Postischemic infusion of Cu/Zn superoxide dismutase or SOD:Tet451 reduces cerebral infarction following focal ischemia/reperfusion in rats

Oxygen-free radicals play a major role in neuronal cell injury following cerebral ischemia/reperfusion. The free-radical scavenging enzyme, Cu/Zn superoxide dismutase (SOD-1), ameliorates various types of brain injury resulting from temporary CNS ischemia. We have compared the cerebroprotective properties of human SOD-1 (hSOD-1) with a novel recombinant SOD-1 hybrid protein, SOD:Tet451, composed of hSOD-1 linked to the neuronal binding fragment of tetanus toxin (TTxC). Following 2 h of temporary middle cerebral artery occlusion, rats infused with equivalent activities of either hSOD-1 or SOD:Tet451 for the initial 3 h of reperfusion showed reductions in cerebral infarct volume of 43 and 57%, respectively, compared to saline-treated controls (P < 0.01). Serum hSOD-1 concentrations in rats receiving SOD:Tet451 were seven-fold higher than those in rats receiving the native enzyme. Animals treated with SOD:Tet451 also demonstrated an extended persistence of hSOD-1 in the bloodstream during drug washout as compared to animals given free enzyme. Immunohistochemical examination of brain sections from an SOD:Tet451-treated ischemic rat showed positive immunoreactivity in the ipsilateral cerebral cortex using either anti-TTxC or anti-human SOD-1 antibodies. Our results document that both hSOD-1 and SOD:Tet451 significantly reduce brain infarct volume in a model of transient focal ischemia/reperfusion in rats. Additionally, our findings suggest that the cerebroprotective effects of SOD-1 may be enhanced by neuronal targeting as seen with the hybrid protein SOD:Tet451.     

A role of Hsp60 in autoimmune diabetes: analysis in a transgenic model

A pathogenic role for self-reactive cells against the stress protein Hsp60 has been proposed as one of the events leading to autoimmune destruction of pancreatic beta cells in the diabetes of nonobese diabetic (NOD) mice. To examine this hypothesis, we generated transgenic NOD mice carrying a murine Hsp60 transgene driven by the H-2E alpha class II promoter. This would be expected to direct expression of the transgene to antigen-presenting cells including those in the thymus and so induce immunological tolerance by deletion. Detailed analysis of Hsp60 expression revealed that the endogenous gene is itself expressed strongly in thymic medullary epithelium (and weakly in cortex) yet fails to induce tolerance. Transgenic mice with retargeted Hsp60 showed overexpression of the gene in thymic cortical epithelium and in bone marrow-derived cells. Analysis of spontaneous T-cell responses to a panel of self and heterologous Hsp60 antigens showed that tolerance to the protein had not been induced, although responses to an immunodominant 437-460 epitope implicated in disease were suppressed, probably indicating an epitope shift. This correlated with changes in disease susceptibility: insulitis in transgenic mice was substantially reduced so that pathology rarely progressed beyond periislet infiltration. This was reflected in a substantial reduction in hyperglycemia and disease. These data indicate that T cells specific for some epitopes of murine Hsp60 are likely to be involved in the islet-cell destruction that occurs in NOD mice.     

Expression of cyclin D1 in epithelial tissues of transgenic mice results in epidermal hyperproliferation and severe thymic hyperplasia

To study the involvement of cyclin D1 in epithelial growth and differentiation and its putative role as an oncogene in skin, transgenic mice were developed carrying the human cyclin D1 gene driven by a bovine keratin 5 promoter. As expected, all squamous epithelia including skin, oral mucosa, trachea, vaginal epithelium, and the epithelial compartment of the thymus expressed aberrant levels of cyclin D1. The rate of epidermal proliferation increased dramatically in transgenic mice, which also showed basal cell hyperplasia. However, epidermal differentiation was unaffected, as shown by normal growth arrest of newborn primary keratinocytes in response to high extracellular calcium. Moreover, an unexpected phenotype was observed in the thymus. Transgenic mice developed a severe thymic hyperplasia that caused premature death due to cardio-respiratory failure within 4 months of age. By 14 weeks, the thymi of transgenic mice increased in weight up to 40-fold, representing 10% of total body weight. The hyperplastic thymi had normal histology revealing a well-differentiated cortex and medulla, which supported an apparently normal T-cell developmental program based on the distribution of thymocyte subsets. These results suggest that proliferation and differentiation of epithelial cells are under independent genetic controls in these organs and that cyclin D1 can modulate epithelial proliferation without altering the initiation of differentiation programs. No spontaneous development of epithelial tumors or thymic lymphomas was perceived in transgenic mice during their first 8 months of life, although they continue under observation. This model provides in vivo evidence of the action of cyclin D1 as a pure mediator of proliferation in epithelial cells.     

