Search

Background The cardiovascular phenotype is poorly characterized in treated pediatric hypertension. Cardiovascular magnetic resonance imaging (MRI) can be used to better characterize both cardiac and vascular phenotype in children with hypertension. Objective To use MRI to determine the cardiac and vascular phenotypes of different forms of treated hypertension and compare the results with those of healthy children. Materials and methods Sixty children (15 with chronic renal disease with hypertension, 15 with renovascular hypertension, 15 with essential hypertension and 15 healthy subjects) underwent MRI with noninvasive blood pressure measurements. Cardiovascular parameters measured include systemic vascular resistance, total arterial compliance, left ventricular mass and volumetric data, ejection fraction and myocardial velocity. Between-group comparisons were used to investigate differences in the hypertension types. Results Renal hypertension was associated with elevated vascular resistance (P≤0.007) and normal arterial compliance. Conversely, children with essential hypertension had normal resistance but increased compliance (P=0.001). Renovascular hypertension was associated with both increased resistance and compliance (P≤0.03). There was no difference in ventricular volumes, mass or cardiac output between groups. Children with renal hypertension also had lower systolic and diastolic myocardial velocities. Conclusion Cardiovascular MRI may identify distinct vascular and cardiac phenotypes in different forms of treated childhood hypertension. Future studies are needed to investigate how this may inform further optimisation of blood pressure treatment in different types of hypertension. Electronic supplementary material The online version of this article (10.1007/s00247-019-04393-6) contains supplementary material, which is available to authorized users.

Background Children with chronic kidney disease (CKD) have increased cardiovascular mortality. Identifying high-risk children who may benefit from further therapeutic intervention is difficult as cardiovascular abnormalities are subtle. Although transthoracic echocardiography may be used to detect sub-clinical abnormalities, it has well-known problems with reproducibility that limit its ability to accurately detect these changes. Cardiovascular magnetic resonance (CMR) is the reference standard method for assessing blood flow, cardiac structure and function. Furthermore, recent innovations enable the assessment of radial and longitudinal myocardial velocity, such that detection of sub-clinical changes is now possible. Thus, CMR may be ideal for cardiovascular assessment in pediatric CKD. This study aims to comprehensively assess cardiovascular function in pediatric CKD using CMR and determine its relationship with CKD severity. Methods A total of 120 children (40 mild, 40 moderate, 20 severe pre-dialysis CKD subjects and 20 healthy controls) underwent CMR with non-invasive blood pressure (BP) measurements. Cardiovascular parameters measured included systemic vascular resistance (SVR), total arterial compliance (TAC), left ventricular (LV) structure, ejection fraction (EF), cardiac timings, radial and longitudinal systolic and diastolic myocardial velocities. Between group comparisons and regression modelling were used to identify abnormalities in CKD and determine the effects of renal severity on myocardial function. Results The elevation in mean BP in CKD was accompanied by significantly increased afterload (SVR), without evidence of arterial stiffness (TAC) or increased fluid overload. Left ventricular volumes and global function were not abnormal in CKD. However, there was evidence of LV remodelling, prolongation of isovolumic relaxation time and reduced systolic and diastolic myocardial velocities. Conclusion Abnormal cardiovascular function is evident in pre-dialysis pediatric CKD. Novel CMR biomarkers may be useful for the detection of subtle abnormalities in this population. Further studies are needed to determine to prognostic value of these biomarkers. Electronic supplementary material The online version of this article (10.1186/s12968-018-0444-0) contains supplementary material, which is available to authorized users.

