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Topic:Developmental Biology

Developmental biology in horses involves the study of the processes by which horses grow and develop from a single fertilized egg into a fully formed organism. This field encompasses various stages, including embryonic development, fetal growth, and postnatal maturation. Researchers in this area examine cellular differentiation, gene expression, and morphogenetic movements that contribute to the formation of tissues and organs in equine species. Key topics include the molecular mechanisms that regulate developmental pathways, the influence of genetic and environmental factors on development, and the identification of developmental disorders. This page compiles peer-reviewed research studies and scholarly articles that explore the fundamental aspects of equine developmental biology, offering insights into the mechanisms driving normal and abnormal development in horses.
Effects of season on placental, foetal and neonatal development in horses.
Theriogenology    April 22, 2017   Volume 97 98-103 doi: 10.1016/j.theriogenology.2017.04.027
Beythien E, Aurich C, Wulf M, Aurich J.Seasonal changes in metabolic rate have been shown in horses and we hypothesized that this leads to the birth of smaller foals early in the year. Mares and their foals were assigned to three groups by day of foaling within the year (e.g. 1 January = day 1): Group 1 (n = 10) day 40-65, group 2 (n = 8) day 67-92, group 3 (n = 9) day 94-121. Groups did not differ with regard to parity. In foals, height at withers and body weight were determined on days 1-5 and weekly until 12 weeks of age. Chest circumference, distances fetlock to carpus, carpus to elbow, poll to nose and crown-rump lengt...
Placentation in the plains zebra (Equus quagga).
Reproduction, fertility, and development    March 31, 2017   Volume 29, Issue 11 2225-2234 doi: 10.1071/RD16475
Allen WRT, Stansfield F, Wilsher S.The placenta and fetal gonads of 12 pregnant plains zebra (Equus quagga), estimated to be between 81 and 239 days of gestation, were examined. The diffuse, microcotyledonary zebra placenta appeared, developmentally, to be 3-4 weeks behind its counterpart in horse pregnancy and this, together with the presence of small and long-lived endometrial cups, low levels of zebra chorionic gonadotrophin in maternal serum and few accessory corpora lutea in the maternal ovaries during the first half of gestation, made zebra pregnancy more similar to donkey than horse pregnancy. Zebra fetal gonads enlarged...
Dynamics of 5-methylcytosine and 5-hydroxymethylcytosine during pronuclear development in equine zygotes produced by ICSI.
Epigenetics & chromatin    March 15, 2017   Volume 10 13 doi: 10.1186/s13072-017-0120-x
Heras S, Smits K, De Schauwer C, Van Soom A.Global epigenetic reprogramming is considered to be essential during embryo development to establish totipotency. In the classic model first described in the mouse, the genome-wide DNA demethylation is asymmetric between the paternal and the maternal genome. The paternal genome undergoes ten-eleven translocation (TET)-mediated active DNA demethylation, which is completed before the end of the first cell cycle. Since TET enzymes oxidize 5-methylcytosine to 5-hydroxymethylcytosine, the latter is postulated to be an intermediate stage toward DNA demethylation. The maternal genome, on the other ha...
Evaluation of equine oocyte developmental competence using polarized light microscopy.
Reproduction (Cambridge, England)    March 10, 2017   Volume 153, Issue 6 775-784 doi: 10.1530/REP-17-0125
Bertero A, Ritrovato F, Evangelista F, Stabile V, Fortina R, Ricci A, Revelli A, Vincenti L, Nervo T.The purpose of this study was to observe -matured equine oocytes with an objective computerized technique that involves the use of a polarized light microscope (PLM) in addition to the subjective morphological evaluation obtained using a classic light microscope (LM). Equine cumulus-oocyte complexes (COCs,  = 922) were subjected to different maturation times (24, 36 or 45 h), however, only 36-h matured oocytes were analyzed using CLM. The 36-h matured oocytes that reached maturity were parthenogenetically activated to evaluate the quality and meiotic competence. Average maturation perce...
Therapeutic Effects of Horseback Riding Interventions: A Systematic Review and Meta-analysis.