The action of Bax and bcl-2 on T cell selection

T cell development and selection in the thymus are shaped by the induction of apoptosis. However, a direct role in T cell development and selection for any of the molecules known to regulate apoptosis has remained controversial. We have studied the effect of bax and bcl-2 transgenes in recombination activation gene 1-deficient (RAG-1(-/-)) mice transgenic for the major histocompatibility complex class I-restricted F5 T cell receptor. Overexpression of a bax transgene in the thymus seriously impairs the production of mature T cells, whereas bcl-2 overexpression greatly promotes it. The effect of bax and bcl-2 overexpression on antigen-induced negative selection was studied using fetal thymic organ cultures. This analysis showed that Bcl-2 strongly inhibits negative selection, whereas Bax does not affect it. Our data directly show that Bcl-2 family members have specific roles in T cell selection and also lend support to the hypothesis that Bax and Bcl-2 can antagonize each other's action in a certain apoptosis pathway while in another they can be functionally nonreciprocal.     

Transgenic mice and knockout mutants in the study of oxidative stress in brain injury

A rapid increase in the need to explore the molecular basis of cellular function and injury in the central nervous system has led neuroscientists to employ transgenic mouse technology. The successful making of transgenic mice (Tg) overexpressing human CuZn-superoxide dismutase (SOD-1) activity has made it possible to investigate the role of oxygen free radicals in ischemic and traumatic brain injury in a molecular fashion. It has been demonstrated that the 3-fold increase in SOD-1 transgene activity in SOD-1 Tg mice offers protection against cerebral ischemia and reperfusion in two different models of focal cerebral ischemia, as compared to nontransgenic wild-type littermates. Studies involving traumatic brain injury have also demonstrated that acute injuries, including brain edema and blood-brain barrier permeability, are significantly reduced in SOD-1 Tg mice. Furthermore, chronic neurological deficits, such as beam walking, beam balance, and body weight, are significantly improved in these transgenic animals following traumatic brain injury. In addition to the SOD-1 Tg mice being a useful tool for the study of CNS injury, targeted disruption of the mouse gene for mitochondrial manganese SOD (SOD-2) has been successful. These SOD-2 knockout mutant mice, in addition to the recently developed knockout mutants of neuronal nitric oxide synthase (NOS), are believed to offer a unique opportunity to elucidate the oxidative mechanisms in brain injury following stroke and trauma.     

T-maze spontaneous alternation rate is decreased in S100!b transgenic mice.

S100β, a calcium-binding brain specific protein, may affect both brain development and hippocampal long-term potentiation. S100β levels are elevated in Down's syndrome (DS), and the gene for S100β is located on chromosome 21, which is duplicated in DS. To test the hypothesis that elevated levels of S100β cause behavioral alterations in a mammalian system, 3 transgenic mouse lines with multiple copies of the human gene for S100β were derived and behaviorally tested. The spontaneous alternation behavior of transgenic and normal littermate mice was compared in a T-maze during a 15-trial test. The overall alternation rate was found to be significantly decreased in the transgenic mice compared with their normal littermates. The S100β transgenic mouse model offers one of the first opportunities to investigate the relation between overexpression of a human chromosome 21 gene product and abnormal behavior and brain function. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Zinc and metallothioneins on cellular immune effectiveness during liver regeneration in young and old mice