Background—Conventional cardiopulmonary exercise testing can objectively measure exercise intolerance but cannot provide comprehensive evaluation of physiology. This requires additional assessment of cardiac output and arteriovenous oxygen content difference. We developed magnetic resonance (MR)–augmented cardiopulmonary exercise testing to achieve this goal and assessed children with right heart disease.Methods and Results—Healthy controls (n=10) and children with pulmonary arterial hypertension (PAH; n=10) and repaired tetralogy of Fallot (n=10) underwent MR-augmented cardiopulmonary exercise testing. All exercises were performed on an MR-compatible ergometer, and oxygen uptake was continuously acquired using a modified metabolic cart. Simultaneous cardiac output was measured using a real-time MR flow sequence and combined with oxygen uptake to calculate arteriovenous oxygen content difference. Peak oxygen uptake was significantly lower in the PAH group (12.6±1.31 mL/kg per minute;P=0.01) and trended toward lower in the tetralogy of Fallot group (13.5±1.29 mL/kg per minute;P=0.06) compared with controls (16.7±1.37 mL/kg per minute). Although tetralogy of Fallot patients had the largest increase in cardiac output, they had lower resting (3±1.2 L/min per m2) and peak (5.3±1.2 L/min per m2) values compared with controls (resting 4.3±1.2 L/min per m2and peak 6.6±1.2 L/min per m2) and PAH patients (resting 4.5±1.1 L/min per m2and peak 5.9±1.1 L/min per m2). Both the PAH and tetralogy of Fallot patients had blunted exercise–induced increases in arteriovenous oxygen content difference. However, only the PAH patients had significantly reduced peak values (6.9±1.3 mlO2/100 mL) compared with controls (8.4±1.4 mlO2/100 mL;P=0.005).Conclusions—MR-augmented cardiopulmonary exercise testing is feasible in both healthy children and children with cardiac disease. Using this novel technique, we have demonstrated abnormal exercise patterns in oxygen uptake, cardiac output, and arteriovenous oxygen content difference.

A protein component which accounts for 10% of the total protein of isolated cell membranes of Micrococcus lysodeikticus has been partially characterized. The protein can be released into solution from the membrane complex and in this “soluble” state it possesses Ca++-dependent adenosine triphosphatase activity. This enzyme has been purified about 50-fold, by a procedure based on its association-dissociation properties with the membrane. The progressive purification and degree of homogeneity have been demonstrated by examining the various stages in the soluble ATPase preparation by disc electrophoresis and by gel diffusion against membrane antiserum. On treatment with 8 M urea or 1% sodium dodecyl sulfate followed by disc electrophoresis, clear evidence of dissociation was obtained, indicating the existence of subunits. The purified protein has a sedimentation coefficient of 14–15 S as determined by sucrose gradient centrifugation or analytical ultracentrifugation. A sedimentation coefficient of 3.5 S has been estimated for the main product of the protein dissociated by sodium dodecyl sulfate again an indication of the dissociation of the protein into subunits, probably identical on the basis of molecular weight. The “soluble” enzyme was activated by Ca++, optimum activation being observed at ATP/Ca++ ratio of 1/1 and pH 7.5 under which conditions 50–60 μmoles of ATP were hydrolyzed per mg protein per 10 min at 37°. The ATPase was quite stable at 4°. The Ca++-dependent ATPase activity was inhibited by ADP, sodium azide, and Mg++. When the membrane-ATPase complex remains stable, no hydrolytic activity for ATP is shown unless trypsin activation. Our system shows similarities with the “allotopic” properties described for some mitochondrial enzymes. From this and previous work in our laboratory a close relationship between the “stalked particles” seen in M. lysodeikticus cell membranes by electron microscopy and ATPase appears to be evident.

We have evaluated four hepatitis E virus (HEV) specific antibody assays, using sequential samples taken from 86 rhesus monkeys at intervals for up to 86 weeks after they had been infected with different doses of HEV. The animals are a common experimental model of hepatitis E. The large collection of sequential samples used avoids uncertainties encountered in previous studies regarding the precise infection status of study subjects and minimizes bias due to the individuality of response to infection. One assay (YES IgG) was produced with synthetic peptides; the others (E2 IgM, E2 IgG, and GL IgG) were produced with recombinant antigens. The results were compared with the viral RNA contents of the serum and stool samples and the occurrence of these virological and immunological markers in the course of the infection was temporally related to the development of hepatitis. Diagnostic utility of the markers was assessed according to their response rates and prevalence at different times in the course of infection. All the animals produced E2 IgG and developed viremia and all but one also produced E2 IgM and excreted the virus in stool, whereas response rates for the other antibodies were lower and decreased with virus dose. Hepatitis occurred over a period of 4 weeks between 3 and 7 weeks after infection. Virological activity occurred mainly during the incubation period and the prevalence of viral markers declined rapidly after the onset of hepatitis. Production of the E2 antibodies immediately preceded the onset of hepatitis, and this was followed about one week later by production of the other antibodies. Seroprevalence E2 IgM reached a peak value 3 weeks after the onset of hepatitis, whereas seroprevalence of GL IgG and YES IgG peaked after the disease had subsided. E2 IgG persisted in all animals for the entire duration of the experiment of up to 86 weeks and possibly beyond and, thus, can serve as a useful epidemiological marker of HEV infection.

