathews L, Akinde MA, Hall DB, Peroni JF.Mesenchymal stromal cells (MSCs) support wound healing through immunomodulation and by promoting tissue organization. Cell tracking studies demonstrate that MSCs must 'home' to injuries in order to participate in reparative processes. It has been reported that in vivo cell tracking labels, including superparamagnetic iron oxide particles (SPIOs), can alter MSC homing. Homing is thought to occur in coordinated steps including tethering, activation, arrest, diapedesis, and migration, guided by chemotactic stimuli. Equine distal limb injuries, which are commonly treated with MSCs, possess a disti...
Beaumont RE, Flood C, Guest DJ.Interleukin 1 beta (IL-1β) is upregulated following a tendon injury and in vitro studies have shown that it leads to numerous negative effects on tendon cell function and gene expression. IL-1β activates nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and we hypothesised that inhibiting NF-κB activation would mediate the negative effects of IL-1β on equine tendon cells in 3-dimensional (3D) cultures. Results: Here, we tested three inhibitors of NF-κB signalling (Bortezomib, BAY11-7082 and Wedelolactone) along withTJ-M2010-5, an inhibitor of MyD88, which is a critic...
Wójtowicz A, Sadowska A, Molcan T, Słyszewska M, Drzewiecka EM, Tobolski D, Ferreira-Dias G, Szóstek-Mioduchowska A.Mare endometrosis is a chronic degenerative condition of the endometrium, primarily characterized by fibrosis, involving interactions among fibroblasts, immune cells, and epithelial cells regulated by cytokines and growth factors. T helper (Th)1 and Th2 cells seem to play a pivotal role in fibrosis. However, their roles in equine endometrial fibrosis remain unknown. This study explores Th1 and Th2 cell distribution across different stages of endometrium histopathological Kenney and Doig categories; and evaluated their secretome effects on non-fibrotic endometrium derived fibroblast functional ...
Larson MK, Gaffney C, Hoagland C, Jayawickrama J, Kamm JL.To determine the xenogen-free serum source that provides the greatest number of live equine mesenchymal stromal cells (MSCs) while maintaining the MSC phenotype. Unassigned: Equine bone marrow-derived MSCs from 8 horses were cultured for 7 days in media containing one of the following serum treatments: 10% xenogeneic serum, 10% or 20% commercial allogeneic equine serum, 10% autologous serum, 10% equine pooled platelet lysate (PPL), or a staged media reduction of xenogeneic media. Live cell numbers, MSC viability, and MSC immunophenotype were compared. Unassigned: The use of 10% commercial allo...
Muñoz-García CC, Soriano-Campos MP, Luis-Calero M, Gallardo-Soler A, González-Fernández L, Macías-García B.The use of mesenchymal stromal cells (MSCs) in equine reproduction is increasing its interest in the treatment of specific pathologies. MSCs have been isolated from follicular aspirates obtained during transvaginal oocyte aspiration in women, offering a novel source for autologous therapies in reproductive treatments. However, this approach has not been tested in mares despite the common use of transvaginal oocyte aspiration for oocyte collection to produce equine embryos in vitro. Our study aimed to investigate the feasibility of isolating MSCs from equine ovarian follicular aspirates obtaine...
Del Prete C, Gaspari G, Kosior MA, Merlo B, Iacono E, Longobardi C, Martino NA, Dell'Aquila ME, Damiano S, Cocchia N, Gasparrini B, Lange-Consiglio A.Over the years, regenerative therapies have emerged as promising alternatives for persistent breeding-induced endometritis. In vitro studies testing the effects of these therapies on equine endometrial cells are still scarce. This study aimed to evaluate in vitro the effect of Wharton's jelly (WJ) mesenchymal stromal/stem cell (MSCs)-derived conditioned medium (WJ-CM) and platelet-rich plasma (PRP) on equine endometrial cells, with or without lipopolysaccharide (LPS)-induced inflammation. The WJ-CM was obtained after 24 h of starvation in Ringer's lactate of WJ-MSCs and PRP was prepared using...
de la Fuente A, Scoggin C, Bradecamp E, Martin-Pelaez S, van Heule M, Troedsson M, Daels P, Meyers S, Dini P.Maturation is a critical step in the development of an oocyte, and it is during this time that the oocyte advances to metaphase II (MII) of the meiotic cycle and acquires developmental competence to be fertilized and become an embryo. However, in vitro maturation (IVM) remains one of the limiting steps in the in vitro production of embryos (IVP), with a variable percentage of oocytes reaching the MII stage and unpredictable levels of developmental competence. Understanding the dynamics of oocyte maturation is essential for the optimization of IVM culture conditions and subsequent IVP outcomes....