Partial hepatectomy in young mice (pHx) induces thymic atrophy, disregulation of thymocytes subsets and a strong accumulation of zinc in thymic tissue after 1-2 days of liver regeneration. Zinc is relevant for good immune functioning. Restoration of zinc into both the thymus and thymocytes subsets in the late period of liver regeneration is observed in young pHx mice. These findings have suggested a link between the thymus and the liver influencing T-cell functions and involving zinc. This kind of link could be relevant in aging because thymic involution, negative crude zinc balance and crippled immune functions are constant events. The preminence of a liver extrathymic T-cell pathway after pHx or during aging has been suggested. Thus the study of pHx in young and old mice may offer a good model to better understand the role played both by thymic involution and by liver extrathymic T-cell pathway and the role of zinc in these physiological processes during aging. Young pHx mice after 1-2 days of liver regeneration show: reduced thymic endocrine activity, increment of double negative (DN) thymocytes subsets, impairment of peripheral immune efficiency (PHA, NK activity and IL-2) and negative crude zinc balance, which are all restored in the late period of liver regeneration. By contrast the thymic and peripheral immune defects and the negative crude zinc balance, already present in old sham mice, are not modified during liver regeneration in old pHx mice. Circulating leukocytes and lymphocytes are not significantly modified both in young and old pHx mice as compared to respective sham controls. Zinc may also be crucial for extrathymic T-cells pathway, being preminent in aging, rather than in young age, due to its metallothioneins (MT) binding capacity. MT are significantly increased in young pHx and in aging inducing a low zinc-free quota for thymic and peripheral immune efficiency in young pHx mice, and for extrathymic T-cell pathway, in old age. Thus low zinc bioavailability, due to MT, may play a pivotal role, not only for thymocytes but also for liver extrathymic T-cell pathway.     

Results of the Wilke operation to stop drooling in cerebral palsy

Chronic treatment with lithium chloride produced significant involution of the thymus gland with histological evidence of reduced cellularity due to loss of thymic lymphocytes and a significant reduction in the weight of the gland in normal and adrenalectomized mice. Lithium also increased corticosterone levels in normal mice without changes in adrenal weights. The involution of the thymus gland is most likely due to an effect of lithium on the gland, and it is not mediated by adrenocortical mechanisms or stress.     

Cross-clade immune responses after immunization with a whole-killed gp120-depleted human immunodeficiency virus type-1 immunogen in incomplete Freund''s adjuvant (HIV-1 immunogen, REMUNE) in human immunodeficiency virus type-1 seropositive subjects

Down syndrome (DS) is associated with mental retardation, immune disorders and congenital heart diseases. Although it is usually caused by the presence of an extra chromosome 21, a subset of the diagnostic phenotypic features may be caused by the presence of the band 21q22, called the "Down syndrome region". Many proteins important for the immune and nervous systems as CuZn-superoxide dismutase (SOD-1), CD18-beta chain of LFA-1, interferon receptor, APP-amyloid precursor protein, protein S-100 beta are coded by chromosome 21. Overexpression of these molecules may contribute to the thymic derangement that results in anomalous maturation leading to functionally impaired T cells. Many factors have been shown to contribute to the immune deficiency which results in high susceptibility to infections, high rate of malignancies, and autoimmune phenomena in persons with DS. The main disorders in the immune system include thymus abnormalities, changes in cell-mediated immunity, phagocytosis, antibodies-mediated immunity and a high prevalence of autoantibodies in persons with DS. Furthermore, the duplication of chromosome 21 genes may generate most of the pathological changes in the central nervous system. There is an increased prevalence of seizure disorders. Such widespread alterations in the cortical areas seem to account for specific impairments observed in short-term and long-term memory, language skills, and cognitive and learning processes. If all principles of optimal health care and adequate education were followed without exception for persons with DS, then the quality of their life could be improved significantly and they would be able to become productive citizens in the society. (Tab. 5, Fig. 3, Ref. 42.)     

Tamoxifen exerts testosterone-dependent and independent effects on thymic involution

Circulating testosterone concentrations and seminal vesicles weights, as well as thymus and spleen weights and histology were assessed in male Wistar rats from the infantile to post-pubertal period. The widely used anti-estrogenic agent tamoxifen was then administered in adult intact and castrated male rats and its long-term effects on thymic involution and splenic growth were examined. The results showed that: (1) age-related involution of the male thymus from the juvenile period through puberty to post-puberty depends on the rising testosterone levels and represents mainly a decrease of thymic lymphoid-cell elements; (2) tamoxifen administration reverses thymic involution in intact adult male rats and this effect is related to a dose-dependent, tamoxifen-induced castration and decrease of testosterone levels; (3) the changes of circulating testosterone levels, either resulting from maturity, or induced by tamoxifen or by castration, have a minimal effect on splenic growth and weight; and (4) in contrast to intact animals, administration of tamoxifen at pharmacological doses to adult castrated rats results in thymic regression. Underscoring the critical role of testosterone on thymic involution, these findings show that tamoxifen is able to reverse ageing changes in the thymus by suppressing testosterone production, while conversely, exerts thymolytic effects in the absence of androgens.     

