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Identification and synthesis of an antigenic determinant common to human chorionic somatomammotropin and human growth hormone.

Human chorionic somatomammotropin (hCS) and human growth hormone (hGH), despite their different biological activities, show a remarkable degree of homology in their primary structure, which explains their immunological cross-reactivity. On the basis of the hydrophilicity profiles, we predicted that the sequence 165-174 would correspond to an antigenic determinant common to both hormones. The sequence 165-174 was synthesized in the solid phase and the ability to bind antibodies to hCS was tested by a radioimmunoassay at each step of the synthesis, without detaching the peptide from the resin. We found that the immunological sequence able to bind antibodies to hCS is that corresponding to 167-174. In similar experiments, we showed that the sequence 166-174 is able to bind antibodies to hGH. In the plaque-forming cell test using the synthetic fragment 166-174 bound to sheep red blood cells, we also observed that the ratio between the number of cells forming antibodies to hCS and to the 166-174 peptide was always between 0.45 and 0.73, thus suggesting that the 166-174 peptide represents a major determinant of hCS.

Amino Acid Sequence↗

Identification of novel cps locus polymorphisms in nontypable group B Streptococcus.

Group B Streptococcus (GBS) is an important pathogen responsible for a variety of diseases in newborns and the elderly. A clinical GBS isolate is considered nontypable (NT) when serological methods fail to identify it as one of nine known GBS serotypes. Eight clinical isolates (designated A1-A4, B1-B4) showed PFGE profiles similar to that of a GBS serotype V strain expressing R1, R4 surface proteins. These unique isolates were further characterized by immunologic and genetic methods. Rabbit sera to isolates A1 and A2 reacted weakly with concentrated HCl extracts of A1-A4 isolates, but not with those of B1-B4 isolates. In addition, a type V capsular polysaccharide (CPS) inhibition ELISA revealed that cell wall extracts from isolates A1-A4, but not from B1-B4, expressed low but measurable amounts of type V CPS. Molecular serotyping with PCR analysis showed that all eight isolates contained a type V-specific CPS gene (cpsO) and harboured the gene encoding the surface protein Alp3. Multilocus sequence typing identified isolate A1 as belonging to a new sequence type (ST) designated ST-173, whereas the other seven isolates keyed to ST-1. Sequencing of the 18 genes (17 736 bp) in the cps locus showed that each NT isolate harboured one to three unique polymorphisms, and also identified an IS1381 element in cpsE of the B4 isolate. Collectively, genetic and immunologic analyses revealed that these NT isolates expressing R1, R4 proteins have a genetic profile consistent with that of type V, an emergent, antigenically diverse and increasingly prevalent GBS serotype.

Animals↗

Antigen non-specific immunological tests in progressive sensorineural hearing loss.

The purpose of the present study was to determine the validity of antigen non-specific immunological tests in order to develop a risk-clinical profile of patients with immune-mediated inner ear pathology. The study was carried out on 38 patients with progressive sensorineural hearing loss and a control group of 26 subjects. Serological testing was used to determine cell activation complexes. In patients with hearing loss, T-lymphocyte subpopulations were found to have an increase in cytotoxic T-cells when compared to T-helper, but only for interleukin-2 receptor expression. This difference was significant between patients and the control group. Present findings suggest that T-lymphocyte "early" marker is of particular interest for developing a risk-clinical profile of patients with sensorineural progressive hearing loss of immune origin.

Adolescent↗

[Cellular immune reactions in the victims of explosive injuries to the skull and brain].

Specific features in the immunological status of 49 wounded with explosion injuries of neurosurgical profile are as follows: in light injuries--it is hyperergic character of the immune system and its inadequacy to the injuries of a middle degree it is a distinct parallelism between the dynamics of the immune status indices and the functional state of the CNS depending on the severity of the injury-an extremely flabby with the inhibiting of main immunological indices at a low but permanent level.

Adolescent↗

Salvage therapy with abacavir and other reverse transcriptase inhibitors for human immunodeficiency-associated encephalopathy.

