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Ilya Frolov - One of the best experts on this subject based on the ideXlab platform.
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analysis of venezuelan equine encephalitis virus capsid protein function in the inhibition of cellular transcription
Journal of Virology, 2007Co-Authors: Natalia Garmashova, Svetlana Atasheva, Wenli Kang, Scott C Weaver, Elena I Frolova, Ilya FrolovAbstract:The genus Alphavirus in the family Togaviridae includes a number of important human and animal pathogens (15). Alphaviruses are currently classified into six antigenic complexes and are widely distributed in both the New World and the Old World. They are efficiently transmitted by mosquitoes, in which they cause a persistent, lifelong infection with a minimal effect on biological functions. In vertebrates, alphavirus infection is acute and in many cases characterized by high-titer viremia, rash, fever, and encephalitis until the death of the infected host or clearance of the virus by the immune system. The encephalitogenic alphaviruses, including Venezuelan (VEEV), eastern (EEEV), and western (WEEV) equine encephalitis viruses, represent a continuous public health threat in the United States (41, 48-50). They circulate in Central, South, and North America and have the ability to cause fatal disease in humans and in horses and other domestic animals. During VEEV epizootics, equine mortality can reach 83%, and in humans, the virus produces a severe temporary immunodeficiency and a greatly debilitating, sometimes fatal disease (42). The overall mortality rate is below 1%, but neurological disease, including disorientation, ataxia, Mental Depression, and convulsions, can be detected in up to 14% of all infected individuals, especially children (23). Also described are sequelae of VEEV-related clinical encephalitis in humans (10, 27). The VEEV genome is represented by a single-stranded RNA molecule of positive polarity that is almost 12 kb in length. It mimics the structure of cellular mRNAs, with a cap at the 5′ terminus and a poly(A) tail at the 3′ end of the RNA. The VEEV genome has been cloned in a cDNA form (24) that allows a wide variety of genetic manipulations to be undertaken. The only experiMental vaccine against VEEV infection that has been used extensively in humans was developed 4 decades ago by serial passaging of the virulent subtype IAB Trinidad Donkey VEEV strain in guinea pig heart cell cultures (3). Over 8,000 humans have been vaccinated during the past 4 decades (2, 6, 37), and the cumulative data indicate that nearly 40% of vaccinated people develop a disease with some symptoms typical of those seen with natural VEEV infection, including a febrile systemic illness and other adverse effects (2, 3, 21). No effective antivirals have been developed against any alphavirus, including VEEV. In spite of the continuous threat of VEEV epidemics, the biology of this virus has been studied less intensively than those of other, less pathogenic alphaviruses, such as Sindbis (SINV) and Semliki Forest (SFV) viruses. These viruses can be readily manipulated in a low-biocontainment environment and represent good models for studying the mechanisms of alphavirus replication, virus-host interactions, and encephalitis development (14). However, important differences in pathogenesis and the severity of human and veterinary diseases suggest that these viruses may not be ideal models for encephalitis. Moreover, the results from recent studies of the Old World (SINV and SFV) and the New World (VEEV and EEEV) alphaviruses (1, 9, 11-13, 36, 46) demonstrated that both of these groups have developed the ability to interfere with cellular transcription and to use this effect as a means of downregulating cellular antiviral response. However, the mechanisms of transcription inhibition appear to be fundaMentally different; while the Old World alphaviruses use nsP2 to inhibit cellular transcription (11), the more encephalitogenic VEEV and EEEV use their capsid proteins for the same function (1, 12). Expression of the latter proteins by different vectors is sufficient for induction of cell death and cytopathic effects (CPE) in tissue culture, and the development of these phenomena strongly correlates with the inhibition of transcription of cellular mRNA and rRNA. Moreover, the replacement of the structural protein genes in the VEEV genome by those derived from SINV has made the chimeric virus significantly less cytopathic and incapable of interfering with the development of an antiviral response in cells having no defect in alpha/beta interferon induction and signaling (12). In this study, we continued our investigation of the VEEV capsid protein-dependent inhibition of cellular transcription. Our data demonstrate that (i) transcriptional shutoff and CPE development in capsid protein-expressing cells are determined by a short CVEE peptide, CVEE33-68, located in the N-terminal part of the protein; (ii) inhibition of transcription and CPE development correlate with the presence of the entire capsid protein or the indicated peptide on the nuclear membrane, which suggests their interaction with the nuclear pore complex (NPC); (iii) the mutations in the peptide might lead to the accumulation of capsid protein in either the cytoplasm or the nucleus; and (iv) VEEV variants encoding either the chimeric capsid protein with the amino-terminal fragment (amino acids [aa] 1 to 110) replaced by the corresponding SINV capsid-coding fragment or the VEEV capsid protein with the frameshift mutations changing a short peptide in the N terminus are highly attenuated but capable of very efficient replication in vitro in cells having defects in alpha/beta interferon signaling. This study provides new data at the cellular level on the mechanism of VEEV cytopathogenicity. These findings can be used for development of new, attenuated VEEV variants and may be applicable to other New World encephalitogenic alphaviruses.
