Biomedical subjects

A C Cuello

Publications and source records attributed to A C Cuello.

At least 91 records · Page 5Linked to original sources

PubMed1992

Glial fibrillary acidic protein immunoreactivity following cortical devascularizing lesion.

Disruption of a restricted area of the pia-arachnoid compromises vascular irrigation of the underlying cortex, leading to infarction of the tissue. This study was undertaken to examine the effects of such brain injury on glial cells. Adult male Wistar rats were processed for glial fibrillary acidic protein immunoreactivity at 1, 4, 7, 15 and 30 days after undergoing cortical devascularization. One day post-lesion, glial fibrillary acidic protein-positive cells were observed only at the lesion site. Glial fibrillary acidic protein-positive cells were present in the ipsilateral remaining cortex, distant from the wound, between days 4 and 15, and were also evident in the thalamus beginning 4 days post-lesion. These diverse temporal patterns of GFAP immunoreactivity in different brain structures suggest that various mechanisms can mediate increased GFAP immunostaining following injury.

Animals↗

PubMed1992

Epidermal growth factor affects both glia and cholinergic neurons in septal cell cultures.

The effects of epidermal growth factor on high density primary cultures of fetal (embryonic day 17) rat septal cells were examined. Under serum-free conditions, the continuous exposure of these cultures to epidermal growth factor for seven days significantly decreased choline acetyltransferase (EC 2.3.1.6) activity in a dose-dependent manner. Maximal decreases were observed from 1 to 10 ng/ml epidermal growth factor. This effect was completely abolished by the addition of anti-epidermal growth factor antibodies. The epidermal growth factor-mediated decrease in choline acetyltransferase activity was culture-time dependent, being first detectable after five days of factor application and may likely represent an inhibition of the spontaneous increase in enzyme activity that occurs with time in culture. Concomitant with changes in enzyme activity, epidermal growth factor produced a significant and proportional decrease in the number of acetylcholinesterase-positive neurons. This decrease in acetylcholinesterase-positive cells did not reflect a decrease in cholinergic cell survival as nerve growth factor could restore the number of acetylcholinesterase-positive neurons in epidermal growth factor-treated cultures to control levels. Furthermore, in these high-density cultures, epidermal growth factor did not affect general neuronal survival, while it did produce an increase in the number and intensity of glial fibrillary acidic protein-immunoreactive astroglia as well as in the number of macrophage-like cells. The proliferative response of these non-neuronal cells to epidermal growth factor, as assessed by [3H]thymidine incorporation, was evident after three days of epidermal growth factor application, persisted thereafter, and could be antagonized by the inclusion of the antimitotic 5-fluorodeoxyuridine. Furthermore, 5-fluorodeoxyuridine completely blocked the epidermal growth factor-mediated decrease in choline acetyltransferase activity. However, when epidermal growth factor was tested in pure glial cultures, it only directly induced proliferation of astrocytes. These results suggest that the proliferative response of either one or both of these glial cell types in the mixed cultures may be indirectly affecting cholinergic cell expression.

Acetylcholinesterase↗

PubMed1992

Enkephalin-immunoreactive nociceptive neurons in the cat spinal cord.

In this combined electrophysiological-ultrastructural study in the cat spinal cord, we detected enkephalin-like immunoreactivity using internally radio-labelled monoclonal antibodies in functionally characterized neurons which had been filled intracellularly with horseradish peroxidase. Of the 4 neurons included in this study, two were positive for enkephalin immunoreactivity; one was a nociceptive specific neuron in lamina I, the other a wide dynamic range neuron in lamina V. The other two cells were devoid of immunoreactivity for enkephalin; one was a wide dynamic range neuron and the other was a non-nociceptive neuron. These results thus provide a morphological substrate within the spinal dorsal horn for the release of an endogenous opioid following administration of substance P or noxious cutaneous stimulation.

Animals↗

PubMed1992

Recovery of nucleus basalis cholinergic neurons by grafting NGF secretor fibroblasts.

