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L Probert

Publications and source records attributed to L Probert.

At least 37 records · Page 2Linked to original sources
PubMed1997

TNF and related molecules: trends in neuroscience and clinical applications.

A clear message to emerge from the recent 6th International TNF Congress is that TNF-alpha, LT-alpha and possibly other TNF family members, are important integral mediators of the CNS stress response to threatened homeostasis and that either excessive or insufficient TNF-alpha production can have significant consequences upon correct CNS functioning. Experimental data are particularly relevant in light of recent evidence which shows some linkage between polymorphisms in the human TNF-alpha/LT-alpha locus and susceptibility to CNS infection and thus highlight such cytokine pathways as promising therapeutic targets for the management of certain CNS diseases. Judging by the significant advances that have been made recently in the field of cytokine neurobiology, and the currently explosive development of transgenic and gene mutant mouse models with which to probe the cellular and molecular mechanisms of TNF-alpha and LT-alpha action within the CNS, we can look forward to shortly understanding more precisely the important and diverse roles that these cytokines play in CNS health and disease.

Animals↗
PubMed1997

TNF-alpha transgenic and knockout models of CNS inflammation and degeneration.

Tumour necrosis factor-alpha (TNF-alpha) plays a central role in inflammatory events including those taking place in the central nervous system (CNS), and has been implicated as a key pathogenic mediator in several human inflammatory, infectious and autoimmune CNS disorders. Using transgenic and gene knockout mice we have investigated the role of deregulated TNF-alpha production in the CNS. We show that the overexpression of wild-type murine or human TNF-alpha transgenes by resident CNS astrocytes or neurons in sufficient to trigger a neurological disorder characterised by ataxia, seizures and paresis, with histopathological features of chronic CNS inflammation and white matter degeneration. Furthermore, we show that transmembrane human TNF-alpha is sufficient to trigger CNS inflammation and degeneration when overexpressed by astrocytes but not by neurons, indicating that target cells mediating the neuroinflammatory activities of TNF-alpha localise in the vicinity of astrocytes rather than neurons. Our results establish that both soluble and transmembrane molecular forms of TNF-alpha can play critical roles in vivo in the pathogenesis of CNS inflammation and demyelination, and validate TNF-alpha transgenic and mutant mice as important models for the further study of related human CNS diseases.

Animals↗
PubMed1996

Dissection of the pathologies induced by transmembrane and wild-type tumor necrosis factor in transgenic mice.

With increasing awareness that seemingly diverse immune-mediated diseases involve similar pathogenetic mechanisms, and the identification of a growing number of key effector molecules, it is becoming possible to design and generate effective transgenic models for such diseases. Tumor necrosis factor (TNF) plays a prominent role in immune and host defense responses and there is strong evidence that abnormal TNF production contributes to disease initiation and progression in rheumatoid arthritis, systemic inflammatory response syndrome, diabetes, multiple sclerosis, and many other immune-mediated disorders. The generation of TNF transgenic mice, in which TNF production is deregulated, has provided us with direct evidence that, in vivo, this cytokine can indeed trigger the development of such complex disease phenotypes. Transgenic mice that have been engineered to overexpress human or murine TNF molecules in peripheral joints, T cells, or neurons of the central nervous system represent important animal models for human rheumatoid arthritis, systemic inflammation, and multiple sclerosis, respectively. In addition to establishing a central role for TNF in such diseases, these animal models have already proved valuable for identifying additional important disease-effector molecules, and for gaining an insight into the complex in vivo mechanisms that are involved in disease pathogenesis. For example, in the case of arthritis, TNF has been found to transmit its pathogenic effects entirely through interleukin-1, which may therefore represent an additional important target for therapeutic intervention in the human disease. In summary, TNF transgenic models of human disease are expected to serve as important in vivo tools for defining details of disease pathogenesis, potential targets for therapeutic intervention and for evaluating the possible involvement of additional genetic and environmental factors on the disease state.

Animals↗
PubMed1996

Neurobehavioral alterations in developing transgenic mice expressing TNF-alpha in the brain.