BACKGROUND: HIV-associated encephalopathy (HIV-AE) is a severe neurologic condition that affects HIV-infected children. The potential benefit of antiretroviral (ARV) agents with good cerebrospinal fluid (CSF) penetration remains to be defined. Abacavir (ABC) achieves good CSF concentrations and studies of high-dose ABC showed benefit in adults with HIV dementia. The present study evaluated the safety and virologic, immunologic and neuropsychological responses of an ARV regimen including high-dose ABC in children with HIV-AE. METHODS: Children between 3 months and 18 years old and abacavir-naive with HIV-AE and virologic failure were eligible. RESULTS: : Seventeen children (16 ARV-experienced) were enrolled and 14 children completed 48 weeks of therapy. The overall tolerability was good; 2 children had a possible hypersensitivity reaction. At week 48, 53% and 59% of the children achieved HIV RNA levels below the limit of quantitation in plasma and CSF, respectively. The median (25%-75% range) change of HIV RNA from baseline to week 48 was -2.29 (-0.81 to -2.47) log10 copies/mL in plasma and -0.94 (0 to -1.13) log10 copies/mL in CSF. The mean increases in CD4 (+/-standard error of mean) cell count and CD4% were 427 (+/-169) cells/mm and 8% (+/-2), respectively. Concentrations of soluble tumor necrosis factor receptor II were reduced in plasma and CSF. Children less than 6 years of age demonstrated significant neuropsychological improvement at week 48. CONCLUSIONS: In the present study with a limited number of children, highly active ARV therapy including high-dose ABC showed a safety profile similar to standard dose ABC and provided clinical, immunologic and virologic response in children with HIV-AE at week 48. Children less than 6 years of age also demonstrated significant neuropsychological improvement.

AIDS Dementia Complex↗

Characterization and profiling of hepatic cytochromes P450 and phase II xenobiotic-metabolizing enzymes in beluga whales (Delphinapterus leucas) from the St. Lawrence River Estuary and the Canadian Arctic.

Cytochromes P450 (CYP, phase I) and conjugating (phase II) enzymes can be induced by and influence the toxicokinetics (metabolism) and toxicity of xenobiotic contaminants in exposed organisms. Beluga whale (Delphinapterus leucas) from the endangered St. Lawrence (SL) River Estuary population exhibit deleterious health effects and various severe pathologies that have been associated with contaminant exposure. In contrast, such effects (e.g. reproductive and immunological impairment) are generally less frequent in less exposed populations in the Canadian Arctic (CA). In the present study, opportunistic sampling resulted in the collection immediately after death of liver tissue from a single female neonate SL beluga (SL6) and male and female CA beluga (n=10) from the Arviat region of western Hudson Bay, in addition to sampling of stranded carcasses of male and female SL beluga (n=5) at least 12 h postmortem. We immunologically characterized cross-reactive proteins of hepatic microsomal CYP1A, CYP2B, CYP3A, CYP2E, epoxide hydrolase (EH) and uridine diphosphoglucuronosyl transferase (UDPGT) isozymes. Cross-reactive proteins were found in all SL and CA beluga using anti-rat CYP1A1, anti-rainbow trout CYP3A, anti-human CYP2E1, anti-rabbit EH and anti-human UDPGT1A1 polyclonal antibodies (Abs), whereas faintly cross-reactive CYP2B proteins were only found in SL6 and the CA samples using an anti-rabbit CYP2B1 Ab. In corresponding catalytic activity assessments, only SL6 and all CA beluga microsomal samples exhibited CYP1A-mediated 7-ethoxyresorufin O-deethylase (EROD) activity (51-260 pmol/mg/min), CYP3A-mediated activity (113-899 pmol/mg/min) based on the formation of 6beta-hydroxytestosterone using a testosterone hydroxylase assay, and UDPGT activity (830-4956 pmol/mg/min) based on 1-naphthylglucuronide formation. The marginal cross-reactivity with the anti-CYP2B1 Ab and lack of catalytically measurable hydroxytestosterone isomers associated with CYP2B-type activity in all the SL and CA animals is suggestive of low CYP2B-type enzyme expression in beluga. The absence of measurable total P450 enzyme levels and catalytic activities in samples from the stranded SL belugas suggested catalytically inactive enzymes as a consequence of tissue degradation related due to the time delay of sample collection after death. However, all SL and CA animals demonstrated similar, immunologically cross-reactive phase I and II hepatic enzyme profiles, which is suggestive of the importance of metabolism in the toxicokinetics and fate of xenobiotics in animals from both populations

Animals↗

The development of methods for immunophenotypic and lymphocyte function analyzes for assessment of Southern sea otter (Enhydra lutris nereis) health.