Influence of detergents on the function of cloned potassium channels

The human thymus is a lymphoepithelial organ in which T cells develop during fetal life. After maturation and selection in the fetal thymic microenvironment, T cells emigrate to peripheral lymphoid tissues such as the spleen, gut, and lymph nodes, and establish the peripheral T cell repertoire. Although the thymus has enormous regenerative capacity during fetal development, the regenerative capacity of the human postnatal thymus decreases over time. With the advent of intensive chemotherapy regimens for a variety of cancer syndromes, and the discovery that infection with the Human Immunodeficiency Virus (HIV) leads to severe loss of CD4+ T cells, has come the need to understand the role of the human thymus in reconstitution of the immune system in adults. During a recent study of the thymus in HIV infection, we observed many CD8+ T cells in AIDS thymuses that had markers consistent with those of mature effector cytotoxic T cells usually found in peripheral immune tissues, and noted these CD8+ effector T cells were predominately located in a thymic zone termed the thymic perivascular space. This article reviews our own work on the thymus in HIV-1 infection, and discusses the work of others that, taken together, suggest that the thymus contains peripheral immune cell components not only in the setting of HIV infection, but also in myasthenia gravis, as well as throughout normal life during the process of thymus involution. Thus, the human thymus can be thought of as a chimeric organ comprised of both central and peripheral lymphoid tissues. These observations have led us to postulate that the thymic epithelial atrophy and decrease in thymopoiesis that occurs in myasthenia gravis, HIV-1 infection, and thymic involution may in part derive from cytokines or other factors produced by peripheral immune cells within the thymic perivascular space.     

Exhaustive scanning approach to screen all the mitochondrial tRNA genes for mutations and its application to the investigation of 35 independent patients with mitochondrial disorders

There have been several reports that thymoma in human is a progressive disease, and that thymoma and thymic carcinoma form a continuum. We established a stable line of SV40T transgenic mice, which consistently produced thymic epithelial tumours progressing to thymic carcinoma within a predictable time span. Using this animal model and a morphological approach, thymic epithelial tumour progression was studied with reference to sequential changes at different time points in animals aged from 3 to 32 weeks. At all ages, SV40T was expressed in the nuclei of thymic epithelial cells; in these transgenic mice we observed the entire spectrum from cortical type thymoma to thymic carcinoma. Thymic size tended to increase with ageing in SV40T TG mice. While younger mice had predominantly cortical (organoid) or cortical thymoma, older mice had well-differentiated thymic carcinoma (WDTC) or poorly differentiated thymic carcinoma. When SV40T TG mice (248 line) reached a certain age, carcinoma of the thymus was present in all of them. Cortical-type thymoma became malignant within a predictable time span, suggesting a cortical thymoma-carcinoma sequence. When the mice were 9 weeks of age, the thymuses formed gross masses compatible with cortical thymoma. At 14 weeks of age, WDTC appeared against the background of cortical thymoma. Poorly differentiated thymic carcinoma was found after 15 weeks and affected all animals over 23 weeks of age. Most thymic carcinomas coexisted in varying proportions with cortical-type thymoma. Medullary thymomas did not develop in the mice, and no transition from medullary-type thymomas to thymic carcinomas was observed. In this SV40T transgenic mouse model, thymic carcinoma is clearly preceded by cortical-type thymoma. These transgenic mice may provide an interesting model for the progression from cortical thymoma to WDTC and/or high-grade carcinoma.     

Case report: non-extraction treatment with functional and mechanical therapy of a Class II division 1 malocclusion

The cardinal sign of acute stress is thymic involution, which subsequently attenuates the activity of immunocompetent cells, notably T-lymphocytes, macrophages and NK cells. Sodium diethyldithiocarbamate (DTC), a low molecular weight sulphur compound, may function as a thymic hormone to induce precursor cells to become functionally mature T-lymphocytes. The studies were conducted on Balb/c mice exposed to restraint stress twice for 12 h at 24 h intervals. DTC at a dose of 20 mg/kg or calf thymus extract (TFX) at a dose of 10 mg/kg were injected i.p. four times at 24 h intervals prior to the exposure. It has been found that restraint stress markedly reduces the number of thymocytes which is concomitant with reduction in the weight of the thymus. In our study the changes sustained for 10 days of the observation. Besides, alterations in proliferative response of the thymocytes stimulated in vitro with concanavalin A (Con A) and phytohemagglutinin (PHA) were observed. The proliferative response of thymocytes to Con A was reduced 24 h after the exposure to restraint stress, but between days 4 and 7 it was found at increased levels, which decreased again on day 10. In contrast, the proliferative response of thymocytes to PHA was depressed for the entire 10 day period of the observation. It has been found that DTC and TFX administered to mice prior to restraint stress successfully counteract stress-induced immunosuppression, albeit TFX exerts stronger protective and regenerating impact on the thymus than DTC. TFX totally inhibits the suppressive effect of stress on proliferative activity of the thymocytes stimulated in vitro with Con A and PHA, stimulates restoration of thymic cells and increases the weight of the thymus. In contrast, DTC is not able to counteract the decrease in proliferative response of thymocytes to PHA.     

