Two-photon imaging of remyelination of spinal cord axons by engrafted neural precursor cells in a viral model of multiple sclerosis.

Neural precursor cells (NPCs) offer a promising approach for treating demyelinating diseases. However, the cellular dynamics that underlie transplanted NPC-mediated remyelination have not been described. Using two-photon imaging of a newly developed ventral spinal cord preparation and a viral model of demyelination, we describe the motility and intercellular interactions of transplanted mouse NPCs expressing green fluorescent protein (GFP) with damaged axons expressing yellow fluorescent protein (YFP). Our findings reveal focal axonal degeneration that occurs in the ventral side of the spinal cord within 1 wk following intracranial instillation with the neurotropic JHM strain of mouse hepatitis virus (JHMV). Axonal damage precedes extensive demyelination and is characterized by swelling along the length of the axon, loss of YFP signal, and transected appearance. NPCs engrafted into spinal cords of JHMV-infected mice exhibited diminished migration velocities and increased proliferation compared with transplanted cells in noninfected mice. NPCs preferentially accumulated within areas of axonal damage, initiated direct contact with axons, and subsequently expressed the myelin proteolipid protein gene, initiating remyelination. These findings indicate that NPCs transplanted into an inflammatory demyelinating microenvironment participate directly in therapeutic outcome through the wrapping of myelin around damaged neurons.

Interactions of Hepatitis B Virus Infection with Nonalcoholic Fatty Liver Disease: Possible Mechanisms and Clinical Impact.

Hepatitis B virus (HBV) infection is a major etiology of chronic liver disease worldwide. In the past decade, nonalcoholic fatty liver disease (NAFLD) has emerged as a common liver disorder in general population. Accordingly, the patient number of chronic hepatitis B (CHB) concomitant with NAFLD grows rapidly. The present article reviewed the recent studies aiming to explore the relationship between CHB and NAFLD from different aspects, including the relevant pathogenesis of CHB and NAFLD, the intracellular molecular mechanisms overlaying HBV infection and hepatic steatosis, and the observational studies with animal models and clinical cohorts for analyzing the coincidence of the two diseases. It is concluded that although numerous cross-links have been suggested between the molecular pathways in HBV infection and NAFLD pathogenesis, regarding whether HBV infection can substantially interfere with the occurrence of NAFLD or vice versa in the patients, there is still far from a conclusive agreement.

Establishment of Duplex SYBR Green I-based Real-time PCR Assay for Simultaneous Detection of Duck Hepatitis A Virus-1 and Duck Astrovirus-3.

Duck viral hepatitis (DVH) mainly affects ducklings under 1 month of age, causes liver necrosis, enlargement, and hemorrhage, and is highly lethal, seriously jeopardizing the duck industry. The prevalence of duck hepatitis A virus (DHAV-1) and duck astrovirus type 3 (DAstV-3) is increasing, and coinfection is common. Moreover, the similar clinical characteristics of the DHAV-1 and DAstV-3 infections and the high frequency of coinfection make diagnosis difficult. In this study, to establish a method for the rapid, simultaneous detection of DHAV-1 and DAstV-3, two pairs of specific primers were designed according to their conserved gene regions. An SYBR® Green I-based qPCR assay was successfully established that can quickly and differentially detect the two viruses. Moreover, the assay is highly specific and does not show cross-reaction with other common viruses. The detection limit of the method is 7.34 × 101 copies/µl and 3.78 × 101 copies/µl for DHAV-1 and DAstV-3, respectively, indicating high sensitivity. A total of 34 clinical samples were tested using the established method; the positive rates for DHAV-1 and DAstV-3 were 14.71% and 8.82%, respectively, and that for coinfection was 2.94% (1/34), which was better than that obtained with conventional PCR. In summary, the SYBR Green I-based qPCR assay established in this study has high specificity, good sensitivity and accuracy, high feasibility, and is rapid. Thus, it can be a powerful tool for the coinfection detection of DHAV-1 and DAstV-3 and for future epidemiologic studies.