We assessed 5 EBV specific assays for their capacity to effect serologic diagnosis of suspected NPC. The assays were the immunofluorescent assays, VCA IgA and EA IgA, the enzyme-linked immunosorbent assays specific for EBNA 1 IgA or zta IgG and an EBV DNA assay. Serum samples were taken from 218 symptomatic NPC patients presenting consecutively at a public hospital in Hong Kong, 51 of whom were subsequently diagnosed as having NPC; 4 had EBV-associated lung cancer with similar serology as NPC. The remaining patients included 23 who had other cancers and 140 who had other diseases. Objectives of serodiagnosis under such clinical settings, therefore, are to both exclude and predict a diagnosis of NPC. None of the assays individually can meet both requirements adequately, however. The difficulty was best overcome by combining EBNA 1 IgA and zta IgG. It was shown that 68.3% of the patients gave a confirmed test results, negative or positive, by both tests. A confirmed negative result was associated with a negative predictive value of 99.1%, providing a clear indication to exclude a diagnosis of NPC; a confirmed positive result was associated with a positive predictive value of 86.8%, providing a clear indication to proceed with diagnostic work-up of NPC. The remaining patients gave equivocal test results, being positive for one or the other test, which were associated with a positive predictive value of 43.3% and 24.2%, respectively. © 2003 Wiley-Liss, Inc.

A 23 kDa peptide of the major structural protein of the hepatitis E virus (HEV) expressed in E. coli was found to naturally interact with one another to form homodimers and the peptide was recognized strongly in its dimeric form by HEV reactive human sera. To determine if the peptide may confer protection against HEV infection, three monkeys were immunized with the purified peptide and three were given placebo. Both groups of animals were challenged with 10(5) genome equivalent dose of the homologous strain of HEV. All control animals excreted the virus for 10-12 days beginning 5 days after the infection. The viral genome was also present in the peripheral blood monocyte (PBMC) samples from two animals, but it was not detected in the plasma samples from any of the animals. The infection in two control animals was accompanied by HEV seroconversion. Immunization was found to abrogate HEV stool excretion in two animals and reduced the viral excretion to one day in the third. None of the immunized animals showed detectable HEV in plasma or PBMC samples nor did the animals showed evidence of HEV seroconversion. These results suggested that immunization with the bacterially expressed peptide may prevent experimental infection of primates with the homologous strain of HEV.

Human cytomegalovirus (HCMV), human herpesvirus-6 (HHV-6), and human herpesvirus-7 (HHV-7) DNA in peripheral blood leukocytes (PBL) of 61 bone marrow transplant recipients was monitored weekly during the first 12 weeks post-transplantation by a nested polymerase chain reaction (PCR). Thirty-seven (61%), 17 (28%), and 32 (53%) of patients had one or more PBL specimens positive for HCMV, HHV-6 or HHV-7 DNA, respectively. HHV-7 DNA in PBL during the early post-transplant period was associated with a longer time to neutrophil engraftment (mean 28.8 days vs 19.8 days; P = 0.01). In two patients who failed to engraft, HHV-6 DNA and HHV-7 DNA was detected in plasma and PBL, respectively, early in their post-transplant period. Patients with HCMV disease were more likely to have concurrent HHV-7 DNA in PBL prior to onset of disease than were patients with asymptomatic HCMV infection, suggesting that HHV-7 may be a cofactor in the progression from HCMV infection to HCMV disease. In the 17 patients (179 specimens) in whom viral DNA in plasma was studied (in addition to PBL), a positive result was found only in 3. In each, viral DNA in plasma appeared to correlate with clinically significant disease. HHV-7 DNA in plasma was associated with encephalitis in an allograft recipient.