American journal of physical medicine & rehabilitation    March 3, 2017   Volume 96, Issue 10 717-725 doi: 10.1097/PHM.0000000000000726
Stergiou A, Tzoufi M, Ntzani E, Varvarousis D, Beris A, Ploumis A.Equine-assisted therapies, such as therapeutic riding and hippotherapy, are believed to have positive physical and emotional effects in individuals with neuromotor, developmental, and physical disabilities. The purpose of this review was to determine whether therapeutic riding and hippotherapy improve balance, motor function, gait, muscle symmetry, pelvic movement, psychosocial parameters, and the patients' overall quality of life. Methods: In this study, a literature search was conducted on MEDLINE, CINAHL, MBASE, SportDiscus, Cochrane Database of Systematic Reviews, Cochrane Controlled Trial...
Translational Research for Occupational Therapy: Using SPRE in Hippotherapy for Children with Developmental Disabilities.
Occupational therapy international    February 19, 2017   Volume 2017 2305402 doi: 10.1155/2017/2305402
Weissman-Miller D, Miller RJ, Shotwell MP.Translational research is redefined in this paper using a combination of methods in statistics and data science to enhance the understanding of outcomes and practice in occupational therapy. These new methods are applied, using larger data and smaller single-subject data, to a study in hippotherapy for children with developmental disabilities (DD). The Centers for Disease Control and Prevention estimates DD affects nearly 10 million children, aged 2-19, where diagnoses may be comorbid. Hippotherapy is defined here as a treatment strategy in occupational therapy using equine movement to achieve...
Blastocyst development after intracytoplasmic sperm injection of equine oocytes vitrified at the germinal-vesicle stage.
Cryobiology    February 13, 2017   Volume 75 52-59 doi: 10.1016/j.cryobiol.2017.02.004
Canesin HS, Brom-de-Luna JG, Choi YH, Ortiz I, Diaw M, Hinrichs K.We evaluated the meiotic and developmental competence of GV-stage equine oocytes vitrified under different conditions. In a preliminary study, using dimethyl sulfoxide (D), ethylene glycol (EG) and sucrose (S) as cryoprotectants, the maturation rate was higher for cumulus-oocyte complexes (COCs) held overnight before vitrification (37%) than for those vitrified immediately (14%; P < 0.05). Thereafter, all COCs were held overnight before vitrification. In Experiment 1 we compared 1 min (1m) and 4 min (4m) exposure to vitrification and warming solutions; oocytes that subsequently matured wer...
Tissue organization alters gene expression in equine induced trophectoderm cells.
General and comparative endocrinology    February 2, 2017   Volume 247 174-182 doi: 10.1016/j.ygcen.2017.01.030
Reinholt BM, Bradley JS, Jacobs RD, Ealy AD, Johnson SE.Rapid morphological and gene expression changes occur during the early formation of a mammalian blastocyst. Critical to successful retention of the blastocyst and pregnancy is a functional trophectoderm (TE) that supplies the developing embryo with paracrine factors and hormones. The contribution of TE conformational changes to gene expression was examined in equine induced trophoblast (iTr) cells. Equine iTr cells were cultured as monolayers or in suspension to form spheres. The spheres are hollow and structurally reminiscent of native equine blastocysts. Total RNA was isolated from iTr monol...
An open trial of equine-assisted therapy for children exposed to problematic parental substance use.
Health & social care in the community    February 1, 2017   Volume 25, Issue 3 1247-1256 doi: 10.1111/hsc.12427
Tsantefski M, Briggs L, Griffiths J, Tidyman A.Children exposed to problematic parental substance use (PPSU) often face a number of deleterious developmental outcomes, yet these children are less likely to become known to child protection and welfare services. Although there is a growing evidence base for equine-assisted therapy (EAT) as an effective treatment modality for atypically developing children and adolescents, scant research has explored the benefit of EAT for children exposed to PPSU. The current study is the first to explore the benefit of EAT for children exposed to PPSU in Victoria, Australia. Five 12-week EAT programmes were...
Scleraxis Is Essential for Tendon Differentiation by Equine Embryonic Stem Cells and in Equine Fetal Tenocytes.
Stem cells and development    January 24, 2017   Volume 26, Issue 6 441-450 doi: 10.1089/scd.2016.0279
Bavin EP, Atkinson F, Barsby T, Guest DJ.The transcription factor scleraxis is required for tendon development and is upregulated during embryonic stem cell (ESC) differentiation into tenocytes. However, its role beyond early embryonic development is not defined. We utilized a short hairpin RNA to knock down scleraxis expression in ESCs and adult and fetal tenocytes. No effect on growth or morphology was observed in two-dimensional cultures. However, scleraxis knockdown in fetal tenocytes significantly reduced COL1A1, COMP, and SOX9 gene expression. Scleraxis knockdown in adult tenocytes had no effect on the expression of these genes...