Artículo regular­Establecimiento de un ensayo dúplex de PCR en tiempo real basado en SYBR Green I para la detección simultánea del virus de la hepatitis A del pato-1 y del astrovirus del pato tipo 3. La hepatitis viral del pato (DVH) afecta principalmente a los patitos menores de 1 mes de edad, causa necrosis hepática, agrandamiento y hemorragia, y es altamente letal, lo que pone en grave peligro la industria del pato. La prevalencia del virus de la hepatitis A del pato (DHAV-1) y del astrovirus del pato tipo 3 (DAstV-3) está aumentando y la coinfección es común. Además, las características clínicas similares de las infecciones por el virus de la hepatitis A del pato y el astrovirus del pato tipo 3 así como la alta frecuencia de coinfección dificultan el diagnóstico. En este estudio, para establecer un método para la detección rápida y simultánea por el virus de la hepatitis A del pato y el astrovirus del pato tipo 3, se diseñaron dos pares de iniciadores específicos según sus regiones génicas conservadas. Se estableció con éxito un ensayo cuantitativo de PCR basado en SYBR® Green I que pudo detectar rápida y diferencialmente los dos virus. Además, el ensayo es muy específico y no muestró reacción cruzada con otros virus comunes. El límite de detección del método fue de 7.34 × 101 copias/µl y de 3.78 × 101 copias/µl para el virus de la hepatitis A del pato y para el astrovirus del pato tipo 3, respectivamente, lo que indica una alta sensibilidad. Se analizaron un total de 34 muestras clínicas utilizando el método establecido; las tasas positivas para el virus de la hepatitis A del pato y para el astrovirus del pato tipo 3 fueron del 14.71% y 8.82%, respectivamente y la de coinfección fue del 2.94% (1/34), que fue mejor que la obtenida con el método de PCR convencional. En resumen, el ensayo cuantitativo de PCR basado en SYBR Green I establecido en este estudio tiene alta especificidad, buena sensibilidad y precisión, alta viabilidad y es rápido. Por lo tanto, puede ser una herramienta poderosa para la detección de coinfecciones con el virus de la hepatitis A del pato y astrovirus del pato tipo 3 y para futuros estudios epidemiológicos.

High incidence of glomerulonephritis associated with inclusion body hepatitis in broiler chickens: routine histopathology and histomorphometric studies.

During the routine histologic evaluation of an outbreak of inclusion body hepatitis (IBH) in Mississippi broilers, a high incidence of renal enlargement and glomerulonephropathy was observed in the birds presenting classic hepatic pathology. Characteristic intranuclear adenoviral inclusion bodies were demonstrated in the livers of these birds, and fowl adenovirus was identified by viral isolation and by PCR. The glomerular lesions were consistent with proliferative or membranoproliferative forms of glomerulonephritis. Histomorphometric evaluations were performed to generate a more quantitative analysis of altered glomerular size and cellularity, to detect statistically significant borderline changes, and to get a clearer insight into the incidence of the glomerular alterations. Marked increases in both the average glomerular size (area) and the total glomerular cellularity were observed for the affected glomeruli relative to normal controls. The average glomerular area values for normal glomeruli in the peripheral subcapsular cortical and central cortical kidney regions were 1791 microm2 and 5302 microm2, respectively. In contrast, glomerular measurements for kidneys exhibiting glomerulonephritis by routine histopathology, had average values for the two regions of 4429 microm2 and 11,063 microm2. The average glomerular cell counts for the two regions in controls were 44 and 107 cells/ glomeruli, while averages for birds with glomerulonephritis were 85 and 193 cells/glomeruli. The proportion of IBH-associated glomeruli greater than two standard deviations above the mean glomerular size of the normal controls was 52% for the central region and 62% for the peripheral region.

Complete genome analysis of avian hepatitis E virus from chicken with hepatic rupture hemorrhage syndrome.

Since 2016, severe outbreaks of hepatic rupture hemorrhage syndrome (HRHS) associated with infections of tentative novel avian hepatitis E virus (HEV) have emerged in chickens in China, causing increased mortality and decreased laying rate in adult hens and disturbing the hatching and breeding of chicks. To further identify the genotype and gain a better understanding of the genetic properties of the avian HEV responsible for that, a strain from Hebei province was isolated, purified and sequenced in this study. Results identified a novel avian HEV genotype, sharing 79.5-86.9% identities with other published avian HEV strains, and having higher identities with Orthohepevirus A HEV strains. More importantly, the new isolate contains various amino-acid substitutions in its functional proteins, including methyltransferase, helicase, RNA-dependent RNA polymerase. The data presented in this report will enhance the current understanding of the genetic diversity of the avian HEV and provide additional insight into the critical factors that determine the pathogenicity.