The effects of nerve growth factor (NGF) secreted from genetically modified fibroblasts was studied in vitro, using dissociated septal cells and in vivo, in rats bearing unilateral cortical devascularizing lesions. Transfected fibroblasts expressing nerve growth factor (NGF) were co-cultured with rat embryonic cholinergic cells of the septal region. This in vitro system showed that NGF secretor cells produce biologically active NGF, as determined by increasing choline acetyltransferase (ChAT) activity in septal culture after seven days. The potential therapeutic value of applying grafts of transfected fibroblasts expressing NGF in the model of retrograde atrophy of cholinergic neurons of the nucleus basalis magnocellularis (NBM) was assessed following partial devascularizing lesions of the cerebral cortex. We observed an increase in ChAT activity in the remaining cortex and a partial protection of the ipsilateral NBM, as determined by morphometric and biochemical studies.

Animals↗

PubMed1991

Similarities in the ultrastructural distribution of nerve growth factor receptor-like immunoreactivity in cerebellar Purkinje cells of the neonatal and colchicine-treated adult rat.

The intracellular distribution of nerve growth factor receptor immunoreactivity was examined by electron microscopy in the cerebellum of adult and postnatal day 12 rats. The very faint immunostaining in Purkinje cells of naive adult animals was greatly amplified after colchicine treatment. Neonatal cerebellum, in contrast, contained prominent immunoreactivity in both Purkinje cells and germinal cells of the external granular layer. Intracellular distribution of the nerve growth factor receptor reaction product was very similar in Purkinje cells of both neonatal and colchicine-treated adult animals. It was consistently present along the perikaryal cell membrane, in segments of the rough endoplasmic reticulum and Golgi complex. Numerous membrane-bound aggregates of immunoreactive vesicles resembling multivesicular bodies (secondary lysosomes) were scattered throughout the cell soma, although less frequently in neonatal rats. Bulbous expansions along the proximal axons of colchicine-treated Purkinje cells were filled with such immunoreactive multivesicular bodies. These cells also displayed evidence of nerve growth factor receptor internalization in the form of immunoreactive coated vesicles situated near the cell membrane. In addition to the staining in Purkinje cells, neonatal cerebellum contained high amounts of nerve growth factor receptor reaction product along the cell membrane of germinal cells in the external granular layer. Although Purkinje cells of naive adult animals possessed little or no cell membrane-related nerve growth factor receptor immunoreactivity, reaction product was sometimes seen in cisternae of the rough endoplasmic reticulum and Golgi apparatus. These findings provide electron microscopic immunocytochemical evidence of nerve growth factor receptor synthesis, internalization, and catabolism in noncholinergic neurons of the central nervous system.

Aging↗

PubMed1991

Two distinct monoclonal antibodies raised against mouse beta nerve growth factor. Generation of bi-specific anti-nerve growth factor anti-horseradish peroxidase antibodies for use in a homogeneous enzyme immunoassay.

Two hybridomas producing monoclonal antibodies against mouse beta nerve growth factor (NGF) were obtained from the fusion of hyperimmune splenocytes from rats immunized with polymerized beta-NGF and Sp2/0.Ag mouse myeloma cells. The monoclonal antibodies coded IgG 24 and 30 produced and secreted by the hybrid cells are both of the IgG2a subclass. Both monoclonal antibodies are capable of recognizing native NGF coated on microassay plates as well as the denatured factor on Western blots. However, only IgG 30 has been found to block NGF-induced process outgrowth from the rat pheochromocytoma cell line (PC12) as well as NGF-induced increase in choline acetyltransferase activity in rat primary septal cell cultures. In addition, only IgG 30 was able to detect immunocytochemically NGF-immunoreactive sites in fixed tissue. And, finally, IgG 24 could not compete for IgG 30 binding to immobilized native NGF. Consequently, it appears that these antibodies are recognizing different epitopes on the NGF molecule. Neither monoclonal antibody displayed any crossreactivity with serum albumin, aprotinin, epidermal growth factor or insulin. A hybrid-hybridoma producing bi-specific anti-NGF anti-horseradish peroxidase (HRP) monoclonal antibodies was generated from the fusion of an azaguanine resistant anti-HRP hybridoma, coded RAP2.Ag and the anti-NGF IgG 30 hybridoma treated with emetine. The potential merits of using these bi-specific antibodies in combination with their mono-specific anti-NGF parent in a homogeneous sandwich immunoassay for the quantitation of NGF are discussed.

Animals↗

PubMed1991

Production and immunocytochemical application of a highly sensitive and specific monoclonal antibody against rat dopamine-beta-hydroxylase.