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recombinant sindbis venezuelan equine encephalitis virus is highly attenuated and immunogenic
Journal of Virology, 2003Co-Authors: Slobodan Paessler, Scott C Weaver, Rafik Fayzulin, Michael Anishchenko, Ivorlyne P Greene, Ilya FrolovAbstract:Venezuelan equine encephalitis virus (VEEV) is a member of the Alphavirus genus in the Togaviridae family. VEEV is an enveloped virus with a nonsegmented, positive-sense RNA genome of approximately 11.5 kb. The 5′ two-thirds of the genome encodes four nonstructural proteins (nsP1 to nsP4) that form an enzyme complex required for viral replication (45). After viral RNA entry into the cytoplasm, a nonstructural polyprotein is translated directly from the viral genome and utilized in the production of a full-length, negative-sense replicative RNA intermediate. This RNA is then used as a template for synthesis of positive-sense genomic RNA and for transcription of a subgenomic 26S RNA. The ca. 4-kb subgenomic RNA corresponds to the 3′ one-third of the viral genome and is translated into a structural polyprotein that is proteolytically cleaved into the capsid and envelope glycoproteins E2 and E1 (39). Two hundred forty copies of the capsid protein combine with the genomic viral RNA to form an icosahedral nucleocapsid. Finally, the nucleocapsid buds from the plasma membrane to acquire a lipid envelope with embedded protein spikes containing E1-E2 heterodimers (42, 45). VEEV was a significant human and equine pathogen for much of the past century, and recent epidemics (40, 50) indicate that VEEV still represents a serious public health threat. Furthermore, VEEV is a potential biological warfare and terrorism threat. Alphaviruses in the Venezuelan equine encephalitis (VEE) complex are serologically classified into six distinct antigenic subtypes (48, 51, 52). Historically, only members of subtypes IAB and IC were associated with major epidemics and equine epizootics. During the most recent major outbreak in Venezuela and Colombia (1995) involving subtype IC VEEV, about 100,000 human cases occurred, with more than 300 fatal encephalitis cases estimated (41). VEEV infection of mice leads to a biphasic disease with initial replication in lymphoid tissues followed by viremia and penetration into the central nervous system (CNS), where the virus replicates until the death of the infected animal (16-18, 24). The result of the CNS infection is acute meningoencephalitis that leads to massive death of neuronal cells (6). During VEEV epizootics, equine mortality due to encephalitis can reach 83%; in humans, while the overall mortality rate is low (<1%), neurological disease including disorientation, ataxia, Mental Depression, and convulsions can be detected in up to 14% of infected individuals, especially children (25). Sequelae of VEE-related clinical encephalitis in humans and rats are also described (16, 32). The predominant pathological findings in fatal human VEE cases reveal the following: (i) in the CNS, edema, congestion, hemorrhages, vasculitis, meningitis and encephalitis; (ii) in the lungs, interstitial pneumonia, alveolar hemorrhage, congestion, and edema; (iii) in lymphoid tissue, follicular necrosis and lymphocyte depletion; and (iv) in the liver, diffuse hepatocellular degeneration (10, 11, 26). A small animal (mouse) model for VEE-induced encephalitis and lymphotropism is well established (8, 9, 24, 30); however, mice do not develop the pulmonary (24) and hepatic symptoms that were described at high rates (91 and 61%, respectively) in fatal human VEE cases (10). The live-attenuated TC-83 vaccine strain (TC-83) was developed 4 decades ago by serial passaging of the virulent, subtype IAB Trinidad donkey (TrD) VEEV strain in guinea pig heart cell cultures (2). Currently, TC-83 is still the only VEEV strain available for vaccination of laboratory workers and military personnel. More than 8,000 humans have been vaccinated during the past 4 decades (1, 5, 36). The cumulative data unambiguously demonstrated that nearly 40% of vaccinated people develop a disease with some symptoms typical of natural VEEV infection, including febrile, systemic illness and other adverse effects (1). The TC-83 vaccine has also been used for immunization of equines and can lead to febrile clinical illness with viremia levels that are potentially sufficient to infect mosquitoes and initiate a transmission cycle (23). These findings indicate significant residual virulence of TC-83 strain in both humans and horses (1, 23). Residual virulence has also been detected in mice, where TC-83 was uniformly lethal for the C3H/HeN strain after intracerebral (i.c.) inoculation and produced clinical illness in BALB/c and C3H/HeN mice for almost 14 days after subcutaneous (s.c.) inoculation (34). A formalin-inactivated version of the TC-83 vaccine, C-84, is used for individuals who fail to seroconvert to TC-83 (36). However, repeated boosters with C-84 are required to generate and maintain an effective immune response, and the induced neutralizing antibodies do not persist for a long time. A more promising candidate vaccine was described recently, when attenuation of the VEEV TrD strain was achieved by the introduction of lethal mutations into the PE2 furin cleavage site of an infectious cDNA clone, followed by selection of a second-site suppressor mutation in the E1 glycoprotein (7, 22). This virus is highly attenuated for laboratory rodents. This strategy minimized the potential for direct reversion to virulence. However, the potential for reversion to wild-type virulence via compensatory mutations remains. To further overcome the problem of possible reversion to wild-type virulence of live-attenuated VEEV vaccine strains, we have applied an alternative approach using a recombinant virus capable of inducing a protective immune response. Utilizing the genome of Sindbis virus (SINV), a relatively human-nonpathogenic alphavirus, as a vector, we developed a chimeric SIN/VEE virus expressing all of the structural proteins of the TC-83 strain. This recombinant virus, SIN-83, replicated to high titers in cell culture and protected mice against lethal challenge with high doses of recently isolated, naturally circulating subtype IC and ID strains of VEEV. A distinguishing feature of this SIN-83 chimeric virus is its inability to cause any detectable clinical disease in either adult or weanling mice after either s.c. or i.c. inoculation.