Regeneration of oligodendroglia during recovery from demyelinating disease.

Infection of mice with the JHM strain of mouse hepatitis virus causes demyelination as a result of a cytolytic infection of oligodendroglia. In recovery, animals show remyelination, which could result either from surviving oligodendrocytes extending their territory or by generation of new oligodendroglia. Electron microscopic autoradiographic studies with 3H-labeled thymidine demonstrate that the cells associated with remyelination are newly generated oligodendroglia.

Cross-species hybridization of woodchuck hepatitis viral infection-induced woodchuck hepatocellular carcinoma using human, rat and mouse oligonucleotide microarrays.

BACKGROUND AND AIM: We aimed to evaluate the transcriptional characteristics of viral infection-induced woodchuck hepatocellular carcinoma (HCC), to compare the use of human, rat and mouse gene arrays for cross-species hybridization, and to look into gene expression profiles in woodchuck HCC by the combined use of these arrays. METHODS: Commercially available human, rat and mouse oligonucleotide microarrays were used to determine the gene expression profiles on the same woodchuck liver samples. Differentially expressed genes between HCC and the surrounding hepatic tissues found in the arrays were selected for quantitative reverse transcription polymerase chain reaction. RESULTS: Despite the difference in the number of the probes from each array, the percentage of genes that were detectable was similar. Stringent microarray data analysis using both supervised and unsupervised methods identified 281 differentially expressed genes via the human array with a false discovery rate (FDR) of 0.99%, 107 genes via the rat array with an FDR of 1.85% and 78 genes via the mouse array with an FDR of 7.41%. Eleven genes were differentially changed in all three arrays that include the upregulation of NPM1, H2AFZ, EEF1G, HNRPAB, RPS18, EIF5, CKS2, ARIH1, RPS12 and RPS10, and the downregulation of EGR1. The quantitative reverse transcription polymerase chain reaction with woodchuck-specific primers confirmed the reliability of the microarray results. CONCLUSION: This study further demonstrated the utility of cross-species hybridization of microarrays on woodchuck HCC. A combined use of three types of arrays identified more differential genes in HCC than individual arrays with the human array providing the richest information among the three arrays used.

A mouse model linking viral hepatitis and salivary gland dysfunction.

OBJECTIVE: Viral hepatitis is known to cause xerostomia in humans, but this has not been reported in an animal model. We report a severe, acute, highly reproducible saliva deficiency occurring in BALB/c mice as a result of experimental viral hepatitis. MATERIALS AND METHODS: BALB/c mice, splenectomized or carrying genetic mutations to detect immunological contributions to the saliva deficiency syndrome, were infected intraperitoneally with a non-lethal dose of murine cytomegalovirus. Pilocarpine-stimulated saliva volumes were determined between 0 and 15 days after infection. Salivary gland, liver, spleen, and sera were analyzed for the presence of virus, cytokines, inflammatory infiltrates, and tissue damage. RESULTS: Saliva deficiency was detectable 2 days after cytomegalovirus infection, peaked at 88% below normal by day 7, and resolved partially in all mice by 15 days postinfection as sialoadenitis increased. Neither salivary gland viral titers, sialoadenitis, splenectomy, nor systemic inflammatory markers correlated with hyposalivation severity. Elevated liver enzymes did correlate with hyposalivation, and mice genetically resistant to murine cytomegalovirus-induced hepatitis were significantly protected. CONCLUSIONS: Murine cytomegalovirus-induced salivary gland dysfunction is biphasic, with an acute hepatitis-associated phase and a later sialoadenitis-associated phase. Acute murine cytomegalovirus infection of BALB/c mice may provide a model for investigation of hepatitis-associated xerostomia.

A PDGFRß-based score predicts significant liver fibrosis in patients with chronic alcohol abuse, NAFLD and viral liver disease.