Deletion of 2.7 kb near HOXD3 in an Arabian horse with occipitoatlantoaxial malformation.
Animal genetics    January 23, 2017   Volume 48, Issue 3 287-294 doi: 10.1111/age.12531
Bordbari MH, Penedo MCT, Aleman M, Valberg SJ, Mickelson J, Finno CJ.In the horse, the term occipitoatlantoaxial malformation (OAAM) is used to describe a developmental defect in which the first cervical vertebra (atlas) resembles the base of the skull (occiput) and the second cervical vertebra (axis) resembles the atlas. Affected individuals demonstrate an abnormal posture and varying degrees of ataxia. The homeobox (HOX) gene cluster is involved in the development of both the axial and appendicular skeleton. Hoxd3-null mice demonstrate a strikingly similar phenotype to Arabian foals with OAAM. Whole-genome sequencing was performed in an OAAM-affected horse (O...
Developmental expression of B cell molecules in equine lymphoid tissues.
Veterinary immunology and immunopathology    December 13, 2016   Volume 183 60-71 doi: 10.1016/j.vetimm.2016.12.004
Prieto JMB, Tallmadge RL, Felippe MJB.Identification and classification of B cell subpopulations has been shown to be challenging and inconsistent among different species. Our study tested aspects of ontogeny, phenotype, tissue distribution, and function of equine CD5 B cells, which represented a greater proportion of B cells early in development and in the peritoneal cavity. CD5 and CD5 B cells differentially expressed B cell markers (CD2, CD21, IgM) measured using flow cytometry, but similar mRNA expression of signature genes (DGKA, FGL2, PAX5, IGHM, IL10) measured using quantitative RT-PCR. Sequencing lambda light chain segment...
Preattachment Embryos of Domestic Animals: Insights into Development and Paracrine Secretions.
Annual review of animal biosciences    December 5, 2016   Volume 5 205-228 doi: 10.1146/annurev-animal-022516-022900
Sandra O, Charpigny G, Galio L, Hue I.In mammalian species, endometrial receptivity is driven by maternal factors independently of embryo signals. When pregnancy initiates, paracrine secretions of the preattachment embryo are essential both for maternal recognition and endometrium preparation for implantation and for coordinating development of embryonic and extraembryonic tissues of the conceptus. This review mainly focuses on domestic large animal species. We first illustrate the major steps of preattachment embryo development, including elongation in bovine, ovine, porcine, and equine species. We next highlight conceptus secret...
Microvesicle-mediated Wnt/β-Catenin Signaling Promotes Interspecies Mammary Stem/Progenitor Cell Growth.
The Journal of biological chemistry    October 12, 2016   Volume 291, Issue 47 24390-24405 doi: 10.1074/jbc.M116.726117
Bussche L, Rauner G, Antonyak M, Syracuse B, McDowell M, Brown AMC, Cerione RA, Van de Walle GR.Signaling mechanisms that regulate mammary stem/progenitor cell (MaSC) self-renewal are essential for developmental changes that occur in the mammary gland during pregnancy, lactation, and involution. We observed that equine MaSCs (eMaSCs) maintain their growth potential in culture for an indefinite period, whereas canine MaSCs (cMaSCs) lose their growth potential in long term cultures. We then used this system to investigate the role of microvesicles (MVs) in promoting self-renewal properties. We found that Wnt3a and Wnt1 were expressed at higher levels in MVs isolated from eMaSCs compared wi...
In Vitro and In Vivo Development of Horse Cloned Embryos Generated with iPSCs, Mesenchymal Stromal Cells and Fetal or Adult Fibroblasts as Nuclear Donors.