Scott C Weaver - One of the best experts on this subject based on the ideXlab platform.
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analysis of venezuelan equine encephalitis virus capsid protein function in the inhibition of cellular transcription
Journal of Virology, 2007Co-Authors: Natalia Garmashova, Svetlana Atasheva, Wenli Kang, Scott C Weaver, Elena I Frolova, Ilya FrolovAbstract:The genus Alphavirus in the family Togaviridae includes a number of important human and animal pathogens (15). Alphaviruses are currently classified into six antigenic complexes and are widely distributed in both the New World and the Old World. They are efficiently transmitted by mosquitoes, in which they cause a persistent, lifelong infection with a minimal effect on biological functions. In vertebrates, alphavirus infection is acute and in many cases characterized by high-titer viremia, rash, fever, and encephalitis until the death of the infected host or clearance of the virus by the immune system. The encephalitogenic alphaviruses, including Venezuelan (VEEV), eastern (EEEV), and western (WEEV) equine encephalitis viruses, represent a continuous public health threat in the United States (41, 48-50). They circulate in Central, South, and North America and have the ability to cause fatal disease in humans and in horses and other domestic animals. During VEEV epizootics, equine mortality can reach 83%, and in humans, the virus produces a severe temporary immunodeficiency and a greatly debilitating, sometimes fatal disease (42). The overall mortality rate is below 1%, but neurological disease, including disorientation, ataxia, Mental Depression, and convulsions, can be detected in up to 14% of all infected individuals, especially children (23). Also described are sequelae of VEEV-related clinical encephalitis in humans (10, 27). The VEEV genome is represented by a single-stranded RNA molecule of positive polarity that is almost 12 kb in length. It mimics the structure of cellular mRNAs, with a cap at the 5′ terminus and a poly(A) tail at the 3′ end of the RNA. The VEEV genome has been cloned in a cDNA form (24) that allows a wide variety of genetic manipulations to be undertaken. The only experiMental vaccine against VEEV infection that has been used extensively in humans was developed 4 decades ago by serial passaging of the virulent subtype IAB Trinidad Donkey VEEV strain in guinea pig heart cell cultures (3). Over 8,000 humans have been vaccinated during the past 4 decades (2, 6, 37), and the cumulative data indicate that nearly 40% of vaccinated people develop a disease with some symptoms typical of those seen with natural VEEV infection, including a febrile systemic illness and other adverse effects (2, 3, 21). No effective antivirals have been developed against any alphavirus, including VEEV. In spite of the continuous threat of VEEV epidemics, the biology of this virus has been studied less intensively than those of other, less pathogenic alphaviruses, such as Sindbis (SINV) and Semliki Forest (SFV) viruses. These viruses can be readily manipulated in a low-biocontainment environment and represent good models for studying the mechanisms of alphavirus replication, virus-host interactions, and encephalitis development (14). However, important differences in pathogenesis and the severity of human and veterinary diseases suggest that these viruses may not be ideal models for encephalitis. Moreover, the results from recent studies of the Old World (SINV and SFV) and the New World (VEEV and EEEV) alphaviruses (1, 9, 11-13, 36, 46) demonstrated that both of these groups have developed the ability to interfere with cellular transcription and to use this effect as a means of downregulating cellular antiviral response. However, the mechanisms of transcription inhibition appear to be fundaMentally different; while the Old World alphaviruses use nsP2 to inhibit cellular transcription (11), the more encephalitogenic VEEV and EEEV use their capsid proteins for the same function (1, 12). Expression of the latter proteins by different vectors is sufficient for induction of cell death and cytopathic effects (CPE) in tissue culture, and the development of these phenomena strongly correlates with the inhibition of transcription of cellular mRNA and rRNA. Moreover, the replacement of the structural protein genes in the VEEV genome by those derived from SINV has made the chimeric virus significantly less cytopathic and incapable of interfering with the development of an antiviral response in cells having no defect in alpha/beta interferon induction and signaling (12). In this study, we continued our investigation of the VEEV capsid