To evaluate applications of highly purified recombinant EBV DNAase in the diagnosis and prognosis of NPC, we tested sera from patients with NPC, other EBV-associated diseases and EBV-seropositive and -seronegative healthy subjects by immunoblotting and DNAase inhibitory assay. The results were compared with those obtained by the conventional immunofluorescence assays against the EBV-specified early antigens and capsid antigens. The antigenic specificity of the immunoblotting assay for IgG antibody against the viral enzyme, but not that for the IgA antibody, was correlated with DNAase-inhibitory activity of the sera and their titers of IgG antibodies against the viral early antigens. Purified IgA as well as IgG from NPC sera inhibited enzyme activity with similar efficiency. The use of highly purified viral DNase has increased the sensitivity of detection of the corresponding antibodies by immunoblotting, with the IgG antibody being detected in all but one, and IgA antibody in all but 2, of the 174 NPC sera tested. The IgG antibody was also commonly detected in the other groups of control sera, while the IgA antibody was detected in about 10% of African Burkitt's lymphoma and Algerian Hodgkin's lymphoma patients and less than 3% of the other control subjects. These results suggest that IgA antibody against recombinant EBV DNAase may be useful in the diagnosis of NPC, but the level of this antibody did not appear to be related to clinical stages of this cancer.

An extended follow-up study of hepatitis C virus (HVC) infection was conducted in Guangzhou and its nearby regions on patients hospitalized with acute hepatitis. Routine screening of blood donors for HCV was not yet instituted at the time of this study. HCV was found to be a common cause of the disease, and the infection had a close association with recent histories of blood transfusion. Sequential sera obtained from patients during hospitalization and after discharge were tested for the presence of HCV antibodies by the first and the second generation of commercial test kits, for levels of alanine aminotransferase (ALT), and for the presence of HCV-RNA. The development of HCV antibodies in some of the patients may be delayed for protracted period following clinical onset. HCV-RNA was only intermittently detectable both before and after seroconversion. Six of 33 patients studied showed seroreversion and 5 of them were accompanied by loss of HCV-RNA and serum ALT returned to normal levels. The disease persisted in 24 of 27 patients without seroreversion, accompanied by intermittent detection of HCV-RNA throughout the protracted course of the infection. Our results indicate that both EIA for detection of HCV antibodies and PCR for serum HCV-RNA should be used in combination for reliable diagnosis of HCV infection and screening of blood for transfusion.

A new immunoglobulin M (IgM) enzyme-linked immunosorbent assay (ELISA) based on the recombinant Epstein-Barr virus (EBV) matrix protein was developed. Compared to indirect immunofluorescence for the detection of IgM antibody to the EBV capsid antigen on clinical specimens, the sensitivity and specificity of the new IgM ELISA were 96 and 96%, respectively.

It was evaluated its antigenicity, immunogenicity and efficacy of a candidate recombinant hepatitis E virus (HEV) vaccine, referred hitherto as HEV 239 vaccine. The vaccine peptide has a 26 amino acids extension from the N terminal of another peptide, E2, of the HEV capsid protein, which has been shown to protect monkeys against HEV infection previously. The vaccine peptide is similar as E2 in that: first, the vaccine peptide migrates predominantly as dimer in SDS-PAGE and it is dissociated into monomers by heating; second, its dimeric form of which predominantly recognized by HEV reactive human serum; and third, it shows the same pattern of reaction as E2 with a panel of eight monoclonal antibodies that had been raised against E2. In contrast to E2, the vaccine peptide aggregates to form particles of 13 nm mean radius, and consequently, it is more than 240 times more immunogenic than E2. Using alum as adjuvant, immunizing dose determined in mice was 80-250 ng for the vaccine and >60 microg for E2. Rhesus monkeys twice vaccinated with a 10 microg or a 20 microg formulation of this vaccine showed essentially the same antibody response, whereas the response to a 5 microg formulation was delayed but reached similar antibody levels. All the three vaccine formulations afford complete protection against infection with 10(4) genome equivalent dose of the homologous genotype 1 virus. At higher virus dose of 10(7), the same vaccine formulation partially protected against the infection and completely protected against hepatitis. The efficacy of the vaccine was essentially the same for the homologous genotype 1 virus and heterologous genotype 4 virus.