PloS one    October 12, 2016   Volume 11, Issue 10 e0164049 doi: 10.1371/journal.pone.0164049
Olivera R, Moro LN, Jordan R, Luzzani C, Miriuka S, Radrizzani M, Donadeu FX, Vichera G.The demand for equine cloning as a tool to preserve high genetic value is growing worldwide; however, nuclear transfer efficiency is still very low. To address this issue, we first evaluated the effects of time from cell fusion to activation (<1h, n = 1261; 1-2h, n = 1773; 2-3h, n = 1647) on in vitro and in vivo development of equine embryos generated by cloning. Then, we evaluated the effects of using different nuclear donor cell types in two successive experiments: I) induced pluripotent stem cells (iPSCs) vs. adult fibroblasts (AF) fused to ooplasts injected with the pluripotency-inducin...
The Calcium-Sensing Receptor and the Reproductive System.
Frontiers in physiology    August 30, 2016   Volume 7 371 doi: 10.3389/fphys.2016.00371
Ellinger I.Active placental transport of maternal serum calcium (Ca(2+)) to the offspring is pivotal for proper development of the fetal skeleton as well as various organ systems. Moreover, extracellular Ca(2+) levels impact on distinct processes in mammalian reproduction. The calcium-sensing receptor (CaSR) translates changes in extracellular Ca(2+)-concentrations into cellular reactions. This review summarizes current knowledge on the expression of CaSR and its putative functions in reproductive organs. CaSR was detected in placental cells mediating materno-fetal Ca(2+)-transport such as the murine int...
A model for the development and growth of the parasitic stages of Parascaris spp. in the horse.
Veterinary parasitology    August 28, 2016   Volume 228 108-115 doi: 10.1016/j.vetpar.2016.08.019
Leathwick DM, Sauermann CW, Donecker JM, Nielsen MK.Literature documenting the growth and development of Parascaris spp. infections was used to develop a model describing worm dynamics in the young horse. The model incorporates four main variables; the rate at which larvae migrate through host tissues to return to the small intestine, the proportion of migrating larvae which succeed in returning to the small intestine, the rate of growth in size of maturing and adult worms and the survival rate of maturing and adult worms. In addition, the number of eggs laid each day by adult female worms is calculated as a function of worm size (length) and i...
Age-Related Changes in Locomotor Performance Reveal a Similar Pattern for Caenorhabditis elegans, Mus domesticus, Canis familiaris, Equus caballus, and Homo sapiens.
The journals of gerontology. Series A, Biological sciences and medical sciences    August 16, 2016   Volume 72, Issue 4 455-463 doi: 10.1093/gerona/glw136
Marck A, Berthelot G, Foulonneau V, Marc A, Antero-Jacquemin J, Noirez P, Bronikowski AM, Morgan TJ, Garland T, Carter PA, Hersen P, Di Meglio JM....Locomotion is one of the major physiological functions for most animals. Previous studies have described aging mechanisms linked to locomotor performance among different species. However, the precise dynamics of these age-related changes, and their interactions with development and senescence, are largely unknown. Here, we use the same conceptual framework to describe locomotor performances in Caenorhabditis elegans, Mus domesticus, Canis familiaris, Equus caballus, and Homo sapiens. We show that locomotion is a consistent biomarker of age-related changes, with an asymmetrical pattern througho...
Central Nervous System and Vertebrae Development in Horses: a Chronological Study with Differential Temporal Expression of Nestin and GFAP.
Journal of molecular neuroscience : MN    August 15, 2016   Volume 61, Issue 1 61-78 doi: 10.1007/s12031-016-0805-9
Rigoglio NN, Barreto RS, Favaron PO, Jacob JC, Smith LC, Gastal MO, Gastal EL, Miglino MA.The neural system is one of the earliest systems to develop and the last to be fully developed after birth. This study presents a detailed description of organogenesis of the central nervous system (CNS) at equine embryonic/fetal development between 19 and 115 days of pregnancy. The expression of two important biomarkers in the main structure of the nervous system responsible for neurogenesis in the adult individual, and in the choroid plexus, was demonstrated by Nestin and glial fibrillary acid protein (GFAP) co-labeling. In the 29th day of pregnancy in the undifferentiated lateral ventricle...
Convergent evolution of pregnancy-specific glycoproteins in human and horse.