During development, neuronal circuitry and memory formation are associated with the synthesis and release of several biological mediators, including cytokines. Among the numerous cytokines, the role of tumor necrosis factor-alpha (TNF-alpha) in neurobehavioral development is largely unknown. Thus, the recently generated transgenic mice expressing murine TNF-alpha in the brain represent a valid animal model for investigating the role of TNF-alpha in neurobehavioral processes. Using these mice, we showed that an overexpression of murine TNF-alpha increases grooming in the novel object investigation test, decreases rearing as a reaction to novel olfactory cues, and produces a retardation of passive avoidance acquisition while enhancing the thermal response in the hot-plate test, a task regulated by both peripheral and central mechanisms. The possibility that these effects are associated with endogenous changes in concentration of the NGF, known to be modulated by TNF-alpha, is discussed.

Animals↗
PubMed1995

Spontaneous inflammatory demyelinating disease in transgenic mice showing central nervous system-specific expression of tumor necrosis factor alpha.

Cytokines are now recognized to play important roles in the physiology of the central nervous system (CNS) during health and disease. Tumor necrosis factor alpha (TNF-alpha) has been implicated in the pathogenesis of several human CNS disorders including multiple sclerosis, AIDS dementia, and cerebral malaria. We have generated transgenic mice that constitutively express a murine TNF-alpha transgene, under the control of its own promoter, specifically in their CNS and that spontaneously develop a chronic inflammatory demyelinating disease with 100% penetrance from around 3-8 weeks of age. High-level expression of the transgene was seen in neurons distributed throughout the brain. Disease is manifested by ataxia, seizures, and paresis and leads to early death. Histopathological analysis revealed infiltration of the meninges and CNS parenchyma by CD4+ and CD8+ T lymphocytes, widespread reactive astrocytosis and microgliosis, and focal demyelination. The direct action of TNF-alpha in the pathogenesis of this disease was confirmed by peripheral administration of a neutralizing anti-murine TNF-alpha antibody. This treatment completely prevented the development of neurological symptoms, T-cell infiltration into the CNS parenchyma, astrocytosis, and demyelination, and greatly reduced the severity of reactive microgliosis. These results demonstrate that overexpression of TNF-alpha in the CNS can cause abnormalities in nervous system structure and function. The disease induced in TNF-alpha transgenic mice shows clinical and histopathological features characteristic of inflammatory demyelinating CNS disorders in humans, and these mice represent a relevant in vivo model for their further study.

Animals↗
PubMed1995

The type I interleukin-1 receptor acts in series with tumor necrosis factor (TNF) to induce arthritis in TNF-transgenic mice.

The pro-inflammatory cytokines tumor necrosis factor (TNF) and interleukin-1 (IL-1) have been shown to be primary mediators in the pathogenesis of chronic inflammatory joint diseases. However, the relative contributions of these molecules to the development and progression of disease is not known. In the present study, we have investigated the involvement of the type I IL-1 receptor in the development and progression of chronic arthritis in a previously described TNF-transgenic mouse model of this disease. A neutralizing monoclonal antibody to the murine type I IL-1 receptor was injected intraperitoneally three times a week from birth to 9 weeks of age. In the positive control group of untreated transgenic mice the incidence of arthritis was 100% after 9 weeks of age. The injection of normal hamster IgG did not influence the incidence or development of arthritis. In contrast, the injection of antibody to the type I IL-1 receptor completely prevented the development of arthritis. Clinical evaluation of disease was confirmed histologically, anti-receptor antibody-treated mice showed no sign of arthritic change. Moreover, the analysis of sera taken at the end of the study period showed that the neutralization of arthritis by IL-1 receptor antibody treatment was accompanied by significantly lower levels of circulating human TNF compared to the control groups and to untreated transgenic mice prior to disease onset. Taken together, these results show that IL-1 signaling blockade exerts a direct negative feedback effect on TNF production. Our results show that in TNF-transgenic mice the IL-1 receptor accepts the whole pathogenic load of TNF, thereby acting as a potent downstream mediator in the pathogenesis of chronic arthritis.

Animals↗
PubMed1995

The mouse/human chimeric monoclonal antibody cA2 neutralizes TNF in vitro and protects transgenic mice from cachexia and TNF lethality in vivo.