The dimeric form of the recombinant peptide (E2), comprising amino acid 394-606 of the capsid protein of hepatitis E virus (HEV), is strongly recognized by HEV reactive human serum, and when used as a vaccine, it protects rhesus monkeys against experimental HEV infection. In this work, the relationship of E2 to HEV has been probed using three murine monoclonal antibodies, 8C11, 13D8 and 8H3, all of which react predominantly against the E2 dimer, and can effect immune capture of the virus as well. 8C11 and 8H3 were further found to neutralize HEV infectivity in animals. Cross-blocking patterns between these antibodies discerned two spatially separate antigenic domains, one identified by 8C11 and 13D8, and the other, by 8H3. Kinetic studies using BIAcore biosensor suggest that the epitope to which 8H3 is directed is partially masked, and thus has limited access by the native antibody. However, this is not the case with the smaller Fab. Access to the 8H3 epitope was enhanced by the binding of 8C11, and inhibited by the binding of 13D8 to a distal site on the peptide. Similar to the effects of binding 8H3 to E2, 8C11 was found to enhance immune capture by 8H3, while 13D8 was inhibitory. Moreover, 8C11 and 8H3 act synergistically to neutralize HEV infectivity. The parallel cross-reaction patterns that these antibodies exhibit against the peptide and the virus, respectively, implicate two interacting conformationally dependent neutralization sites on the HEV particle. These sites might cooperate in the adsorption and penetration of the HEV virus.

The geographically constrained distribution of Epstein-Barr virus (EBV)-associated nasopharyngeal carcinoma (NPC) in southeast Asian populations suggests that both viral and host genetics may influence disease risk. Although susceptibility loci have been mapped within the human genome, the role of viral genetics in the focal distribution of NPC remains an enigma. Here we report a molecular phylogenetic analysis of an NPC-associated viral oncogene, LMP1, in a large panel of EBV isolates from southeast Asia and from Papua New Guinea, Africa, and Australia, regions of the world where NPC is and is not endemic, respectively. This analysis revealed that LMP1 sequences show a distinct geographic structure, indicating that the southeast Asian isolates have evolved as a lineage distinct from those of Papua New Guinea, African, and Australian isolates. Furthermore, a likelihood ratio test revealed that the C termini of the LMP1 sequences of the southeast Asian lineage are under significant positive selection pressure, particularly at some sites within the C-terminal activator regions. We also present evidence that although the N terminus and transmembrane region of LMP1 have undergone recombination, the C-terminal region of the gene has evolved without any history of recombination. Based on these observations, we speculate that selection pressure may be driving the LMP1 sequences in virus isolates from southeast Asia towards a more malignant phenotype, thereby influencing the endemic distribution of NPC in this region., published_or_final_version

Epstein-Barr virus (EBV)-specific immunoblot analysis was used to reveal the molecular diversity of immunoglobulin (Ig) G and IgA antibody responses against Epstein-Barr nuclear antigen (EBNA), early antigen (EA), and viral capsid antigen (VCA) in serum samples from patients with nasopharyngeal carcinoma (NPC) and control subjects, by use of immunofluorescence assay (IFA). Control donors (n=150) showed IgG responses to few EBV proteins--VCA-p18, VCA-p40, EBNA1, and Zebra--and sporadically weak IgA reactivity to EBNA1 and VCA-p18. Patients with NPC stage 1 (n=6) had similar response patterns. Patients with NPC stage 2-4 (n=132) showed significantly more diverse IgG and IgA responses to EA and VCA proteins--VCA-p18/-p40, EBNA1, Z-encoded broadly reactive activator, and EAd-p47/54, -DNAse, -thymidine kinase, and -p138. No correlation was found between IFA titers and the number of EBV proteins recognized by IgG or IgA. Our results reveal dissimilarity between EBV polypeptides recognized by IgG and IgA antibodies, which suggests independent B cell triggering events.

Detection of Epstein-Barr Virus (EBV) DNA by PCR in serum had a sensitivity of 80%, a specificity of 94%, and positive and negative predictive values of 95 and 79%, respectively, for the diagnosis of primary EBV infection. We suggest that this is a useful addition to the panel of tests used for this purpose., published_or_final_version

A 23 kDa peptide locating to amino acid residues 394 to 604 of the major Hepatitis E Virus (HEV) structural protein was expressed in E. coli. This peptide was found to interact naturally with one another to form homodimers and it was recognized strongly and commonly in its dimeric form by HEV reactive human sera. The antigenic activity associated with the dimeric form was abrogated when the dimer was dissociated into monomer and the activity was reconstituted after the monomer was re-associated into dimer again. The dimeric form of the peptide elicited a vigorous antibody response in experimental animals and the resulting antisera were found to cross-react against HEV, effecting an efficient immune capture of the virus. These results attributed the antigenic activity associated with the dimeric form of the peptide to conformational antigenic determinants generated as a result of interaction between the peptide molecules. It is suggested that some of these antigenic determinants may be expressed by the HEV capsid and raised the possibility of this bacterially expressed peptide as an HEV vaccine candidate.