A highly sensitive and specific monoclonal antibody against the enzyme dopamine beta-hydroxylase (DBH) from rat was produced and coded DBH 41. The generated hybridoma secreted immunoglobulins of mouse IgG1 subtype, as determined by radial immunodiffusion. This antibody, characterized by immunoblotting against a crude rat DBH preparation, was found to specifically recognize two bands of molecular weight 70 and 75 kDa corresponding to the soluble and membrane bound forms of the enzyme, respectively. With regard to species specificity, the anti-DBH antibody recognizes only the rat DBH molecule as it exhibits no cross-reactivity with either mouse, human, rabbit, guinea pig, cat or bovine DBH. Comparative immunocytochemical localization of DBH and TOH immunoreactivity was performed in different brain regions and we found that the DBH 41 antibody specifically stained DBH-containing neurons and fibers in the rat central nervous system (CNS). The high sensitivity of the DBH 41 antibody permitted us to detect immunologically the presence of the enzyme even in areas where only scattered DBH-containing fibers were present.

Animals↗

PubMed1991

Light and electron microscopic distribution of nerve growth factor receptor-like immunoreactivity in the skin of the rat lower lip.

Nerve growth factor receptor distribution in the skin of the adult rat was studied by immunocytochemistry with the use of the monoclonal antibody 192-IgG. Immunoreaction occurred in a patchy pattern in the epidermis and outer root sheaths of hair follicles, as well as in nerves and in capsulated and non-capsulated sensory receptors. Electron microscopic analysis revealed that the areas of patchy epithelial immunostaining corresponded to the plasma membrane of groups of keratinocytes, which were always associated with intra-epithelial nerve fibres. Immunostaining was also associated with Merkel cells, hair follicle receptors, and the capsular cells of capsulated receptors. In the nerves, immunostaining was detected in perineurial cells and axons. After sensory denervation of the skin, immunostaining decreased considerably in the epithelia, as well as in the small nerves, although short segments of increased staining were observed in the latter. In larger nerves, immunostaining was markedly increased and mainly associated with newly formed Schwann cell processes. After sympathetic denervation, the pattern of immunostaining was identical to controls, except for augmented immunoreactivity in short segments of degenerating nerves around blood vessels and smooth muscle fibres. From these observations, it is suggested that the epithelial and perineurial cells synthesize and express nerve growth factor receptors, while receptors present in nerve fibres originate from the nerve cell bodies in the gasserian ganglion.

Animals↗

PubMed1991

A progressive deposition of paired helical filaments (PHF) in the brain characterizes the evolution of dementia in Alzheimer's disease. An immunocytochemical study with a monoclonal antibody against the PHF core.

Using the monoclonal antibody (mAb) 6.423 which recognizes epitopes of the pronase-resistant core of paired helical filaments (PHF), we studied postmortem frontal cortex from Alzheimer's disease (AD) patients with short (Group II) and long (Group III) histories of clinical dementia. Four cases with clinically unconfirmed dementia and a postmortem diagnosis of AD (Group I) were also studied. In Group I, the 6,423 mAb was negative whereas in Group II, the antibody recognized primarily neurofibrillary tangles (NFT). In contrast, brains in Group III contained a dense network of 6,423-immunoreactive (IR) thread-like structures ("ghost" neurites) and plaque-like structures with granular appearance, in addition to NFT. The number of 6,423-IR structures appeared to be related to the duration of clinical dementia and the age of onset. Furthermore, "ghost" neurites were more abundant in young AD cases. The possible significance of the 6,423-IR pattern in the pathogenesis of AD is discussed.

Adult↗

PubMed1991

Substance P- and enkephalin-like immunoreactivities are colocalized in certain neurons of the substantia gelatinosa of the rat spinal cord: an ultrastructural double-labeling study.