protein-dependent inhibition of cellular transcription. Our data demonstrate that (i) transcriptional shutoff and CPE development in capsid protein-expressing cells are determined by a short CVEE peptide, CVEE33-68, located in the N-terminal part of the protein; (ii) inhibition of transcription and CPE development correlate with the presence of the entire capsid protein or the indicated peptide on the nuclear membrane, which suggests their interaction with the nuclear pore complex (NPC); (iii) the mutations in the peptide might lead to the accumulation of capsid protein in either the cytoplasm or the nucleus; and (iv) VEEV variants encoding either the chimeric capsid protein with the amino-terminal fragment (amino acids [aa] 1 to 110) replaced by the corresponding SINV capsid-coding fragment or the VEEV capsid protein with the frameshift mutations changing a short peptide in the N terminus are highly attenuated but capable of very efficient replication in vitro in cells having defects in alpha/beta interferon signaling. This study provides new data at the cellular level on the mechanism of VEEV cytopathogenicity. These findings can be used for development of new, attenuated VEEV variants and may be applicable to other New World encephalitogenic alphaviruses.
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recombinant sindbis venezuelan equine encephalitis virus is highly attenuated and immunogenic
Journal of Virology, 2003Co-Authors: Slobodan Paessler, Scott C Weaver, Rafik Fayzulin, Michael Anishchenko, Ivorlyne P Greene, Ilya FrolovAbstract:Venezuelan equine encephalitis virus (VEEV) is a member of the Alphavirus genus in the Togaviridae family. VEEV is an enveloped virus with a nonsegmented, positive-sense RNA genome of approximately 11.5 kb. The 5′ two-thirds of the genome encodes four nonstructural proteins (nsP1 to nsP4) that form an enzyme complex required for viral replication (45). After viral RNA entry into the cytoplasm, a nonstructural polyprotein is translated directly from the viral genome and utilized in the production of a full-length, negative-sense replicative RNA intermediate. This RNA is then used as a template for synthesis of positive-sense genomic RNA and for transcription of a subgenomic 26S RNA. The ca. 4-kb subgenomic RNA corresponds to the 3′ one-third of the viral genome and is translated into a structural polyprotein that is proteolytically cleaved into the capsid and envelope glycoproteins E2 and E1 (39). Two hundred forty copies of the capsid protein combine with the genomic viral RNA to form an icosahedral nucleocapsid. Finally, the nucleocapsid buds from the plasma membrane to acquire a lipid envelope with embedded protein spikes containing E1-E2 heterodimers (42, 45). VEEV was a significant human and equine pathogen for much of the past century, and recent epidemics (40, 50) indicate that VEEV still represents a serious public health threat. Furthermore, VEEV is a potential biological warfare and terrorism threat. Alphaviruses in the Venezuelan equine encephalitis (VEE) complex are serologically classified into six distinct antigenic subtypes (48, 51, 52). Historically, only members of subtypes IAB and IC were associated with major epidemics and equine epizootics. During the most recent major outbreak in Venezuela and Colombia (1995) involving subtype IC VEEV, about 100,000 human cases occurred, with more than 300 fatal encephalitis cases estimated (41). VEEV infection of mice leads to a biphasic disease with initial replication in lymphoid tissues followed by viremia and penetration into the central nervous system (CNS), where the virus replicates until the death of the infected animal (16-18, 24). The result of the CNS infection is acute meningoencephalitis that leads to massive death of neuronal cells (6). During VEEV epizootics, equine mortality due to encephalitis can reach 83%; in humans, while the overall mortality rate is low (<1%), neurological disease including disorientation, ataxia, Mental Depression, and convulsions can be detected in up to 14% of infected individuals, especially children (25). Sequelae of VEE-related clinical encephalitis in humans and rats are also described (16, 32). The predominant pathological findings in fatal human VEE cases reveal the following: (i) in the CNS, edema, congestion, hemorrhages, vasculitis, meningitis and encephalitis; (ii) in the lungs, interstitial pneumonia, alveolar hemorrhage, congestion, and edema; (iii) in lymphoid tissue, follicular necrosis and lymphocyte depletion; and (iv) in the liver, diffuse hepatocellular degeneration (10, 11, 26). A small animal (mouse) model for VEE-induced encephalitis and lymphotropism is