BACKGROUND: Platelet Derived Growth Factor Receptor beta (PDGFRß) has been associated to hepatic stellate cell activation and has been the target of multiple therapeutic studies. However, little is known concerning its use as a diagnostic agent. METHODS: Circulating PDGFRß levels were analysed in a cohort of patients with liver fibrosis/cirrhosis due to chronic alcohol abuse, viral hepatitis, or non-alcoholic fatty liver disease (NAFLD). The diagnostic performance of PDGFRß as individual blood parameter, or in combination with other metabolic factors was evaluated. FINDINGS: sPDGFRß levels are progressively increased with increasing fibrosis stage and the largest difference was observed in patients with significant fibrosis, compared to no or mild fibrosis. The accuracy of sPDGFRß-levels predicting fibrosis could be increased by combining it with albumin levels and platelet counts into a novel diagnostic algorithm, the PRTA-score, generating a predictive value superior to Fib-4, APRI, and AST/ALT. The sPDGFRß levels and the PRTA-score are independent of liver disease aetiology, thus overcoming one of the major weaknesses of current non-invasive clinical and experimental scores. Finally, we confirmed the diagnostic value of sPDGFRß levels and the PRTA-score in an independent patient cohort with NAFLD which was staged for fibrosis by liver biopsy. INTERPRETATION: The PRTA-score is an accurate tool for detecting significant liver fibrosis in a broad range of liver disease aetiologies. FUND: Vrije Universiteit Brussel, the Institute for the Promotion of Innovation through Science and Technology in Flanders (IWT-Flanders) (HILIM-3D; SBO140045), and the Fund of Scientific Research Flanders (FWO).

Differential gene expression profiles in acute hepatic failure model in mice infected with MHV-3 virus intervened by anti-hepatic failure compound.

Differential gene expression profiles in Balb/cJ mouse model of acute hepatic failure infected with MHV-3 virus intervened by anti-hepatic failure compound (AHFC) and the changes of cytokines regulated by genes were investigated. The Balb/cj mice were divided into AHFC-intervened group and control group randomly. Acute hepatic failure model of Balb/cJ mice infected with MHV-3 virus was established. The survival rate in the two groups was observed. It was found that the survival rate in the AHFC-intervened group and control group was 90% and 50% respectively 48 h after intraperitoneal injection of MHV-3 (P<0.05). Before and after the experiment, the cytokines in peripheral blood of the survival mice were determined, and RNA was extracted from survival mouse liver tissue for the analysis of the differential gene expression by a 36 kb mouse oligonucleotide DNA array. In all the genes of microarray there were 332 genes expressed differently in the two groups, in which 234 genes were up-regulated and 78 genes down-regulated. Through clustering analysis, the differential expression of immune related genes, including TNF receptor superfamily, Kctd9, Bcl-2, Fgl2, IL-8, IL-6, IFN-gamma, TNF-alpha etc. might be related with the curative effectiveness of AHFC. It was suggested that AHFC can balance the immune state of mouse model of acute hepatic failure infected with MHV-3 virus mainly through regulating the expression of immune related genes, decrease the immune damage and inhibit liver cell apoptosis of mouse acute hepatic failure model obviously so as to increase the survival rate of mouse models of acute hepatic failure.

An optimized method for enumerating CNS derived memory B cells during viral-induced inflammation.

BACKGROUND: CNS inflammation resulting from infection, injury, or neurodegeneration leads to accumulation of diverse B cell subsets. Although antibody secreting cells (ASC) within the inflamed CNS have been extensively examined, memory B cell (Bmem) characterization has been limited as they do not secrete antibody without stimulation. Moreover, unlike human Bmem, reliable surface markers for murine Bmem remain elusive. NEW METHOD: Using a viral encephalomyelitis model we developed a modified limiting dilution in vitro stimulation assay to convert CNS-derived virus specific Bmem into ASC. COMPARISON WITH EXISTING METHODS: Stimulation methods established for lymphoid tissue cells using prolonged stimulation with viral lysate resulted in substantial ASC loss and minimal Bmem to ASC conversion of CNS-derived cells. By varying stimulation duration, TLR activators, and culture supplements, we achieved optimal conversion by culturing cells with TLR7/8 agonist R848 in the presence of feeder cells for 2days. RESULTS: Flow cytometry markers CD38 and CD73 characterizing murine Bmem from lymphoid tissue showed more diverse expression patterns on corresponding CNS-derived B cell subsets. Using the optimized TLR7/8 stimulation protocol, we compared virus-specific IgG Bmem versus pre-existing ASC within the brain and spinal cord. Increasing Bmem frequencies during chronic infection mirrored kinetics of ASC. However, despite initially similar Bmem and ASC accumulation, Bmem prevailed in the brain, but were lower than ASC in the spinal cord during persistence. CONCLUSION: Simultaneous enumeration of antigen-specific Bmem and ASC using the Bmem assay optimized for CNS-derived cells enables characterization of temporal changes during microbial or auto-antigen induced neuroinflammation.