The finding that certain cells of the substantia gelatinosa of the rat spinal cord contain both substance P (SP)- and enkephalin (ENK)-like immunoreactive material offers new insights into the mechanisms of action of these peptides in the processing of nociceptive sensory information. The simultaneous detection of these immunoreactivities was obtained in the superficial dorsal horn of the rat spinal cord at the ultrastructural level using monoclonal antibodies. An internally radiolabeled monoclonal antibody (against SP or ENK) was used to recognize one antigenic site, while the other antigenic site was identified by either a bispecific monoclonal antibody (for SP) or a monoclonal antibody (for ENK). The bispecific anti-SP antibody recognized HRP, whereas a secondary bispecific antibody recognized both the IgG of the anti-ENK monoclonal antibody and HRP. In laminae I-III, SP-like immunoreactivity (SP-LI) and ENK-like immunoreactivity (ENK-LI) were colocalized in a significant number of axonal varicosities, which contained round or pleomorphic synaptic vesicles. Such double-labeled varicosities, however, were not found to be components of synaptic glomeruli. Most of the immunostained boutons of lamina I were SP-like immunoreactive only. In rats pretreated with colchicine, SP-LI and ENK-LI were colocalized in small perikarya of lamina II and in some lamina I cells. These findings indicate that SP and ENK occur in a significant population of interneurons of the superficial dorsal horn. It is suggested that some of these neurons may correspond to stalked cells and release one or the other substance depending on physiological conditions.

Animals↗

PubMed1990

Effects of treatment with microencapsulated monosialoganglioside GM1 on cortical and striatal acetylcholine release in rats with cortical devascularizing lesions.

The present study shows a novel administration form of the monoganglioside GM1, which following microencapsulation in human serum albumin was topically applied on cortical regions damaged by devascularization in rats. The effects of microencapsulated GM1 on extracellular levels of acetylcholine, choline and dopamine in the cortex and in the striatum were analyzed using in vivo microdialysis. Cholinergic neurons in the nucleus basalis magnocellularis were studied immunohistochemically using monoclonal antibodies raised against choline acetyltransferase (ChAT). It was found that cortical devascularizing lesions produced a decrease in extracellular levels of cortical acetylcholine and choline, and retrograde morphological changes in cholinergic neurons in the nucleus basalis magnocellularis. GM1 promoted (1) recovery of the retrograde morphological changes produced by the decortication in the nucleus basalis magnocellularis and (2) a parallel increase in cortical acetylcholine release. No changes were observed in the striatum, nor on cortical or striatal dopamine levels simultaneously measured in the same perfusates.

Acetylcholine↗

PubMed1990

Immunoelectron microscopic evidence of nerve growth factor receptor metabolism and internalization in rat nucleus basalis neurons.

Nerve growth factor receptor (NGFr) immunoreactive neurons of the adult rat nucleus basalis magnocellularis were examined by electron microscopy. Prominent NGFr immunoreactivity (IR) was consistently present along the perikaryal cell membrane and frequently in intracellular sites of protein synthesis and modification such as the rough endoplasmic reticulum and Golgi apparatus, respectively. Immunoperoxidase reaction product was also seen along the nuclear membrane. Membrane-bound aggregates of immunoreactive vesicles were scattered throughout the perikaryon, being more concentrated in the perinuclear region and in the proximal neurites. These may represent either aggregates of receptor-containing vesicles on their way to/from the cell membrane or secondary lysosomes where NGFr reaction product is degraded. Immunostained cytoplasmic vesicles which possessed an electron-dense coat and were adjacent to or contiguous with the plasmalemma probably represented internalized receptor. This ultrastructural study of the subcellular distribution of NGFr-IR in basal forebrain neurons therefore demonstrates sites of receptor metabolism and potential receptor-ligand interaction.

Animals↗

PubMed1990

Choline acetyltransferase-immunoreactive profiles are presynaptic to primary sensory fibers in the rat superficial dorsal horn.

The specific aim of this study was to search for morphological counterparts to the known antinociceptive effects of cholinomimetic drugs at the spinal cord level. For this, the light microscopic and ultrastructural distribution of choline acetyltransferase immunoreactivity was studied in laminae I-III of the rat cervical spinal cord. Immunoreactivity was present in cell bodies in lamina III, and in dendrites and axons of all three laminae. Immunoreactive axonal varicosities were often presynaptic to the central varicosities of type II synaptic glomeruli in lamina II and lamina III, less often presynaptic to the central elements of type I glomeruli in lamina II, and often presynaptic to dendrites in both type I and type II glomeruli. In addition, immunoreactive dendrites were often postsynaptic to the central varicosities of glomeruli of all morphological types. These results indicate that 1) primary sensory fibers excite cholinergic interneurons; 2) the acetylcholine released by the axon terminals of these interneurons modulates both nociceptive and non-nociceptive sensory information at the spinal cord level through both pre- and postsynaptic mechanisms. Furthermore, our results reinforce current ideas on reciprocal sensory interaction between thick and fine afferent fibers.