The pleiotropic cytokine tumour necrosis factor-alpha (TNF) is thought to play a central role in infectious, inflammatory and autoimmune diseases. Critical to the understanding and management of TNF-associated pathology is the development of highly specific agents capable of modifying TNF activity. We evaluated the ability of a high affinity mouse/human chimeric anti-TNF monoclonal antibody (cA2) to neutralize the in vitro and in vivo biological effects of TNF. cA2 inhibited TNF-induced mitogenesis and IL-6 secretion by human fibroblasts, TNF-priming of human neutrophils, and the stimulation of human umbilical vein endothelial cells by TNF as measured by the expression of E-selectin, ICAM-1 and procoagulant activity. cA2 also specifically blocked TNF-induced adherence of human neutrophils to an endothelial cell monolayer. Receptor binding studies suggested that neutralization resulted from cA2 blocking of TNF binding to both p55 and p75 TNF receptors on the cells. In vivo, repeated administration of cA2 to transgenic mice that constitutively express human TNF reversed the cachectic phenotype and prevented subsequent mortality. These results demonstrated that cA2 effectively neutralized a broad range of TNF biological activities both in vitro and in vivo.

Animals↗
PubMed1995

Effect of NGF antibodies on mast cell distribution, histamine and substance P levels in the knee joint of TNF-arthritic transgenic mice.

We have previously shown an increase in nerve growth factor (NGF) levels and in mast cell (MC) distribution in the synovium of patients affected by rheumatoid arthritis. We now report that purified NGF antibodies injected into arthritic transgenic mice carrying the human tumour necrosis factor-alpha (TNF-alpha) gene caused reduction in the number of MCs, as well as a decrease in histamine and substance P levels within the synovium. These observations suggest that NGF antibody might be useful in studying the role of these pro-inflammatory markers in joint arthritis.

Animals↗
PubMed1994

Anaemia and resistance to malaria in transgenic mice expressing human tumour necrosis factor.

Transgenic mice carrying a modified human tumour necrosis factor (huTNF)/beta-globin gene construct linked to the T-cell-specific locus control region of the human CD2 gene express huTNF in their T cells which is released into the circulation and causes the development of a wasting syndrome. We now report that the mice develop anaemia, probably through enhanced erythrophagocytosis rather than inhibition of reticulocyte production. Thus autologous erythrocytes, as well as sheep erythrocytes, were cleared more rapidly from the circulation of transgenic mice than from littermate controls. By contrast, peritoneal macrophages from transgenic mice were less phagocytic in vitro than cells from controls. They also secreted less murine (mu)TNF when stimulated by either bacterial lipopolysaccharide or toxic malarial antigens. The yields of muTNF approached normal levels, however, when these refractory cells from the transgenic mice were stimulated in the presence of a high concentration of indomethacin, suggesting that the production of muTNF was inhibited by enhanced synthesis of prostaglandins. The parasitaemia of transgenic mice infected with Plasmodium yoelii was about 10-fold less at its peak than in controls, although it followed the same time-course, and the multiplication of P. chabaudi was inhibited to an even greater degree. This control of parasitaemia may also be explained by enhancement of macrophage activity, mediated by huTNF acting on the murine p55 receptor, presumably by increasing the removal of parasites by phagocytosis or their killing by toxic products released by the activated macrophages. These observations suggest that a factor in the anaemia of human malaria may be macrophage activation caused by the secretion of TNF that occurs in this disease.

Anemia↗
PubMed1993

Wasting, ischemia, and lymphoid abnormalities in mice expressing T cell-targeted human tumor necrosis factor transgenes.