Large nontransplanted hepatocellular carcinoma in woodchucks: treatment with adenovirus-mediated delivery of interleukin 12/B7.1 genes.

BACKGROUND: Cytokine-based gene therapy strategies efficiently stimulate immune responses against many established transplanted tumors, leading to rejection of the tumor. In this study, we investigated the therapeutic potential of cancer immunotherapy in a clinically more relevant model, woodchucks with primary hepatocellular carcinomas induced by woodchuck hepatitis virus. METHODS: Large (2-5 cm), established intrahepatic tumors were given an injection once with 1 x 10(9) plaque-forming units of AdIL-12/B7.1, an adenovirus vector carrying genes for murine interleukin 12 and B7.1, or of AdEGFP, the control virus, and regression of the tumors was then monitored. Five animals were used in total. RESULTS: In four tumor-bearing animals, the antitumor response was assessed by autopsy and histologic analysis within 1-2 weeks after treatment. In all animals treated with AdIL-12/B7.1 therapy versus AdEGFP therapy, we observed substantial tumor regression (P =.006; two-sided unpaired Student's t test) accompanied by a massive infiltration of T lymphocytes. These tumors also contained increased levels of CD4(+) and CD8(+) T cells and interferon gamma (IFN gamma). In continuously growing tumor nodules given an injection of the control virus or in nontumoral liver, no such effects (i.e., tumor regression and increased levels of CD4(+) and CD8(+) T cells and IFN gamma) were detected. In the fifth animal, monitored for long-term antitumor efficacy by magnetic resonance imaging (MRI) after intratumoral vector administration by MRI guidance, the tumor was almost completely eliminated (> or = 95%) 7 weeks after treatment. CONCLUSION: Adenovirus vector-based immunotherapy appears to be an effective treatment of large nontransplanted (orthotopic) tumors that acquire malignant characteristics in a stepwise process, reflecting the real-world scenario of hepatocellular carcinoma in humans.

Influence of aflatoxin B1 intoxication on duck livers with duck hepatitis B virus infection.

In an attempt to determine the effect of aflatoxin B1 (AFB) intoxication on livers with duck hepatitis B virus (DHBV) infection, domestic ducks were given 0.1 mg of AFB/kg body weight twice a week for a maximum period of 54 weeks employing various experimental designs. The ducks were infected with DHBV by i.v. inoculation of DHBV-positive sera within 24 h posthatch. The livers were examined histologically, immunohistochemically, and ultrastructurally, and the livers and sera were examined by molecular hybridization for DHBV DNA. AFB administration induced hepatocellular necrosis and marked biliary cell proliferation of the periportal areas, and finally liver cirrhosis. On short-term administration, the hepatocytes of DHBV-infected livers revealed a marked increase in incomplete particles of DHBV by immunostaining and electron microscopy, as compared to those without its administration. Long-term AFB administration provoked frequent nodular or cirrhotic changes. There was no significant increase in frequency of these changes in DHBV-positive ducks as compared to DHBV-negative ones. AFB administration induced hepatocellular carcinoma (HCC) in one DHBV-positive duck and in two DHBV-negative ducks. The HCC and cirrhotic livers revealed extrachromosomal but no integrated form of DHBV DNA by Southern blot hybridization analysis. Immunostaining demonstrated a heterogeneous distribution of DHBV from area to area in nodular and cirrhotic livers. Thus, AFB intoxication provoked various liver disorders independent of DHBV infection, and neither a cocarcinogenic effect of AFB and DHBV nor integration of viral DNA into the genome of neoplastic and nonneoplastic tissues was observed in the present experiments. Generally speaking, DHBV infection did not appear to accelerate hepatic disorders induced by AFB intoxication. However, AFB administration altered the DHBV in the liver in terms of its amount and distribution.