DI Stefano DA, Coccoluto L, Panina-Bordignon P, Brambilla E, Ruffini F, Murtaj V, Orlando F, Colombo M, Frigerio C, DI Bona A, Recupero D, Morroni M....Bone grafting is a surgical procedure that involves the use of bone tissue or bone substitutes to repair damaged bone. In dentistry and maxillofacial surgery, bone graft substitutes from various sources are commonly used. Given their critical role in clinical outcomes, it is essential to thoroughly investigate the biological and mechanical properties of these materials. Methods: In this in vitro study, we evaluated the biological properties of two equine-derived bone graft substitutes in comparison to β-tricalcium phosphate. The materials included one equine-derived graft containing hydrolyze...
Schmitz M, Neugebauer E, Full F, Conn KL.Comparative transcriptomics offers a powerful approach to elucidate host-virus interactions across related pathogens, yet systematic evaluations across species-matched cellular systems remain limited. We performed a cross-species RNA sequencing analysis of respective species' cells infected with three alphaherpesviruses-herpes simplex virus 1 (HSV-1), bovine alphaherpesvirus 1 (BHV-1), and equid alphaherpesvirus 1 (EHV-1)-to dissect conserved and virus-specific transcriptional responses. We show that certain orthologous genes and orthologous pathways are differentially regulated upon infection...
Uprety T, Soni S, Sreenivasan C, Hause BM, Naveed A, Ni S, Graves AJ, Morrow JK, Meade N, Mellits KH, Adam E, Kennedy MA, Wang D, Li F.Equine rotavirus species A (ERVA) G3P[12] and G14P[12] are two dominant genotypes that cause foal diarrhoea with a significant economic impact on the global equine industry. ERVA can also serve as a source of novel (equine-like) rotavirus species A (RVA) reassortants with zoonotic potential as those identified previously in 2013-2019 when equine G3-like RVA was responsible for worldwide outbreaks of severe gastroenteritis and hospitalizations in children. One hurdle to ERVA research is that the standard cell culture system optimized for human rotavirus replication is not efficient for isolatin...
Hassanien RT, Thieulent CJ, Carossino M, Li G, Balasuriya UBR.(1) Background: equid alphaherpesvirus-1 (EHV-1) is a highly contagious viral pathogen prevalent in most horse populations worldwide. Genome-editing technologies such as CRISPR/Cas9 have become powerful tools for precise RNA-guided genome modifications; (2) Methods: we designed single guide RNAs (sgRNA) to target three essential (ORF30, ORF31, and ORF7) and one non-essential (ORF74) EHV-1 genes and determine their effect on viral replication dynamics ; (3) Results: we demonstrated that sgRNAs targeting essential lytic genes reduced EHV-1 replication, whereas those targeting ORF74 had a negligi...
Dahlgren AR, Careddu F, Norris JW, Adili R, Kropp EM, Li Q, Holinstat M, Biunno I, Balduini A, Tablin F, Shavit JA, Finno CJ.SEL1L is a well-known protein in the endoplasmic reticulum associated degradation (ERAD) pathway. While it is known to be expressed in platelets, SEL1L has never been shown to play an active role. Here we find evidence that SEL1L regulates platelet function. We first identified SEL1L through the study of Atypical Equine Thrombasthenia (AET), an autosomal recessive platelet disorder found in Thoroughbred horses. A missense variant in (c.1810A>G p.Ile604Val) was found in AET-affected horses, which we show is associated with decreased protein expression. SEL1L is intracellular in equine platelet...
Verstraete MH, Kuijpers F, Holl W, Norris J, van Heule M, Martin-Pelaez S, Heard C, Uribe-Salazar J, Armien A, Conley A, De Spiegelaere W....The endometrium is a hormonally responsive tissue that undergoes cyclic remodeling. Although endometrial organoids have been established in several species, detailed characterization remains limited. Here, we assessed the structural and molecular fidelity of equine endometrial organoids across reproductive cycle stages and during extended culture. Organoids were generated from biopsies collected during estrus and diestrus and analyzed using histology, immunohistochemistry, electron microscopy, and bulk- and single-cell RNA sequencing. Organoids formed polarized cystic structures composed of co...