To evaluate the biologic potential of T cell-specific TNF production in vivo, we have generated transgenic mice constitutively expressing TNF in their T cell compartment. This was achieved by placing a wild-type or a 3'-UTR modified fragment of the human TNF gene under the influence of the T cell-specific, locus control region of the human CD2 gene. Transgenic mice that express human TNF mRNA in T cells develop marked histologic and cellular changes locally in their lymphoid organs and a lethal wasting syndrome associated with widespread vascular thrombosis and tissue necrosis. The extent of pathologic changes and their time of onset appear to reflect levels of transgene expression. Thus, transgenic lines that express the transgene at high levels show both lymphoid organ and systemic abnormalities with wasting. In one transgenic line, mice express lower levels of the transgene and develop normally despite pronounced local lymphoid organ defects, confirming in vivo, the differential potential of localized and systemic TNF action. All pathologic changes could be neutralized by the administration of mAb specific for human TNF. These results demonstrate the important role of T cell-specific TNF production in the development of specific pathology and provide a means by which to evaluate the role of TNF in thymocyte development. Transgenic mice that express TNF constitutively in the T cell compartment offer a unique in vivo system by which to analyze the molecular character of systemic vs contact-dependent and paracrine modes of TNF action. Furthermore, given the species-specific nature of the mouse p75 TNF receptor, it is assumed that the pathology induced by human TNF in these transgenic mice is associated exclusively with p55 TNF receptor signaling. Conceivably, the differential contribution of each of the two TNF receptors in thymus development and TNF-mediated disease can be assessed by comparison of the biologic potential of human vs mouse TNF in the transgenic system developed.

Animals↗
PubMed1993

The synovium of transgenic arthritic mice expressing human tumor necrosis factor contains a high level of nerve growth factor.

We have recently reported that nerve growth factor (NGF) increases in the synovium of patients affected by rheumatoid arthritis and in the synovium of pharmacologically-induced arthritis in animal models. In the present study, we demonstrate that arthritic transgenic mice which carry and express the human TNF gene (Tg197) also express elevated levels of NGF, and that subcutaneous injection of NGF-antibodies attenuates the loss of body weight caused by the development of disease in these mice. Along with our previous findings, which show an increase in the level of NGF during the acute phase of other autoimmune diseases, these results suggest a role of NGF in these pathologies. The functional significance of NGF in rheumatoid arthritis (RA) is currently under study.

Animals↗
PubMed1993

Level of nerve growth factor and distribution of mast cells in the synovium of tumour necrosis factor transgenic arthritic mice.

We have recently reported that nerve growth factor (NGF) increases in the synovium of patients affected by arthritis, as well as in animal models. We report here that the synovium of transgenic arthritic mice expressing human tumour necrosis factor (TNF) contains numerous mast cells (MC) and that their appearance is a phenomenon which was correlated to the local increase in NGF level. These findings provide further evidence that NGF plays a role in inflammation and suggest a functional link between NGF and MC.

Animals↗
PubMed1991

Transgenic mice expressing human tumour necrosis factor: a predictive genetic model of arthritis.

We have generated transgenic mouse lines carrying and expressing wild-type and 3'-modified human tumour necrosis factor (hTNF-alpha, cachectin) transgenes. We show that correct, endotoxin-responsive and macrophage-specific hTNF gene expression can be established in transgenic mice and we present evidence that the 3'-region of the hTNF gene may be involved in macrophage-specific transcription. Transgenic mice carrying 3'-modified hTNF transgenes shows deregulated patterns of expression and interestingly develop chronic inflammatory polyarthritis. Treatment of these arthritic mice with a monoclonal antibody against human TNF completely prevents development of this disease. Our results indicate a direct involvement of TNF in the pathogenesis of arthritis. Transgenic mice which predictably develop arthritis represent a novel genetic model by which the pathogenesis and treatment of this disease in humans may be further investigated.

Animals↗
PubMed1987

The immunocytochemical localisation of 'substance-P-degrading enzyme' within the rat spinal cord.

The regional distribution of a neural endopeptidase ('substance-P-degrading enzyme', SPDE) within the rat spinal cord, as visualised by immunocytochemistry, closely paralleled the distribution of major substance P-containing nerve fibre systems. Thus, SPDE-immunoreactivity was present in laminae I and II of the dorsal horn, in large diameter neurones of the ventral horn and in the ependymal cells which line the central canal. The pattern of immunostaining, particularly in the dorsal horn, is consistent with the view that SPDE may function in the metabolism of substance P.

Animals↗
PubMed1984

The regulatory peptide system of the large bowel in equine grass sickness.