Animals↗

PubMed1990

Derivatives of ganglioside GM1 as neuronotrophic agents: comparison of in vivo and in vitro effects.

Exogenously administered gangliosides have been shown to behave as neuronotrophic/neuritogenic agents in a variety of cell culture systems and animal models, but it is not known whether they operate by the same mechanism in vivo and in vitro. To probe this question we have employed two derivatives of GM1 lacking the negative charge: the methyl ester (GM1-CH3) and the NaBH4 reduction product of the latter (GM1-OH) in which the carboxyl group is replaced by a primary alcohol. Both derivatives proved to be as neuritogenic as GM1 in 3 cell culture systems: neuro-2A cels, PC12 cells and explanted dorsal root ganglia. However, GM1-OH proved ineffective when applied to two animal models involving reduction of cholinergic markers in: (a) hippocampus following lesion of the lateral fimbria and (b) nucleus basalis magnocellularis following cortical lesion; GM1-CH3 showed marginal activity in (a) but more in (b), possibly owing to slow hydrolysis to GM1 which was highly active in both animal models. These results indicate the necessity of a negative change on the ganglioside molecule for in vivo but not in vitro activity and point to different mechanisms for the trophic effects of exogenous gangliosides.

Animals↗

PubMed1990

5-Hydroxytryptamine, substance P, and thyrotropin-releasing hormone in the adult cat spinal cord segment L7: immunohistochemical and chemical studies.

The terminal projections of the descending 5-hydroxytryptamine (5-HT) bulbospinal pathway and the coexistence among 5-HT-, substance P (SP)-, and thyrotropin-releasing hormone (TRH)-like immunoreactivities (LI) in fibers innervating the L7 segment in the cat spinal cord were studied quantitatively and semiquantitatively by use of the indirect double-staining immunofluorescence technique. The content of 5-HT, SP, and TRH in different parts of the spinal cord was determined by use of radioimmunoassay (RIA) (SP and TRH) and high-performance liquid chromatography with electrochemical detection (HPLC-ECD) (5-HT). For all three substances studied, immunoreactive (IR) axon terminals were found in all parts of the gray matter, but with clear regional variation in the density of innervation. Thus, all three substances showed a dense innervation in the motor nucleus, particularly in the ventral part of the nucleus, while the superficial dorsal horn was very densely innervated by SP-IR fibers (laminae I and II) and TRH-IR fibers (laminae II and III). In the motor nucleus, the studied substances coexisted to a very high degree, but some 5-HT-IR fibers (about 10%) lacked peptide-LI and some SP-IR fibers (about 10%) lacked 5-HT-LI while virtually all TRH-IR fibers also contained 5-HT-LI. In the superficial dorsal horn (laminae I-III), no coexistence was detected, while other parts of the gray matter displayed various degrees of coexistence in between those found in the motor nucleus and laminae I-III. The quantitative analysis of IR varicosities in the motor nucleus suggested that the unilateral L7 motor nucleus is innervated by about 55-110 x 10(6) 5-HT-IR nerve terminals, which may indicate as many as 4,000 boutons per descending 5-HT cell body in the brain stem only with this restricted projection. When combing these results with the biochemical data, it could be calculated that the concentration of 5-HT in IR varicosities is about 3-6 x 10(-3) M, while the corresponding figures for SP and TRH was 0.3-0.5 x 10(-3) M and 0.1-0.2 x 10(-3) M, respectively. In cats subjected to spinal cord transection at the lower thoracic level, all 5-HT-IR fibers in the L7 segment had disappeared 44 days after the lesion, indicating a strict suprasegmental origin of 5-HT-IR fibers in this segment.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗

PubMed1990

Production of a bi-specific monoclonal antibody recognizing mouse kappa light chains and horseradish peroxidase. Applications in immunoassays.