The Southern sea otter (Enhydra lutris nereis) is listed as threatened under the Endangered Species Act. The population began a pattern of slow decline in 1995. The decline was attributed to high adult mortality rates with infectious disease being the major cause of death. Multiple pathogens were implicated in these deaths including opportunistic pathogens such as Coccidiodes immitis and Toxoplasma sp. These findings suggested that the immunological health of mature animals in this population might be compromised. The primary goal of this study was to establish techniques for assessing phenotypic and functional baseline data for peripheral blood mononuclear cells (PBMC) in free-ranging sea otters. Standard total and differential white blood cell counts were augmented by emumeration of T and B lymphocyte subsets. Lymphocyte function was determined by both mitogen-induced proliferation and expression of IL-2 receptors. In addition to establishing normal ranges for adult animals, age-related changes were identified in B lymphocyte numbers and cell-surface density of major histocompatability complex class II (MHC II) proteins. The predominant lymphocyte subpopulation in Southern sea otters is the T lymphocyte. Substantial variation among individual animals was observed within the B lymphocyte population both in cell number and density of MHC II expression. Pups had greater numbers of T and B lymphocyte, as well as, greater MHC II expression on B lymphocytes than adults. Mitogen-induced proliferation of peripheral blood mononuclear cells (PBMC) was variable among individual animals with no significant difference in cell response between age class and gender. Concanavalin (ConA) was a more effective mitogen in stimulating proliferation and interleukin (IL)-2 receptor expression than pokeweed. This data can be used to augment routine hematology profiles and aid in the identification of animals with immunologic perturbations.

Age Factors↗

Molecular and immunophenotypical characterization of progressive and regressive leukemia cell lines.

The P815 and P198 cell lines are clonally related mouse mastocytoma cell lines. They differ in their biologic behavior in that P815 is a progressive tumor cell line, whereas P198 is a regressive one. These cell lines have been extensively used as models for the study of tumor-host relationships and tumor immunology. Although some of their biological properties have been well documented, the molecular mechanisms underlying tumor progression or regression have not been completely elucidated. In this study, we characterized the growth behavior and immunophenotype of these two cell lines, and analyzed their gene profiles using a complementary deoxynucleic acid (cDNA) microarray composed of 514 immunologically relevant genes. Our data showed that the two cell lines exhibited quite dissimilar and contrasting growth characteristics when inoculated into syngeneic mice. P815 tumors grew unremittingly, while P198 tumors gradually regressed. From a molecular viewpoint, P815 cells showed a higher expression of genes promoting tumor growth, such as IGF-1, IL-8R, FGFR1, VEGF-A, and VEGF-B. On the other hand, P198 tumor cells expressed CD11b and CD80, which favor the recruitment of lymphocytes and antigen-presenting cells (APCs), as well as the elicitation of antitumor immunity. P198 tumor cells also depicted a higher expression of genes inhibiting tumor growth, such as TNF-alpha, SOCS-1, CIS1, 4-1BB, and GDF-10. In conclusion, our results contribute further information in the understanding of the molecular mechanisms associated with the regression and progression of P815 and P198 tumor cells.

Animals↗

TIP, a T-cell factor identified using high-throughput screening increases survival in a graft-versus-host disease model.

A coordinated effort combining bioinformatic tools with high-throughput cell-based screening assays was implemented to identify novel factors involved in T-cell biology. We generated a unique library of cDNAs encoding predicted secreted and transmembrane domain-containing proteins generated by analyzing the Human Genome Sciences cDNA database with a combination of two algorithms that predict signal peptides. Supernatants from mammalian cells transiently transfected with this library were incubated with primary T cells and T-cell lines in several high-throughput assays. Here we describe the discovery of a T cell factor, TIP (T cell immunomodulatory protein), which does not show any homology to proteins with known function. Treatment of primary human and murine T cells with TIP in vitro resulted in the secretion of IFN-gamma, TNF-alpha, and IL-10, whereas in vivo TIP had a protective effect in a mouse acute graft-versus-host disease (GVHD) model. Therefore, combining functional genomics with high-throughput cell-based screening is a valuable and efficient approach to identifying immunomodulatory activities for novel proteins.