well established (8, 9, 24, 30); however, mice do not develop the pulmonary (24) and hepatic symptoms that were described at high rates (91 and 61%, respectively) in fatal human VEE cases (10). The live-attenuated TC-83 vaccine strain (TC-83) was developed 4 decades ago by serial passaging of the virulent, subtype IAB Trinidad donkey (TrD) VEEV strain in guinea pig heart cell cultures (2). Currently, TC-83 is still the only VEEV strain available for vaccination of laboratory workers and military personnel. More than 8,000 humans have been vaccinated during the past 4 decades (1, 5, 36). The cumulative data unambiguously demonstrated that nearly 40% of vaccinated people develop a disease with some symptoms typical of natural VEEV infection, including febrile, systemic illness and other adverse effects (1). The TC-83 vaccine has also been used for immunization of equines and can lead to febrile clinical illness with viremia levels that are potentially sufficient to infect mosquitoes and initiate a transmission cycle (23). These findings indicate significant residual virulence of TC-83 strain in both humans and horses (1, 23). Residual virulence has also been detected in mice, where TC-83 was uniformly lethal for the C3H/HeN strain after intracerebral (i.c.) inoculation and produced clinical illness in BALB/c and C3H/HeN mice for almost 14 days after subcutaneous (s.c.) inoculation (34). A formalin-inactivated version of the TC-83 vaccine, C-84, is used for individuals who fail to seroconvert to TC-83 (36). However, repeated boosters with C-84 are required to generate and maintain an effective immune response, and the induced neutralizing antibodies do not persist for a long time. A more promising candidate vaccine was described recently, when attenuation of the VEEV TrD strain was achieved by the introduction of lethal mutations into the PE2 furin cleavage site of an infectious cDNA clone, followed by selection of a second-site suppressor mutation in the E1 glycoprotein (7, 22). This virus is highly attenuated for laboratory rodents. This strategy minimized the potential for direct reversion to virulence. However, the potential for reversion to wild-type virulence via compensatory mutations remains. To further overcome the problem of possible reversion to wild-type virulence of live-attenuated VEEV vaccine strains, we have applied an alternative approach using a recombinant virus capable of inducing a protective immune response. Utilizing the genome of Sindbis virus (SINV), a relatively human-nonpathogenic alphavirus, as a vector, we developed a chimeric SIN/VEE virus expressing all of the structural proteins of the TC-83 strain. This recombinant virus, SIN-83, replicated to high titers in cell culture and protected mice against lethal challenge with high doses of recently isolated, naturally circulating subtype IC and ID strains of VEEV. A distinguishing feature of this SIN-83 chimeric virus is its inability to cause any detectable clinical disease in either adult or weanling mice after either s.c. or i.c. inoculation.
Dietmar Fuchs - One of the best experts on this subject based on the ideXlab platform.
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neopterin production tryptophan degradation and Mental Depression what is the link
Brain Behavior and Immunity, 2002Co-Authors: Bernhard Widner, Andreas Laich, B Spernerunterweger, Maximilian Ledochowski, Dietmar FuchsAbstract:The cytokine interferon-gamma stimulates human monocytes/macrophages to release large amounts of neopterin. Increased neopterin concentrations in body fluids of patients are observed during diseases with activated cellular (=TH1-type) immune response such as allograft rejection, virus infections, autoimmune disorders, or malignant tumors but also in neurodegenerative diseases or during pregnancy. In various cells interferon-gamma induces indoleamine 2,3-dioxygenase (IDO) which degrades tryptophan via the kynurenine pathway. Therefore like increased neopterin formation, enhanced tryptophan degradation is observed in diseases concomitant with cellular immune activation. Disturbed metabolism of tryptophan affects biosynthesis of neurotransmitter 5-hydroxytryptamine (serotonin), and it appears to be associated with an increased susceptibility for Depression. In fact, enhanced neopterin concentrations together with increased degradation of tryptophan and low serum levels of tryptophan correlate with neuropsychiatric abnormalities like cognitive decline and depressive symptoms especially in long-lasting and chronic diseases. Activation of IDO could represent an important link between the immunological network and the pathogenesis of Depression.