Contribution of aflatoxin B1 and hepatitis B virus infection in the induction of liver tumors in ducks.

The study of two major risk factors in the development of hepatocellular carcinoma, namely persistent hepatitis virus infection and exposure to dietary aflatoxins, has been hampered by lack of an experimental system. To this end we have used a Pekin duck model to examine the effect of congenital duck hepatitis B virus (DHBV) infection and aflatoxin B1 (AFB1) exposure in the induction and development of liver cancer. AFB1 was administered to DHBV infected or noninfected ducks at two doses (0.08 and 0.02 mg/kg) by i.p. injection once a week from the third month posthatch until they were sacrificed (2.3 years later). Two control groups of ducks not treated with AFB1 (one of which was infected with DHBV) were observed for the same period. Each experimental group included 13-16 ducks. Higher mortality was observed in ducks infected with DHBV and treated with AFB1 compared to noninfected ducks treated with AFB1 and other control ducks. In the groups of noninfected ducks treated with high and low doses of AFB1, liver tumors developed in 3 of 10 and 2 of 10 ducks; in infected ducks treated with the high dose 3 of 6 liver tumors were observed and none in the low dose of AFB1. No liver tumors were observed in the two control groups. Ducks infected with DHBV and treated with AFB1 showed more pronounced periportal inflammatory changes, fibrosis, and focal necrosis compared to other groups. All DHBV carrier ducks showed persistent viremia throughout the observation period. An increase of viral DNA titers in livers and sera of AFB1 treated animals compared to infected controls was frequently observed. No DHBV DNA integration into the host genome was observed, although in one hepatocellular carcinoma from an AFB1 treated duck, an accumulation of viral multimer DNA forms was detected. The metabolism of AFB1 in infected and noninfected duck liver was also examined. The study on the role of DHBV infection and AFB1 in the etiopathogenesis of liver tumors may help to clarify some of the basic mechanisms of carcinogenesis.

Hepatic neoplasms in aflatoxin B1-treated, congenital duck hepatitis B virus-infected, and virus-free pekin ducks.

To assess the effects of the combination of persistent hepadnavirus infection and chemical carcinogen exposure, aflatoxin B1 (AFB) was administered p.o. for 60 days to congenitally duck hepatitis B virus (DHBV)-infected and virus-free Pekin ducks, starting at 3 days of age, during a 28-month study. Hepatic neoplasia occurred only in AFB-dosed ducks. Hepatocellular carcinomas or biliary carcinomas occurred in 4 of 8 DHBV-infected and 3 of 4 DHBV-free ducks, and hepatocellular adenomas developed in 2 DHBV-infected AFB-dosed ducks that survived 20 months or longer. Altered foci of hepatocytes similar to those observed in chemical carcinogen-dosed rodents, characterized by enlarged eosinophilic hepatocytes or vacuolated cytoplasm, occurred in AFB-dosed ducks. Cells in foci or hepatic neoplasms did not contain histochemically detectable gamma-glutamyltranspeptidase but were distinguished from uninvolved parenchyma by altered glycogen content. Immunohistochemical staining indicated that DHBV core antigen persisted in liver, spleen, pancreas, and, to a lesser extent, kidney of most congenitally infected ducks up to 28 months of age. Hepatic neoplasms contained only patches of hepatocytes were detectable viral antigen. Southern blot analysis of restriction endonuclease-digested neoplastic and normal liver DNA revealed high molecular weight forms of DHBV DNA consistent with integration of viral DNA into the genome of hepatic neoplasms from 3 of 4 DHBV-infected ducks but not nontumorous liver. These findings indicate that AFB is a potent hepatic carcinogen in ducks and that persistent congenital DHBV infection did not contribute significantly to the emergence of hepatic neoplasia in ducks under these conditions.

Experimental neuropathology of chronic demyelination induced by a JHM virus variant (DS).