The production of a bi-specific monoclonal antibody that simultaneously recognizes mouse kappa light chains and horseradish peroxidase (HRP) for use as a general developing reagent in a wide variety of immunobased techniques is described. This antibody, named McC10, was produced by the fusion of an aminopterin-sensitive interspecies hybridoma which secretes rat monoclonal antibodies against HRP (RAP2.Ag) and splenocytes from a rat immunized with whole mouse immunoglobulin (Ig)G. The hybrid-hybridoma generated from this fusion expresses and secretes rat Igs of the IgG1 and IgG2a subclasses, as determined by radial immunodiffusion. In competitive binding solid-phase enzymatic assays, McC10 was found to cross-react with all four mouse IgG subclasses as well as mouse kappa light chains. In contrast, in this type of assay, McC10 did not appear to recognize mouse IgA, IgM or lambda light chains. However, IgM-bearing kappa light chains were recognized by immunocytochemistry. Epitope specificity of this bi-specific antibody was more clearly determined on immunoblots where McC10 was found to exclusively recognize mouse kappa light chains and display no cross-reactivity with mouse Ig heavy chains nor with kappa light chains from rat or rabbit. In addition, McC10 was used successfully in two-step immunocytochemistry (ICC) for the localization of enkephalin, nerve growth factor (NGF) receptor and paired helical filament-immunoreactive sites in rat brain, rat skin and human brain, respectively, using mouse IgG's and IgM's as primary antibodies. McC10 compared favourably with peroxidase-anti-peroxidase (PAP) ICC with respect to sensitivity but was markedly superior with respect to specificity when used in fixed human brain or rat skin.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1990

Acetylcholine release in vivo: effects of chronic treatment with monosialoganglioside GM1.

Rats with unilateral cortical devascularizing lesions were treated with the monosialoganglioside GM1 in two different ways. One group of animals received GM1 (5 mg/kg/day, for 7 days), through a permanent cannula implanted intracerebroventricularly (i.c.v.) and connected to an osmotic minipump. The other group was treated with microencapsulated GM1 placed directly onto the surface of the lesioned cortex. The effect of GM1, administered into the lateral ventricle and supracortically, on the release of ACh in vivo was studied, using a microdialysis system combined with sensitive high performance liquid chromatography (HPLC). The release of acetylcholine and choline was studied in the cortex and striatum of the rat under nonstimulated (basal) and KCl (100 mM)-stimulating conditions. The non-stimulated release of acetylcholine was only measurable in the presence of neostigmine and was found to be about 30 microM in the cortex and approximately 10 times greater in the striatum. A large concentration of KCl led to a remarkable increase of acetylcholine in the control (C) and vehicle-treated lesioned groups (V i.c.v., V cap: 11-13 fold), but was greater in the GM1-treated groups (GM1 i.c.v., GM1 cap: 20-25 fold). In contrast, KCl-stimulated release of ACh in striata from GM1-treated lesioned groups was significantly less (5-10 fold), compared to the unlesioned controls (C: 16 fold) and lesioned vehicle-treated rats (V i.c.v. and V cap: 16-18 fold). The release of choline was not increased significantly by large concentrations of KCl present in the perfusion medium. In all experimental groups, and in both structures of the brain, ratios between stimulated and non-stimulated release of choline was between 0.7-1.2.(ABSTRACT TRUNCATED AT 250 WORDS)

Acetylcholine↗

PubMed1990

Three-dimensional reconstruction and quantitative evaluation of devascularizing cortical lesions in the rat.

The use of a reconstruction method suitable for three-dimensional (3-D) representations of the cortex for the quantitative evaluation of unilateral devascularizing cortical lesions is reported, demonstrating that 3-D reconstructions can be used for quantitative evaluation of the extent of cortical lesions, in addition to simply visualizing shapes. The procedure allowed the definition of precise quantitative parameters characterizing the volumes and surface areas of the cortical regions involved. It was shown that digitized serial sections from atlases can be utilized for multiple correlation studies by superimposing morphological and biochemical parameters. More specifically, the enzymatic activity of choline acetyltransferase (ChAT) in the nucleus basalis magnocellularis (NBM), and morphological parameters (number and cross-sectional areas) of the ChAT-positive immunoreactive cholinergic neurons, were related to the numerical values for the volume and surface area of the cortical lesions. The applicability of this method goes beyond the scope illustrated in this study. For example, various morphological anomalies in the brain could be quantified; changes in various cortical and subcortical structures could be followed during development; and the 3-D size, shape and position of a graft could be related to the number of viable foetal cells.

Animals↗