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more rapid method for simultaneous measurement of tryptophan and kynurenine by hplc
Clinical Chemistry, 2002Co-Authors: Andreas Laich, Bernhard Widner, Gabriele Neurauter, Dietmar FuchsAbstract:The essential amino acid l-tryptophan is important in nitrogen balance and the maintenance of muscle mass and body weight in humans (1). Moreover, tryptophan is the precursor for the biosynthesis of the neurotransmitter serotonin (5-hydroxytryptamine). Insufficient availability of tryptophan may increase susceptibility for Mental Depression (2). On activation of cellular immunity, the T-cell-derived cytokine interferon-γ stimulates the enzyme indoleamine-(2,3)-dioxygenase (IDO) in various cells (3)(4). IDO catalyzes the initial step of tryptophan catabolism within the biosynthetic pathway of nicotinamide dinucleotides, and N -formyl-kynurenine is formed as a first intermediate. The kynurenine-to-tryptophan ratio has been a sensitive estimate to monitor the activation status of IDO and of cellular immunity both in vivo and in vitro (3)(5). In patients, a decrease in serum tryptophan and a parallel increase of kynurenine attributable to IDO activation is found in various diseases associated with T-cell activation, such as viral infections, autoimmune disorders, and malignant diseases (3)(5)(6)(7)(8). More recently, activation of IDO in monocytes/macrophages was found to interfere with the proliferative capacity of T cells in response to antigenic stimulation by the withdrawal of tryptophan (9). This finding has attracted immunologic researchers especially to explore the possible involvement of IDO in tolerance induction and in diseases that are associated with acquired immunodeficiency. We have described a reverse-phase HPLC method to quantify serum concentrations of kynurenine and tryptophan in parallel with use of 3-nitro-l-tyrosine as an internal standard (10). Here we report an optimized protocol that uses a shorter HPLC column and a different elution buffer, allowing faster throughput of samples …
Colomé, Lucas Marques - One of the best experts on this subject based on the ideXlab platform.
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Osmotic Demyelination Syndrome after Primary Hypoadrenocorticism Crisis Management
'Universidade Federal do Rio Grande do Sul', 2021Co-Authors: Pöppl, Álan Gomes, Pires, Érico Haas, Barbieri, Claudia Ruga, Colomé, Lucas MarquesAbstract:Background: Primary hypoadrenocorticism is a rare condition resulting from immune-mediated destruction of the adrenal cortices. It can also occur due to necrosis, neoplasms, infarctions and granulomas. The clinical and laboratory changes are due to deficient secretion of glucocorticoids and mineralocorticoids, which leads to electrolyte disorders associated with hyponatremia and hyperkalemia. These disorders can cause hypotension, hypovolemia and shock, putting a patient's life at risk if inadequate hydroelectrolytic supplementation and hormone replacement is provided. Nevertheless, rapid sodium chloride supplementation is contraindicated due to the risk of central pontine myelinolysis induction. The present study aims to describe a thalamic osmotic demyelination syndrome after management of a primary hypoadrenocorticism crisis in a 2-year-old, female West White Highland Terrier. Case: The patient had a presumptive diagnosis of hypoadrenocorticism already receiving oral prednisolone and gastrointestinal protectants in the last 2 days. After prednisolone dose reduction the dog presented a severe primary hypoadrenocorticism crisis treated with intravenous sodium chloride 0.9% solution along with supportive therapy. Four days after being discharged from the hospital, the patient showed severe neurological impairment and went back to the clinic where a neurological examination revealed Mental Depression, drowsiness, ambulatory tetraparesis and proprioceptive deficit of the 4 limbs, postural deficits, and cranial nerves with decreased response. Due to these clinical signs, a magnetic resonance imaging was performed. It showed 2 intra-axial circular lesions, symmetrically distributed in both thalamus sides, with approximately 0.8 cm in diameter each without any other anatomical changes on magnetic resonance imaging. The images were compatible with metabolic lesions, suggesting demyelination. Furthermore, liquor analysis did not show relevant abnormalities, except for a slight increase in density and pH at the upper limit of the reference range. After treatment, the patient had a good neurological evolution secondary to standard primary hypoadrenocorticism treatment, without sequelae. Discussion: In the present case report, primary hypoadrenocorticism gastrointestinal signs seemed to be triggered by a food indiscretion episode, not responsive to the symptomatic therapies employed. The patient´s breed and age (young West White Highland Terrier bitch) is in accordance with the demographic profile of patients affected by the disease, where young females are frequently more affected. Regarding the probable thalamic osmotic demyelination syndrome documented in this case, is important to notice that myelinolysis or demyelination is an exceedingly rare noninflammatory neurological disorder, initially called central pontine myelinolysis, which