A small plaque variant of JHM virus has a markedly reduced ability to kill mice following intracerebral inoculation. Spinal cords of mice surviving 13 to 16 months following acute infection with this variant were examined ultrastructurally. Multiple subpial areas of demyelination in the anterior and lateral white matter were found in five of 13 mice. The lesions had more gliosis, fewer oligodendrocytes, and less remyelination than has been described following other infections with JHM virus. No conclusive evidence of active demyelination or viral-like particles was found. The pathogenesis of the lesions observed may be due to a persistent, attenuated infection of oligodendrocytes or to immunologic processes. These lesions were similar to chronic multiple sclerosis plaques. Therefore, this variant should prove to be a useful tool for studying the long-term effects of viral-induced demyelinating diseases.

Demyelination induced by murine hepatitis virus JHM strain (MHV-4) is immunologically mediated.

The neurotropic mouse hepatitis viruses (MHV), in particular strain JHM (JHMV or MHV-4), cause experimental central nervous system demyelination that pathologically resembles multiple sclerosis, an important human demyelinating disease. The mechanism of JHMV-induced demyelination remains unclear, though its tropism for oligodendrocytes had led to the belief that JHMV causes demyelination by direct lysis of these myelin-producing cells. However, several studies have also implicated the involvement of immune responses in the demyelinating process. In this communication, we present evidence that generalized immunosuppression with gamma irradiation prevents JHMV-induced demyelination, a finding that was not limited to a particular strain of JHMV or to one strain of mouse. In addition, significant paralytic-demyelinating disease was restored to infected, irradiated mice after the adoptive transfer of nylon wool nonadherent splenic cells and appeared to be restricted by the major histocompatibility complex (MHC). These observations indicate that the principal mechanisms of JHMV-induced demyelination are most likely immunopathological.

Effects of regional spinal X-irradiation on demyelinating disease caused by Theiler's virus, mouse hepatitis virus or experimental allergic encephalomyelitis.

The effects of X-irradiation on the course of chronic demyelinating disease were examined in mice with experimental allergic encephalitis (EAE), mouse hepatitis virus (MHV) or Theiler's virus (DAV) infection. One month after the induction of EAE or 2-16 months after inoculation of DAV, exposure of the cervical spinal cord to 20 Gy X-rays caused local exacerbation of disease activity but spinal irradiation did not affect MHV-induced demyelination. In EAE, there was a significant increase in the number of inflammatory cells in the irradiated part of the cord. Mice infected with DAV showed locally increased demyelination and axonal degeneration but no change in the titer of infectious virus within the cord. Thus in DAV infection, as in EAE, the exacerbation of disease seemed to be due to vascular or immunological factors rather than viral reactivation.

Effect of cyclosporin A on an experimental chronic viral infection of the central nervous system.

The effects of cyclosporin A (CsA) on neuropathological lesions induced by a chronic viral infection have been tested in the experimental model of the mouse hepatitis virus 3 (MHV3) infection. Daily injections of CsA (50 mg/kg) inhibited the expression of the MHV3-induced ependymitis, meningitis, hydrocephalus and vasculitis. The effect was preserved even if CsA treatment was initiated 15 days after virus infection but was lost if CsA treatment was given later on or for a shorter period of time. Viral titers in brains of chronically infected mice were not affected by CsA treatment. During the first week following MHV3 infection, CsA treatment increased both the percentage of acute death (31 vs. 10%) and the viral titers in brain and liver of infected mice. In this model, the timing of CsA treatment appeared critical for the balance between its beneficial effect on CNS lesions and the risk of increased acute mortality.

Viruses similar to hepatitis B virus (Icrons).

In 1971, on the basis of the unusual clinical and epidemiologic characteristics of hepatitis B virus, we postulated the existence of a class of viruses that we termed Icrons. An increased understanding of the molecular biology of hepatitis B virus resulted in the discovery, by Summers and his colleagues, of the woodchuck hepatitis virus. This virus is common in the North American woodchuck (Marmota monax) and is associated with primary cancer of the liver in this animal. Subsequently similar viruses were found in Beechey ground squirrels in California by Marion and her coworkers and domesticated (Pekin) ducks from the United States by Mason and his coworkers. In the latter the virus infects the liver and presumably is associated with disease of this organ. The discovery of additional viruses similar to hepatitis B virus in animals other than man and their association with cancer of the liver encourages the continuing search for other virus-cancer associations for which prevention methods might be effective.