Adjuvants, Immunologic↗

Diversity among abortion strains of Chlamydia psittaci demonstrated by inclusion morphology, polypeptide profiles and monoclonal antibodies.

Twenty eight C. psittaci abortion strains had been previously classified in to 4 immunologically distinct groups on the basis of cross-protection experiments in a mouse model. To identify the molecular basis of their immunological divergence 4 representative strains were investigated by cellular, molecular and immunological techniques. An identical pattern was obtained by Alul digestion of the amplified major outer membrane protein gene (MOMP) by the polymerase chain reaction (PCR) of the 4 strains. However, inclusion morphology and polypeptide profiles clearly distinguished one strain, named LLG, and its homologous strain POS from the other prototypes by the presence of a unique protein at 26.5 kDa and the absence of a polypeptide at 23 kDa. Six out of 10 monoclonal antibodies (mAbs) raised against abortion strains failed to react with inclusions of the 2 strains. All 6 mAbs reacted with the chlamydial outer membrane complex (COMC). Two of these mAbs, one against the MOMP and one against an antigen at 90 kDa, did not react with immunoblots of LLG and POS. The data provide direct demonstration of the existence of strain variation in the field and classify strains LLG and POS as a distinct C. psittaci serotype 1-subtype. The antigenic diversity among abortion strains should be taken into consideration when designing a subunit vaccine.

Abortion, Veterinary↗

Genomics in the immune system.

The analysis of gene expression in tissues, cells, and biologic systems has evolved in the last decade from the analysis of a selected set of genes to an efficient high throughput whole-genome screening approach of potentially all genes expressed in a tissue or cell sample. Development of sophisticated methodologies such as microarray technology allows an open-ended survey to identify comprehensively the fraction of genes that are differentially expressed between samples and define the samples' unique biology. This discovery-based research provides the opportunity to characterize either new genes with unknown function or genes not previously known to be involved in a biologic process. The latter category may hold surprises that sometimes urge us to redirect our thinking. Here, we review the impact of large-scale gene expression profiling by DNA-microarray technology on basic and clinical aspects of immunology.

Autoimmune Diseases↗

Efficacy of sublingual immunotherapy in asthma and eczema.

Sublingual immunotherapy (SLIT) is the local route of administration of allergen extracts investigated in several controlled clinical trials. In a number of countries, particularly Italy, France and Spain, this has become common in office practice. At variance with subcutaneous immunotherapy, the knowledge of mechanisms of action of SLIT is still at the beginning: some studies, in animal models, provided interesting information: the dendritic cells of oral mucosa act as efficient antigen-presenting cells and produce IL-12, which directs the immune response towards a Th1 profile away from IgE-Th2 profile. Its clinical efficacy (improvement of symptoms and reduction of drug intake) for both asthma and rhinitis has been assessed in detail for the most common allergens: house dust mites, grass pollen, Parietaria, birch pollen and olive tree. SLIT requires further evaluation concerning the treatment of the extrinsic form of atopic dermatitis. The induction of immunologic tolerance rather than immunoreactivity should be worth pursuing due to the immunologic pathway involved in the pathophysiology of atopic dermatitis. The safety profile of SLIT, derived from the clinical trials and postmarketing surveillance studies, turned out to be satisfactory in adults and children. SLIT represents an important step towards an efficacious and safe treatment of patients with allergic respiratory diseases; nevertheless, further studies are necessary to establish it as a viable alternative to subcutaneous immunotherapy.

Administration, Sublingual↗

Cytokine mRNA profile of peripheral blood mononuclear cells isolated from individuals with Trypanosoma cruzi chronic infection.