Background: Epstein–Barr virus (EBV) has been shown to be the cause of infectious mononucleosis (IM) and has more complicated associations with several malignant diseases. These EBV associated diseases provide a strong incentive for the development of an EBV vaccine. Most primary EBV infection during infancy and early childhood is mild or subclinical. Little is known about its infection in infancy. The pattern of EBV serological response during infancy may be important for vaccine management. Objectives: this study has served to clarify the epidemiology and serology of primary EBV infection during early infancy. Study design: longitudinal serum samples from 66 Hong Kong infants were tested for EBV antibodies by immunofluorescence. Cord blood and sequential serum samples from these infants were taken at birth and then at 4-month intervals up to 2 years of age. Results: maternal antibodies were present at different levels in all cord blood specimens and in serum samples of 8 infants at 4-month of age. Evidenced by VCA-IgG seroconversion, 60.6% (40/66) infants were infected during the first 2 years of life. One episode occurred before 8 months of age but, thereafter and for the remaining 16 months of follow-up until the infants were 2 years of age, the infection occurred at essentially a constant rate affecting about 20% of the remaining seronegative infants every 4 months. Conclusions: the abrupt onset of the infection after a delay of 8 months is a remarkable feature of primary EBV infection during infancy, which implicates a protective role for maternal antibodies. Persisting maternal antibodies may additionally serve to contain the infection once it occurred. This may partly explain why, unlike during adolescence, primary EBV infection early in life is usually asymtomatic.

The age-related prevalence of antibodies to herpesviruses was compared in England and Hong Kong. Altogether 327 sera from England and 266 sera from Hong Kong were tested for antibodies to herpes simplex virus (HSV), varicella-zoster virus (VZV), Epstein-Barr virus (EBV), cytomegalovirus (CMV), and human herpesvirus 6 (HHV-6). Herpesvirus infections were common in both countries but generally were acquired earlier and were more prevalent in Hong Kong. Over 90% of children in Hong Kong were infected with VZV, EBV, and HHV-6 by 8 years of age. HSV and CMV were the least prevalent childhood infections in both countries, although, 30–40% of babies in Hong Kong became infected during the first year of life. CMV infections were rare throughout childhood in the English cohort. Overcrowding and early attendance at kindergarten may aid more efficient and earlier transmission of herpesvirus in Hong Kong. The high prevalence of CMV in particular may have implications for the management of young pregnant women and organ transplant recipients in Hong. © 1994 Wiley-Liss, Inc.

Epstein Barr Virus (EBV) receptor activity in cell free extracts is operationally defined as one which causes a reduction in the effective concentration of the early antigen inducing particles of an EBV preparation when the latter is preincubated with the extract before infection. Such activity was detected in the surface extract of Raji cells and to a lesser extent in that of BJAB cells, both of which are B lymphoblastoid cells that are susceptible to infection with EBV. Receptor activity was not detected in similar extracts of P3HR-1 cells and human diploid fibroblasts neither of which are known to be susceptible to EBV infection. Receptor activity in the Raji cell extract was found to be associated with membranous structures. This may have rendered the activity resistant to treatment with trypsin and sonication. The activity was however abrogated if the extract was exposed to neutral detergent. Binding of receptor activity was observed when Raji cell extract was chromatographed on a column of immobilized EBV. Subsequent electrophoretic analysis showed however that this procedure did not result in an appreciable purification of the receptor activity. Neutral detergent treated extract was similarly chromatographed. The resulting eluates did not contain detectable receptor activity but were less heterogeneous in protein content as compared with that of the original extract. It is not certain at present if these EBV binding proteins are involved in the receptor activity of the extract.