Reproduction (Cambridge, England)    June 8, 2016   Volume 152, Issue 3 171-184 doi: 10.1530/REP-16-0236
Aleksic D, Blaschke L, Mißbach S, Hänske J, Weiß W, Handler J, Zimmermann W, Cabrera-Sharp V, Read JE, de Mestre AM, O'Riordan R, Moore T....Pregnancy-specific glycoproteins (PSGs) are members of the carcinoembryonic antigen cell adhesion molecule (CEACAM) family that are secreted by trophoblast cells. PSGs may modulate immune, angiogenic and platelet responses during pregnancy. Until now, PSGs are only found in species that have a highly invasive (hemochorial) placentation including humans, mice and rats. Surprisingly, analyzing the CEACAM gene family of the horse, which has a non-invasive epitheliochorial placenta, with the exception of the transient endometrial cups, we identified equine CEACAM family members that seem to be rel...
Placental abnormalities in equine pregnancies generated by SCNT from one donor horse.
Theriogenology    May 26, 2016   Volume 86, Issue 6 1573-1582 doi: 10.1016/j.theriogenology.2016.05.017
Pozor MA, Sheppard B, Hinrichs K, Kelleman AA, Macpherson ML, Runcan E, Choi YH, Diaw M, Mathews PM.Placental changes associated with SCNT have been described in several species, but little information is available in this area in the horse. We evaluated the ultrasonographic, gross, and histopathological characteristics of placentas from three successful and five unsuccessful equine SCNT pregnancies, established using cells from a single donor horse. Starting at approximately 6-month gestation, the pregnancies were monitored periodically using transrectal (TR) and transabdominal (TA) ultrasonography (US) to examine the placentas, fetal fluids, and fetuses. Of the five mares that aborted, one...
High Expression of Endogenous Retroviral Envelope Gene in the Equine Fetal Part of the Placenta.
PloS one    May 13, 2016   Volume 11, Issue 5 e0155603 doi: 10.1371/journal.pone.0155603
Stefanetti V, Marenzoni ML, Passamonti F, Cappelli K, Garcia-Etxebarria K, Coletti M, Capomaccio S.Endogenous retroviruses (ERVs) are proviral phases of exogenous retroviruses that have co-evolved with vertebrate genomes for millions of years. Previous studies have identified the envelope (env) protein genes of retroviral origin preferentially expressed in the placenta which suggests a role in placentation based on their membrane fusogenic capacity and therefore they have been named syncytins. Until now, all the characterized syncytins have been associated with three invasive placentation types: the endotheliochorial (Carnivora), the synepitheliochorial (Ruminantia), and the hemochorial pla...
Embryo-maternal communication during the first 4 weeks of equine pregnancy.
Theriogenology    April 21, 2016   Volume 86, Issue 1 349-354 doi: 10.1016/j.theriogenology.2016.04.048
Stout TA.The first month of equine pregnancy covers a period of rapid growth and development, during which the single-cell zygote metamorphoses into an embryo with a functional circulation and precursors of many important organs, enclosed within extraembryonic membranes responsible for nutrient uptake and gaseous exchange. After exiting the oviduct, the conceptus must influence uterine physiology to ensure adequate nutrition and preparation for implantation, while continued development results in the chorioallantois superseding the yolk sac as the primary interface for maternal interaction and exchange...
Comparative genomics of hormonal signaling in the chorioallantoic membrane of oviparous and viviparous amniotes.
General and comparative endocrinology    April 19, 2016   Volume 244 19-29 doi: 10.1016/j.ygcen.2016.04.017
Griffith OW, Brandley MC, Whittington CM, Belov K, Thompson MB.In oviparous amniotes (reptiles, birds, and mammals) the chorioallantoic membrane (CAM) lines the inside of the egg and acts as the living point of contact between the embryo and the outside world. In livebearing (viviparous) amniotes, communication during embryonic development occurs across placental tissues, which form between the uterine tissue of the mother and the CAM of the embryo. In both oviparous and viviparous taxa, the CAM is at the interface of the embryo and the external environment and can transfer signals from there to the embryo proper. To understand the evolution of placental ...
Hippotherapy: Remuneration issues impair the offering of this therapeutic strategy at Southern California rehabilitation centers.
NeuroRehabilitation    April 12, 2016   Volume 38, Issue 4 411-417 doi: 10.3233/NRE-161332
Pham C, Bitonte R.Hippotherapy is the use of equine movement in physical, occupational, or speech therapy in order to obtain functional improvements in patients. Studies show improvement in motor function and sensory processing for patients with a variety of neuromuscular disabilities, developmental disorders, or skeletal impairments as a result of using hippotherapy. Objective: The primary objective of this study is to identify the pervasiveness of hippotherapy in Southern California, and any factors that impair its utilization. Methods: One hundred and fifty-two rehabilitation centers in the Southern Californ...