In recent years, distinct changes in regulatory peptides have been found in a number of gastrointestinal diseases. Grass sickness is a fatal disease of horses for which the etiology has yet to be fully ascertained. In this study, the peptide-containing nerves and ganglionic and mucosal endocrine cells of the ileum, colon and rectum were investigated in horses with sub-acute or chronic grass sickness and compared with normal controls using immunocytochemistry, at both the light and electron microscopical levels, and radioimmunoassay. A substantial loss of both peptide-containing cells and nerves was found in all of the sick horses, particularly in the ileum. Electron microscopy revealed marked degeneration of nerves in the gut wall. Fibers containing granules immunostained for substance P or VIP, using the immunogold staining technique, underwent extensive degranulation in grass sickness, with the formation of multiple vacuoles. Radioimmunoassay of peptide content also showed that the most drastic changes occurred in the ileum. For example, VIP content was significantly reduced from 109 +/- 19.8 (mean +/- SEM) pmoles/g in controls to 6.8 +/- 1.4 pmoles/g in grass sickness (p less than 0.001) and substance P from 65.9 +/- 8.1 to 31.3 +/- 9.5 (p less than 0.02). These results may have applications in the diagnosis and treatment of grass sickness.

Animals↗
PubMed1983

Peptidergic innervation of the human male genital tract.

Four peptides--vasoactive intestinal polypeptide, substance P, somatostatin and a peptide-like avian pancreatic polypeptide--have been found in nerves of the human male genitalia using highly sensitive and specific methods of immunocytochemistry and radioimmunoassay. Five other peptides (met-enkephalin, leu-enkephalin, neurotensin, bombesin and cholecystokinin-8) were absent. Vasoactive intestinal polypeptide was the most abundant peptide, its highest concentration being in the proximal corpus cavernosum. Immunoelectron microscopy localized this peptide to large (97 +/- 20 nm), round, electron-dense granules of p-type nerve terminals. Vasoactive intestinal polypeptide-immunoreactive neuronal cell bodies were found in the prostate gland and the root of the corpus cavernosum. Substance P immunoreactive material was present in smaller concentration and was mainly localized in nerves around the corpuscular receptors of the glans penis. Somatostatin immunoreactive nerves were associated mainly with the smooth muscle of the seminal vesicle and the vas deferens. When antiserum to avian pancreatic polypeptide was applied, certain nerves were stained, particularly in the vas deferens, the prostate gland and the seminal vesicle. However, chromatography detected no pure avian pancreatic polypeptide suggesting the presence of a structurally related substance, possibly neuropeptide Y, which cross-reacts with the avian pancreatic polypeptide antiserum. Similar distributions between vasoactive intestinal polypeptide-immunoreactive and acetylcholinesterase-positive nerves and between avian pancreatic polypeptide-immunoreactive and adrenergic nerves were observed. A general neuronal marker, neuron-specific enolase, was used to investigate the general pattern of the organ's innervation. The abundance and distribution patterns of these peptide-immunoreactive nerves indicate that they may play important roles in the male sexual physiology.

Adult↗
PubMed1983

Double immunogold staining method for the simultaneous ultrastructural localization of regulatory peptides.

Recent studies have suggested that the morphological characteristics of secretory granules contained within endocrine cells and nerves may be determined largely by their chemical composition. The use of the immunogold staining (IGS) method, which is based on the adsorption of colloidal gold to immunoglobulins, has been used in our laboratory to demonstrate a wide range of intracellular antigens at both the light and electron microscope levels. In this study we have applied a modification of the IGS method for the simultaneous detection of two separate antigens in a single tissue section, using a variety of region-specific antisera to different peptides. Peptide antisera were raised in rabbits or in guinea pigs and these were applied simultaneously or sequentially to grid-mounted ultrathin tissue sections. Antigenic sites were visualized at the electron microscope level using antisera raised in goats, adsorbed to gold particles of 12, 20, or 40 nm. Using this technique we have attempted to investigate the coexistence of multiple antigens in single tissue sections, in particular in single granules; the topographic distribution of molecular forms within one single granule or granule population; the heterogeneity of peptidergic neurons and also the heterogeneity of peptide content in morphologically similar granules. The double immunogold staining procedures described here have proved to be extremely effective for the simultaneous ultrastructural localization of two antigens (peptide-peptide; peptide-propeptide) on a single tissue section. The further development of this technique may provide useful information on neuroendocrine cell dynamics in normal and diseased states.

Animals↗