Characterization of immunologic activities during chronic infection with Trypanosoma cruzi is critical for understanding the dynamics of human Chagas' disease. Since cytokine production is mainly regulated by transcription and mRNA stability, quantitative RT-PCR analysis gives an accurate picture of the influences of disease on cytokine profile. Using RT-PCR, the authors analysed the levels of message expression for several cytokines in peripheral blood mononuclear cells (PBMC) freshly isolated from chagasic patients (CP) and non-infected individuals (NI), and in in vitro-stimulated PBMC from CP. Ex vivo analysis showed that mean levels of expression of IL-5, IL-10, IL-13 and IFN gamma were dramatically increased in PBMC from CP, compared to NI. The levels of IL-2 and IL-4 were not significantly different between groups. Analysis of cytokine mRNA production after in vitro culture with parasite-derived antigens (EPI or TRP) or anti-epimastigote antibodies (Id) showed that these two classes of stimuli induced distinct cytokine responses. While EPI or TRP induced higher production of IFN gamma specific message and low IL-10, anti-Id cells produced higher levels of IL-10 and low IFN gamma. The simultaneous presence of antigenic and antibody stimulation in the host during the chronic phase of Chagas' disease could explain the existence of both inflammatory and anti-inflammatory cellular reactivity detected in most patients.

Adult↗

Murine models of Sjögren's syndrome.

Autoimmune MRL/lpr, MRL/+, and NZB/W mice all develop lacrimal gland inflammatory lesions, which consist of focal mononuclear inflammatory cell infiltrates. Each strain has a different immunocytochemical profile, which appears to be related to the underlying immunologic defects present in that mouse. The appearance of these lesions parallels the evolution of the systemic autoimmune disease. The lesions are dynamic over time with the early appearance of CD4+ T cells (helper T cells) for each strain. Subsequently, there is an accumulation of B cells over time in MRL/+ and NZB/W mice. In the two more rapidly evolving mouse models, MRL/lpr and NZB/W, there is a progressive decline in the percentage of CD8+ cells. Conversely, in the slowly evolving MRL/+ lacrimal gland lesions, there is a persistent and unchanging percentage of CD8+ T cells (suppressor/cytotoxic T cells). Autoimmune mice provide models for the human disorder Sjögren's syndrome and a mechanism for better understanding the immunopathogenesis of autoimmune lacrimal gland disease.

Animals↗

Acute myeloid leukemia cell lines MOLM-17 and MOLM-18 derived from patient with advanced myelodysplastic syndromes.

The two acute myelomonocytic leukemia sister cell lines MOLM-17 and MOLM-18 and the Epstein-Barr-virus positive non-malignant B-lymphoblastoid cell lines (B-LCLs) B422 and B423 were established from the bone marrow sample of a 60-year-old Japanese male in the advanced leukemic phase of refractory anemia with excess of blasts, a subtype of myelodysplastic syndromes (MDS). MOLM-17/-18 are proliferatively responsive to the growth factors present in the culture supernatant of the 5637 cell line. The B-LCLs are constitutively growth factor-independent. MOLM-17 and B422 were established at eight months after the initial diagnosis, while MOLM-18 and B423 were derived from a sample one month later. Immunophenotyping of the first leukemia sample revealed a mixed lineage leukemia immunophenotype with positivity for terminal deoxynucleotidyl transferase (TdT), CD13 and CD19; the second sample revealed solely myeloid characteristics with positivity for CD13, CD41 and CD61, whereas TdT was negative. MOLM-17/-18 showed immunomarker profiles typical of the myelomonocytic lineage. The karyotype analysis of MOLM-17/-18 revealed various non-random numerical and structural abnormalities including del(5)(q?), -7, der(11)add(11)(p11.2)add(11)(q23), add(17)(p11.2), add(18)(p11.2), -20, -22 as common aberrations. Treatment with tumor necrosis factor-alpha induced pronounced cellular differentiation of both cell lines into macrophage-like cells. The overall profile of MOLM-17/-18 based on their extensive immunological, cytogenetic and functional characterization suggests that these cell lines together with the paired B-LCLs B422 and B423 may represent scientifically significant in vitro models which could facilitate investigations into the pathobiology of MDS.

Cell Line, Tumor↗

Drug Insight: the use of intravenous immunoglobulin in neurology--therapeutic considerations and practical issues.