Targeted overexpression of phospholamban to mouse atrium depresses Ca2+ transport and contractility

Phospholamban is a small phosphoprotein regulator of the Ca2+-pump of cardiac sarcoplasmic reticulum. Dephosphorylated phospholamban inhibits the Ca2+-pump and depresses contractility, whereas phosphorylation of phospholamban by cAMP-activated mechanisms relieves this inhibition and increases contractility. In order to better understand the function of phospholamban in living systems, a transgenic mouse model was established employing targeted overexpression of phospholamban to the atrium, which normally expresses low levels of the protein. Overexpression was achieved by fusing the alpha-MHC-promoter or the ANF-promoter to the phospholamban gene. Double transgenic mice were created by mating mice positive for each transgene. In single transgenic lineages, phospholamban was overexpressed four to six-fold in left atrium. In the double transgenic mice, phospholamban was overexpressed eight- to nine-fold. In the three transgenic strains. Ca2+ uptake by the sarcoplasmic reticulum was depressed to 22-30% of control values at low ionized calcium. This depression of Ca2+ uptake was largely reversed by addition of a phospholamban monoclonal antibody. In the atrial muscle strips, the time course of contraction was increased in a concentration-dependent manner by overexpression of phospholamban, whereas the basal developed tension was decreased up to 85% by phospholamban-overexpression. In all transgenic lineages, isoproterenol, a beta-adrenoceptor agonist, reversed the depression of contractility caused by overexpression of phospholamban and significantly shortened time parameters to levels approaching control values. These data demonstrate that overexpression of phospholamban in a mammalian myocardial tissue normally deficient in the protein substantially inhibits basal contractility, and furthermore suggest that in myocardial tissues containing high levels of the protein, phosphorylation of phospholamban can account for many of the positive inotropic and lusitropic effects of beta-adrenergic stimulation.     

Thymic alterations in feline GM1 gangliosidosis

GM1 gangliosidosis is an inherited metabolic disease characterized by progressive neurological deterioration with premature death seen in children and numerous animals, including cats. We have observed that thymuses from affected cats greater than seven months of age (GM1 mutant cats) show marked thymic reduction compared to age-matched normal cats. The studies reported here were done to describe alterations in the thymus prior to (less then 90 days of age) and during the development of mild (90 to 210 days of age) to severe (greater than 210 days of age) progressive neurologic disease and to explore the pathogenesis of the thymic abnormality. Although histologic examination of the thymus from GM1 affected cats less than 210 days of age showed no significant differences from age-matched control cats, thymuses from GM1 mutant cats greater than 210 days of age were significantly reduced in size (approximately 3-fold). Histologic sections of lymph nodes, adrenal glands, and spleens from GM1 gangliosidosis-affected cats showed no significant differences. Flow cytometric analyses showed a marked decrease in the percentage of immature CD4+CD8+ thymocytes (p < 0.001) and significantly increased CD4-CD8+ cells (p < 0.01) in GM1 mutant cats greater than 210 days of age when compared to normal age matched cats. Co-labelling with CD4, CD8, and CD5 indicated an increase in the percentage of GM1 mutant cat thymocytes at this age which were CD5high, suggesting the presence of more mature cells. Cytometric analyses of subpopulations of peripheral lymphocytes indicated an increase in CD4-CD8+ cells (p < 0.05) with concurrent decreases in CD4+CD8- and CD4-CD8- cells (which were not significant). Similar analyses of thymocyte and lymphocyte subpopulations from cats < 210 days of age showed no significant differences between GM1 mutant and normal cells. GM1 mutant cats at all ages had increased surface binding of Cholera toxin B on thymocytes, indicating increased surface GM1 ganglioside expression. Increases were highly significant in GM1 mutant cats greater than 210 days of age. In situ labelling for apoptosis was increased in GM1 mutant cats between 90 to 200 days of age when thymic masses were within normal limits. In GM1 mutant cats over 200 days of age, decreased labelling was observed when thymic mass was reduced and the CD4+CD8+ subpopulation, known to be very susceptible to apoptosis, was significantly decreased. These data describe premature thymic involution in feline GM1 gangliosidosis and suggest that increased surface GM1 gangliosides alters thymocyte development in these cats.     