Organogenesis of the Musculoskeletal System in Horse Embryos and Early Fetuses.
Anatomical record (Hoboken, N.J. : 2007)    March 15, 2016   Volume 299, Issue 6 722-729 doi: 10.1002/ar.23339
Barreto Rda S, Rodrigues MN, Carvalho RC, De Oliveira E Silva FM, Rigoglio NN, Jacob JC, Gastal EL, Miglino MA.Musculoskeletal system development involves heterotypical inductive interactions between tendons, muscles, and cartilage and knowledge on organogenesis is required for clarification of its function. The aim of this study was to describe the organogenesis of horse musculoskeletal system between 21 and 105 days of gestation, using detailed macroscopic and histological analyses focusing on essential developmental steps. At day 21 of gestation the skin was translucid, but epithelial condensation and fibrocartilaginous tissues were observed on day 25 of pregnancy. Smooth muscle was seen in lymphat...
Management of the pregnant mare and long-term consequences on the offspring.
Theriogenology    February 4, 2016   Volume 86, Issue 1 99-109 doi: 10.1016/j.theriogenology.2016.01.028
Peugnet P, Robles M, Wimel L, Tarrade A, Chavatte-Palmer P.The study of early developmental conditioning of health and disease in adulthood is particularly relevant in the horse, which is bred mainly to perform in demanding sport challenges. On the basis of this concept, the management of the broodmare could be considered an effective means to produce animals with the desired features. Knowledge on the Developmental Origins of Health and Disease in the equine species remains relatively scarce, with some experimental studies and one single epidemiologic study. Data highlight the determinant role of the maternal environment for postnatal body conformati...
Campylorrhinus lateralis, Bilateral microphthalmia and odontoma temporalis in an Oldenburg Foal.
Reproduction in domestic animals = Zuchthygiene    January 30, 2016   Volume 51, Issue 2 330-334 doi: 10.1111/rda.12665
Casteleyn C, Cornillie P, Tüllmann V, Van Cruchten S, Van Ginneken C.An Oldenburg colt with wry nose was autopsied after having lived for only 30 min. It presented cyanotic oral mucosae, underdeveloped eyes and a right-sided temporal osseous mass. The applicable nomenclature for the defects is discussed, and the potential etiopathogenesis is explored by describing the normal embryonic development of the affected body parts.
Equine-Induced Pluripotent Stem Cells Retain Lineage Commitment Toward Myogenic and Chondrogenic Fates.
Stem cell reports    January 16, 2016   Volume 6, Issue 1 55-63 doi: 10.1016/j.stemcr.2015.12.005
Quattrocelli M, Giacomazzi G, Broeckx SY, Ceelen L, Bolca S, Spaas JH, Sampaolesi M.Induced pluripotent stem cells (iPSCs) hold great potential not only for human but also for veterinary purposes. The equine industry must often deal with health issues concerning muscle and cartilage, where comprehensive regenerative strategies are still missing. In this regard, a still open question is whether equine iPSCs differentiate toward muscle and cartilage, and whether donor cell type influences their differentiation potential. We addressed these questions through an isogenic system of equine iPSCs obtained from myogenic mesoangioblasts (MAB-iPSCs) and chondrogenic mesenchymal stem ce...
A method for isolating and culturing placental cells from failed early equine pregnancies.
Placenta    December 24, 2015   Volume 38 107-111 doi: 10.1016/j.placenta.2015.12.014
Rose BV, Cabrera-Sharp V, Firth MJ, Barrelet FE, Bate S, Cameron IJ, Crabtree JR, Crowhurst J, McGladdery AJ, Neal H, Pynn J, Pynn OD, Smith C....Early pregnancy loss occurs in 6-10% of equine pregnancies making it the main cause of reproductive wastage. Despite this, reasons for the losses are known in only 16% of cases. Lack of viable conceptus material has inhibited investigations of many potential genetic and pathological causes. We present a method for isolating and culturing placental cells from failed early equine pregnancies. Trophoblast cells from 18/30 (60%) failed equine pregnancies of gestational ages 14-65 days were successfully cultured in three different media, with the greatest growth achieved for cells cultured in Amnio...
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