Over the past few years, we have achieved increasing success in the treatment of a number of autoimmune-mediated disorders affecting nerves and muscles. This success is partly attributable to the use of high-dose polyclonal intravenous immunoglobulin (IVIg), which has dramatically changed our treatment options. On the basis of results from controlled, but non-FDA-approved, clinical trials, IVIg is now the treatment of choice for Guillain-Barré syndrome, chronic idiopathic inflammatory demyelinating polyneuropathy and multifocal motor neuropathy; IVIg offers rescue therapy for patients with rapidly worsening myasthenia gravis, and is a second-line therapy for dermatomyositis, stiff-person syndrome, and pregnancy-associated or postpartum multiple sclerosis attacks. The ability of IVIg to treat such immunologically diverse disorders effectively, coupled with its excellent safety profile, has led clinicians to use the drug more liberally, even in diseases for which the data are weak and not evidence-based and in patients with coexisting conditions. Use of IVIg for such indications can increase the risk of complications while raising the cost of the drug. Practical issues regarding dosing and frequency of infusions generate dilemmas in clinical practice. In this article, we review the current indications for IVIg treatment, address practical issues related to the use and costs of the drug, and summarize its mechanisms of action.

Autoimmune Diseases↗

Strategy for monitoring T cell responses to NY-ESO-1 in patients with any HLA class I allele.

NY-ESO-1 elicits frequent antibody responses in cancer patients, accompanied by strong CD8(+) T cell responses against HLA-A2-restricted epitopes. To broaden the range of cancer patients who can be assessed for immunity to NY-ESO-1, a general method was devised to detect T cell reactivity independent of prior characterization of epitopes. A recombinant adenoviral vector encoding the full cDNA sequence of NY-ESO-1 was used to transduce CD8-depleted peripheral blood lymphocytes as antigen-presenting cells. These modified antigen-presenting cells were then used to restimulate memory effector cells against NY-ESO-1 from the peripheral blood of cancer patients. Specific CD8(+) T cells thus sensitized were assayed on autologous B cell targets infected with a recombinant vaccinia virus encoding NY-ESO-1. Strong polyclonal responses were observed against NY-ESO-1 in antibody-positive patients, regardless of their HLA profile. Because the vectors do not cross-react immunologically, only responses to NY-ESO-1 were detected. The approach described here allows monitoring of CD8(+) T cell responses to NY-ESO-1 in the context of various HLA alleles and has led to the definition of NY-ESO-1 peptides presented by HLA-Cw3 and HLA-Cw6 molecules.

Alleles↗

T cells expressing the gammadelta receptor are essential for Th2-mediated inflammation in patients with acute exacerbation of asthma.

OBJECTIVE: T lymphocytes have a central regulatory role in the pathogenesis of asthma. The objective of this study was to characterize immunologically the activation stage of asthma and the functional profile of lymphocytes from induced sputum, with particular emphasis on gammadelta T cells. METHODS: Induced sputum was collected from 10 patients with acute exacerbation of asthma, and from healthy controls. The expression of activation markers on freshly isolated induced sputum lymphocytes and T-cell subsets was analyzed by double immunofluorescent staining and flow cytometry. Fas ligand (FasL) was determined by reverse transcriptase-polymerase chain reaction analysis. The phenotype of gammadelta T-cell subpopulations was tested by A13 and BB3 monoclonal antibodies. In this context, the functional profile of gammadelta T cells was tested in a chromium releasing test. RESULTS: A significantly decreased proportion of alphabeta T cells and an increased proportion of gammadelta T cells, CD56+ cells and CD8+ gammadelta T cells were found in asthma patients compared with healthy controls. In asthmatic patients, there is a significantly increased proportion of T cells expressing CD69 and CD25 antigen. After stimulation of gammadelta T cells, an increased expression of intracellular tumour necrosis factor-alpha, interleukin (IL)-4 and IL10 cytokines were found at higher levels than controls. Interferon-gamma was observed at similar levels in asthma patients and healthy controls. Freshly isolated T-cell receptor (TCR) gammadelta+ cells exhibited an increased percentage of FasL in our patient group. FasL mRNA was detected in TCR gammadelta+ cells before and after IL2 stimulation. TCR gammadelta+ cells were cytotoxic against the K562 cell line. This natural killer activity was mediated by the A13-positive subpopulation. CONCLUSION: The presence of cytokines producing gammadelta cells in induced sputum of asthmatic patients is consistent with regulatory activities. These cells display also cytotoxic function.

Acute Disease↗