We have raised a murine IgM monoclonal antibody (MA6) against immunoprecipitate obtained by reacting the serum of an NPC patient with extract of the Burkitt lymphoma cell line, Raji. It reacts against an antigen (BLCa) which is broadly represented on human B lymphoid tissues and cell lines but is different from the functional B-cell markers such as surface immunoglobulins, Ia products, Fc and complement receptors. BLCa was found to occur on lymphoid cell lines representing all stages of B-cell differentiation. These included the pre-B and null-cell lines, Nalm 6 and Reh, the EBV-transformed lymphoid cell lines and the myeloma cell line. MA6 was reactive against all the Burkitt lymphoma cell lines, whether or not these harbored the EBV genomes, with the exception of P3HR-I. The antibody was found to be selectively reactive against a proportion of peripheral blood B lymphocytes and to stain the B-cell-rich primary follicles and mantle zones of secondary follicles in the lymph node. However, MA6 was not reactive against cell lines of T-lymphocyte, myeloid, monocyte fibroblast or epithelial origin. It did, nevertheless selectively stain tumor cells from a variety of carcinoma tissues originating from different anatomical sites, including the nasopharynx.

Ionic shock treatment in the presence of 10% glycerol is an efficient and selective method for extracting cell surface components from Raji cells and effecting the solubilization of up to 22 polypeptides. The majority of these shock-released polypeptides are accessible to lactoperoxidase radioiodination. Sera from rabbits immunized against these soluble extracts are reactive against Raji cell surface as indicated by indirect membrane immunofluorescence and agglutination assays.

Egg lecithin at 200 μg/ml or greater was found to abrogate blastogenic responses of human leukocyte cultures stimulated with phytohemagglutinin (PHA), concanavalin A (ConA), purified protein derivatives of tuberculin (PPD), candidin or streptokinase and streptodornase (SKSD) while leukocyte aggregation mediated by PHA, however, occurred irrespective of the presence of the lipid. Inhibition of PHA-mediated responses occurred when the lecithin was added simultaneously with the mitogen but the extent of inhibition decreased with delay in the addition of the lipid. When added 48 h after their exposure to PHA, the presence of lecithin did not change the pattern of [3H]TdR incorporation. Prior sonic treatment of an aqueous suspension of lecithin was found to render the lipid more effective in arresting lymphocyte responses than the unsonicated control. In entity, these results indicated that the lipid did not affect the viability of the leukocyte cultures nor did it seem that the lipid may interfere with the binding of the mitogen per se. It was concluded therefore, that inhibition by lecithin may most likely occur early at a time following binding of the mitogen but before the cells are committed to cellular division and it seems that the plasma membrane might be a likely site of action of the lipid.

1. The adenosine analogue toyocamycin caused an inhibition of ribosomal RNA synthesis in cultures of rabbit kidney cells at concentrations as low as 0.1 μg/ml. The synthesis of other species of RNA in the ribosome fraction was relatively resistant to toyocamycin. Amino acid incorporation in intact cells was not suppressed at concentrations as high as 10.0 μg/ml. 2. At a concentration of 10 μg/ml, toyocamycin suppressed interferon synthesis stimulated with polyinosinic · polycytidylic acid (poly(I) · poly(C)). Lower concentrations of the drug that still inhibited the synthesis of ribosomal RNA not only did not suppress, but actually enhanced, the interferon yield. 3. In order to allow incorporation of toyocamycin into interferon mRNA, cultures were stimulated with poly(I) · poly(C) and then incubated for 3 h in the presence of toyocamycin (1 μg/ml) and cycloheximide. Cycloheximide was included to suppress translation of interferon mRNA, while allowing its transcription to take place. Interferon synthesis in toyocamycin-treated cells following the reversal of cycloheximide action was found to be more temperature sensitive than in control cells. In addition, interferon made in toyocamycin-treated cells was more rapidly inactivated at 56 °C than control interferon.