can occur after rapid correction of hyponatremia. It has already been observed in dogs after correction of hyponatremia of different origins, including hypoadrenocorticism and parasitic gastrointestinal disorders. Currently, the terms "osmotic myelinolysis" or “osmotic demyelination syndrome" are considered more suitable when compared to the term "central pontine myelinolysis" since it has been demonstrated in dogs and humans the occurrence of demyelination secondary to the rapid correction of hyponatremia in distinct regions of the central nervous system including pons, basal nuclei, striatum, thalamus, cortex, hippocampus and cerebellum. The present case report emphasizes the difficulties for hormonal confirmation of primary hypoadrenocorticism in a patient already on corticosteroid treatment, as well as proposes that the current term osmotic demyelination syndrome replace the term “central pontine myelinolysis” in veterinary literature related to the management of hypoadrenocorticism crisis.Keywords: Addison Syndrome, hyponatremia, osmotic myelinolysis, magnetic resonance imaging
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Osmotic demyelination syndrome after primary hypoadrenocorticism crisis management
2021Co-Authors: Pöppl, Álan Gomes, Pires, Érico Haas, Barbieri, Claudia Ruga, Colomé, Lucas MarquesAbstract:Background: Primary hypoadrenocorticism is a rare condition resulting from immune-mediated destruction of the adrenal cortices. It can also occur due to necrosis, neoplasms, infarctions, and granulomas. The clinical and laboratory changes are due to deficient secretion of glucocorticoids and mineralocorticoids, which leads to electrolyte disorders associated with hyponatremia and hyperkalemia. These disorders can cause hypotension, hypovolemia and shock, putting a patient’s life at risk if inadequate hydroelectrolytic supplementation and hormone replacement is provided. Nevertheless, rapid sodium chloride supplementation is contraindicated due to the risk of central pontine myelinolysis induction. The present study aims to describe a thalamic osmotic demyelination syndrome after management of a primary hypoadrenocorticism crisis in a 2-year-old, female West White Highland Terrier. Case: The patient had a presumptive diagnosis of hypoadrenocorticism already receiving oral prednisolone and gastrointestinal protectants in the last 2 days. After prednisolone dose reduction the dog presented a severe primary hypoadrenocorticism crisis treated with intravenous sodium chloride 0.9% solution along with supportive therapy. Four days after being discharged from the hospital, the patient showed severe neurological impairment and went back to the clinic where a neurological examination revealed Mental Depression, drowsiness, ambulatory tetraparesis and proprioceptive deficit of the 4 limbs, postural deficits, and cranial nerves with decreased response. Due to these clinical signs, a magnetic resonance imaging was performed. It showed 2 intra-axial circular lesions, symmetrically distributed in both thalamus sides, with approximately 0.8 cm in diameter each without any other anatomical changes on magnetic resonance imaging. The images were compatible with metabolic lesions, suggesting demyelination. Furthermore, liquor analysis did not show relevant abnormalities, except for a slight increase in density and pH at the upper limit of the reference range. After treatment, the patient had a good neurological evolution secondary to standard primary hypoadrenocorticism treatment, without sequelae. Discussion: In the present case report, primary hypoadrenocorticism gastrointestinal signs seemed to be triggered by a food indiscretion episode, not responsive to the symptomatic therapies employed. The patient´s breed and age (young West White Highland Terrier bitch) is in accordance with the demographic profile of patients affected by the disease, where young females are frequently more affected. Regarding the probable thalamic osmotic demyelination syndrome documented in this case, is important to notice that myelinolysis or demyelination is an exceedingly rare noninflammatory neurological disorder, initially called central pontine myelinolysis, which can occur after rapid correction of hyponatremia. It has already been observed in dogs after correction of hyponatremia of different origins, including hypoadrenocorticism and parasitic gastrointestinal disorders. Currently, the terms “osmotic myelinolysis” or “osmotic demyelination syndrome” are considered more suitable when compared to the term “central pontine myelinolysis” since it has been demonstrated in dogs and humans the occurrence of demyelination secondary to the rapid correction of hyponatremia in distinct regions of the central nervous system including pons, basal nuclei, striatum, thalamus, cortex, hypoppocampus, and cerebelum. The present case report emphasizes the difficulties for hormonal confirmation of primary hypoadrenocorticism in a patient already on corticosteroid treatment, as well as proposes that the current term osmotic demyelination syndrome replace the term “central pontine myelinolysis” in veterinary literature related to the management of hypoadrenocorticism crisis
Slobodan Paessler - One of the best experts on this subject based on the ideXlab platform.