The GTPase Rho has a critical regulatory role in thymus development

The present study employs a genetic approach to explore the role of Rho GTPases in murine thymic development. Inactivation of Rho function in the thymus was achieved by thymic targeting of a transgene encoding C3 transferase from Clostridium botulinum which selectively ADP-ribosylates Rho within its effector domain and thereby abolishes its biological function. Thymi lacking functional Rho isolated from C3 transgenic mice were strikingly smaller and showed a marked (90%) decrease in cellularity compared with their normal litter mates. We also observed a similar decrease in levels of peripheral T cells in C3 transgenic mice. Analysis of the maturation status of thymocytes indicated that differentiation of progenitor cells to mature T cells can occur in the absence of Rho function, and both positive and negative selection of T cells appear to be intact. However, transgenic mice that lack Rho function in the thymus show maturational, proliferative and cell survival defects during T-cell development that severely impair the generation of normal numbers of thymocytes and mature peripheral T cells. The present study thus identifies a role for Rho-dependent signalling pathways in thymocyte development. The data show that the function of Rho GTPases is critical for the proliferative expansion of thymocytes. This defines a selective role for the GTPase Rho in early thymic development as a critical integrator of proliferation and cell survival signals.     

Altered brain glucose metabolism in transgenic-PFKL mice with elevated L-phosphofructokinase: in vivo NMR studies

The gene for the liver-type subunit of phosphofructokinase (PFKL) resides on chromosome 21 and is overexpressed in Down syndrome (DS) patients. Transgenic PFKL (Tg-PFKL) mice with elevated levels of PFKL were used to determine whether, as in DS, overexpression of PFKL was also associated with altered sugar metabolism. We found that Tg-PFKL mice had an abnormal glucose metabolism with reduced clearance rate from blood and enhanced metabolic rate in brain. Transgenic-PFKL mice exhibited elevated activity of phosphofructokinase in both blood and brain, as compared to control non-transgenic (ntg) mice. Following glucose infusion, the rate of glucose clearance from the blood of Tg-PFKL mice was significantly slower than that of control ntg mice, although the basal blood glucose levels were similar. However, unlike the slower rate of glucose metabolism in blood, the initial rate of glucose utilization in the brain of the transgenic mice, was 58% faster than in control ntg mice. This was determined by infusion of [1-13C]-glucose followed by in vivo nuclear magnetic resonance (NMR) measurements of brain glucose metabolism. The faster utilization of glucose in Tg-PFKL brain is similar to the increased rate of cerebral glucose metabolism found in the brain of young adult DS patients, which may play a role in the etiology of their cognitive disabilities.     

Up-regulation of protein chaperones preserves viability of cells expressing toxic Cu/Zn-superoxide dismutase mutants associated with amyotrophic lateral sclerosis

Mutations in the Cu/Zn-superoxide dismutase (SOD-1) gene underlie some familial cases of amyotrophic lateral sclerosis, a neurodegenerative disorder characterized by loss of cortical, brainstem, and spinal motor neurons. We present evidence that SOD-1 mutants alter the activity of molecular chaperones that aid in proper protein folding and targeting of abnormal proteins for degradation. In a cultured cell line (NIH 3T3), resistance to mutant SOD-1 toxicity correlated with increased overall chaperoning activity (measured by the ability of cytosolic extracts to prevent heat denaturation of catalase) as well as with up-regulation of individual chaperones/stress proteins. In transgenic mice expressing human SOD-1 with the G93A mutation, chaperoning activity was decreased in lumbar spinal cord but increased or unchanged in clinically unaffected tissues. Increasing the level of the stress-inducible chaperone 70-kDa heat shock protein by gene transfer reduced formation of mutant SOD-containing proteinaceous aggregates in cultured primary motor neurons expressing G93A SOD-1 and prolonged their survival. We propose that insufficiency of molecular chaperones may be directly involved in loss of motor neurons in this disease.