Low to moderate doses of cycloheximide had a stimulatory effect on interferon production in rabbit kidney cell cultures treated with double-stranded polyinosinate-polycytidylate (poly I:poly C). A very marked stimulation occurred in the presence of a dose of cycloheximide inhibiting amino acid incorporation into total cellular protein by about 75%. Higher doses of cycloheximide caused a shift in interferon release towards later intervals and a gradual decrease in the overall degree of stimulation. An even greater increase in the amount of interferon produced was observed if cells were treated with cycloheximide for only 3 to 4 hr immediately after their exposure to poly I:poly C. Under the latter conditions, a rapid burst of interferon production occurred after the reversal of cycloheximide action. Treatment with a high dose of actinomycin D before the reversal of cycloheximide action caused a further increase and a marked prolongation of interferon production. It is postulated that inhibitors of protein synthesis suppress the accumulation of a cellular regulatory protein (repressor) which interacts with the interferon messenger ribonucleic acid mRNA and thereby prevents its translation. Therefore, active interferon mRNA can apparently accumulate in rabbit kidney cells which, after exposure to poly I:poly C, are kept in the presence of an inhibitor of protein synthesis. Some of this accumulated interferon mRNA can be translated during a partial block of cellular protein synthesis, but its most efficient translation occurs after the reversal of the action of the protein synthesis inhibitor.

To facilitate the routine identification of salmonellae and detailed studies of their lipopolysaccharides, we raised murine monoclonal antibodies against these organisms. We raised an immunoglobulin G1 antibody, MO2, which is specific for factor O2. By immunoblotting following sodium dodecyl sulfate-polyacrylamide gel electrophoresis, MO2 was shown to bind only the lipopolysaccharide of Salmonella paratyphi A, giving a ladderlike reaction pattern with regularly spaced reactive bands. MO2 did not react against lipopolysaccharides of other Salmonella serogroups, including those of serogroups B, C, D, E and L. Since the lipopolysaccharides of Salmonella serogroups A, B, D, and E are similar except for the presence of paratose in serogroup A organisms, this dideoxyhexose is therefore believed to be the immunodominant epitope for MO2. Consistent with the latter contention was the finding that periodate oxidation of the S. paratyphi A lipopolysaccharide did not destroy its antigenicity for MO2. In a slide agglutination test, MO2 was found to react specifically against all 12 clinical isolates of S. paratyphi A but not against 98 isolates of other salmonellae or 74 isolates of other bacteria and Candida albicans., link_to_OA_fulltext

To form heterocomplexes, human diploid fibroblast interferon was reacted with an excess of human immunoglobulin in the presence of an optimal concentration of glutaraldehyde. There was virtually no loss of interferon activity due to this treatment and the resulting heterocomplexes possessed the biological properties of both native interferon and immunoglobulin molecules. The apparent molecular weights determined by gel electrophoresis in the presence of sodium dodecyl sulfate were 20,000 for native interferon and 0.5 to 1 X 10(6) for the heterocomplex. Native interferon injected into mice intramuscularly had a serum half life of about 5 hours. However, serum interferon was not detected in mice receiving a similar dose of the heterocomplex.

Antigenic content of a Raji cell surface extract (TS) was studied by sequential abosrption of rabbit antiserum to this extract with different cells. Anti-TS was first allowed to react exhaustively with human blood cells followed by HeLa cells and human diploid fibroblasts. The residual reactivity was studied by stepwise absorption with an EBV-negative B lymphoblastoid cell line, BJAB, and then with B1--19, a clone derived from EBV transformed BJAB cells. These processes delineated three kinds of reactivities in anti-TS having specificities for antigens on B blasts, EBV-determined cell surface antigen(s) and antigens that are exclusive to Raji cells, respectively. These results are compatible with the heterogeneous protein content of TS and with the results of our earlier in vitro cell-mediated immunity studies using a similar preparation of Raji cell TS.

Publisher Summary Interferons are a class of protein capable of suppressing viral replication in competent animal cells. They are produced by a variety of animal cells in response to stimulation with a fairly wide range of materials. This characteristic cell-mediated antiviral action of interferons has proved to be the most useful parameter for the definition of interferons. This chapter reviews information on the general aspects of interferon production and action. It also focuses on the known physicochemical properties of interferons and control of the cellular synthesis of interferons. The chapter also discusses interferon induction by viruses and other agents. Although interferons had once been considered specific products of cell-virus interaction, more recent evidence shows that interferon production by animal cells can also be stimulated by a number of other agents. The still growing list of nonviral interferon inducers includes complex infectious agents, as well as relatively simple chemicals. The relationship between the structure and activity of polynucleotide interferon inducers is also discussed in the chapter.