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recombinant sindbis venezuelan equine encephalitis virus is highly attenuated and immunogenic
Journal of Virology, 2003Co-Authors: Slobodan Paessler, Scott C Weaver, Rafik Fayzulin, Michael Anishchenko, Ivorlyne P Greene, Ilya FrolovAbstract:Venezuelan equine encephalitis virus (VEEV) is a member of the Alphavirus genus in the Togaviridae family. VEEV is an enveloped virus with a nonsegmented, positive-sense RNA genome of approximately 11.5 kb. The 5′ two-thirds of the genome encodes four nonstructural proteins (nsP1 to nsP4) that form an enzyme complex required for viral replication (45). After viral RNA entry into the cytoplasm, a nonstructural polyprotein is translated directly from the viral genome and utilized in the production of a full-length, negative-sense replicative RNA intermediate. This RNA is then used as a template for synthesis of positive-sense genomic RNA and for transcription of a subgenomic 26S RNA. The ca. 4-kb subgenomic RNA corresponds to the 3′ one-third of the viral genome and is translated into a structural polyprotein that is proteolytically cleaved into the capsid and envelope glycoproteins E2 and E1 (39). Two hundred forty copies of the capsid protein combine with the genomic viral RNA to form an icosahedral nucleocapsid. Finally, the nucleocapsid buds from the plasma membrane to acquire a lipid envelope with embedded protein spikes containing E1-E2 heterodimers (42, 45). VEEV was a significant human and equine pathogen for much of the past century, and recent epidemics (40, 50) indicate that VEEV still represents a serious public health threat. Furthermore, VEEV is a potential biological warfare and terrorism threat. Alphaviruses in the Venezuelan equine encephalitis (VEE) complex are serologically classified into six distinct antigenic subtypes (48, 51, 52). Historically, only members of subtypes IAB and IC were associated with major epidemics and equine epizootics. During the most recent major outbreak in Venezuela and Colombia (1995) involving subtype IC VEEV, about 100,000 human cases occurred, with more than 300 fatal encephalitis cases estimated (41). VEEV infection of mice leads to a biphasic disease with initial replication in lymphoid tissues followed by viremia and penetration into the central nervous system (CNS), where the virus replicates until the death of the infected animal (16-18, 24). The result of the CNS infection is acute meningoencephalitis that leads to massive death of neuronal cells (6). During VEEV epizootics, equine mortality due to encephalitis can reach 83%; in humans, while the overall mortality rate is low (<1%), neurological disease including disorientation, ataxia, Mental Depression, and convulsions can be detected in up to 14% of infected individuals, especially children (25). Sequelae of VEE-related clinical encephalitis in humans and rats are also described (16, 32). The predominant pathological findings in fatal human VEE cases reveal the following: (i) in the CNS, edema, congestion, hemorrhages, vasculitis, meningitis and encephalitis; (ii) in the lungs, interstitial pneumonia, alveolar hemorrhage, congestion, and edema; (iii) in lymphoid tissue, follicular necrosis and lymphocyte depletion; and (iv) in the liver, diffuse hepatocellular degeneration (10, 11, 26). A small animal (mouse) model for VEE-induced encephalitis and lymphotropism is well established (8, 9, 24, 30); however, mice do not develop the pulmonary (24) and hepatic symptoms that were described at high rates (91 and 61%, respectively) in fatal human VEE cases (10). The live-attenuated TC-83 vaccine strain (TC-83) was developed 4 decades ago by serial passaging of the virulent, subtype IAB Trinidad donkey (TrD) VEEV strain in guinea pig heart cell cultures (2). Currently, TC-83 is still the only VEEV strain available for vaccination of laboratory workers and military personnel. More than 8,000 humans have been vaccinated during the past 4 decades (1, 5, 36). The cumulative data unambiguously demonstrated that nearly 40% of vaccinated people develop a disease with some symptoms typical of natural VEEV infection, including febrile, systemic illness and other adverse effects (1). The TC-83 vaccine has also been used for immunization of equines and can lead to febrile clinical illness with viremia levels that are potentially sufficient to infect mosquitoes and initiate a transmission cycle (23). These findings indicate significant residual virulence of TC-83 strain in both humans and horses (1, 23). Residual virulence has also been detected in mice, where TC-83 was uniformly lethal for the C3H/HeN strain after intracerebral (i.c.) inoculation and produced clinical illness in BALB/c and C3H/HeN mice for almost 14 days after subcutaneous (s.c.) inoculation (34). A formalin-inactivated version of the TC-83 vaccine, C-84, is used for individuals who fail to seroconvert to TC-83 (36). However, repeated boosters with C-84 are required to generate and maintain an effective immune response, and the induced neutralizing antibodies do not persist for a long time. A more promising candidate vaccine was described recently, when attenuation of the VEEV TrD strain was achieved by the introduction of lethal mutations into the PE2 furin cleavage site of an infectious cDNA clone, followed by selection of a second-site suppressor mutation in the E1 glycoprotein (7, 22). This virus is highly attenuated for laboratory rodents. This strategy minimized the potential for direct reversion to virulence. However, the potential for reversion to wild-type virulence via compensatory mutations remains. To further overcome the problem of possible reversion to wild-type virulence of live-attenuated VEEV vaccine strains, we have applied an alternative approach using a recombinant virus capable of inducing a protective immune response. Utilizing the genome of Sindbis virus (SINV), a relatively human-nonpathogenic alphavirus, as a vector, we developed a chimeric SIN/VEE virus expressing all of the structural proteins of the TC-83 strain. This recombinant virus, SIN-83, replicated to high titers in cell culture and protected mice against lethal challenge with high doses of recently isolated, naturally circulating subtype IC and ID strains of VEEV. A distinguishing feature of this SIN-83 chimeric virus is its inability to cause any detectable clinical disease in either adult or weanling mice after either s.c. or i.c. inoculation.