A long and winding road: the history of Immunology & Cell Biology from 1936 to 1987.

This piece for the 100th year of publication of Immunology & Cell Biology focuses on the contributions of two long-serving Editors-in-Chief, Mark Mitchell (1936-1963) and Derrick Rowley (1963-1987). This was a period of growth and consolidation for the journal, through sometimes challenging and changing times. Some of the notable works published in the journal during this time are also highlighted.

A centenary of service: 100 years of Immunology & Cell Biology.

This year marks the 100th year of the publication of Immunology & Cell Biology since it was first published in March 1924 as the Australian Journal of Experimental Biology and Medical Science. In this Editorial, we recount the journal from its founding, to its focus on immunology, through to the modern era.

Is there a role for antibodies targeting muscarinic acetylcholine receptors in the pathogenesis of schizophrenia?

OBJECTIVE: Muscarinic receptor dysfunction has been suggested to play an important role in the pathophysiology of schizophrenia. Recently, it has also become clear that immune reactivity directed against neurotransmitter receptors may play a pathogenic role in some cases of schizophrenia. The aim of this review is to summarize the case for muscarinic receptor dysfunction in schizophrenia and the evidence supporting the hypothesis that this dysfunction is related to the development of muscarinic receptor-targeting antibodies. METHOD: The article reviews studies of muscarinic receptors and the presence and potential role(s) of anti-muscarinic acetylcholine receptor antibodies in people with schizophrenia. RESULTS: There is accumulating evidence that altered or deficient muscarinic signalling underlies some of the key clinical features of schizophrenia. Although the number of studies investigating anti-muscarinic acetylcholine receptor antibodies in schizophrenia is relatively small, they consistently demonstrate that such antibodies are present in a proportion of patients. This evidence suggests that these antibodies could have pathogenic effects or exist as a biomarker to an unknown pathophysiological process in schizophrenia. CONCLUSION: The presence of elevated levels of anti-muscarinic acetylcholine receptor antibodies may identify a subgroup of people with schizophrenia, potentially informing aetiopathogenesis, clinical presentation and treatment. To date, all studies have examined antibodies in participants with chronic schizophrenia, who have likely received antipsychotic medication for many years. As these medications modulate immune functions and regulate receptor densities, it is recommended that future studies screen for the presence of anti-muscarinic antibodies in people experiencing their first episode of psychosis.

Novel Therapeutics for Multiple Sclerosis Designed by Parasitic Worms.

The evolutionary response to endemic infections with parasitic worms (helminth) was the development of a distinct regulatory immune profile arising from the need to encapsulate the helminths while simultaneously repairing tissue damage. According to the old friend's hypothesis, the diminished exposure to these parasites in the developed world has resulted in a dysregulated immune response that contributes to the increased incidence of immune mediated diseases such as Multiple Sclerosis (MS). Indeed, the global distribution of MS shows an inverse correlation to the prevalence of helminth infection. On this basis, the possibility of treating MS with helminth infection has been explored in animal models and phase 1 and 2 human clinical trials. However, the possibility also exists that the individual immune modulatory molecules secreted by helminth parasites may offer a more defined therapeutic strategy.

Thiopalmitoylation of altered peptide ligands enhances their protective effects in an animal model of multiple sclerosis.

Previously, we have shown that conjugation of a palmitic chain via a thioester bond to a cysteine residue in weakly or nonencephalitogenic or neuritogenic peptides markedly enhances their ability to induce autoimmune disease in an MHC class II-restricted manner. From those studies, however, it was not clear whether thiopalmitoylation of the peptides was merely enhancing their disease-inducing potential or whether the lipid was itself playing a pathogenic role. To investigate this further, we have now tested the effects of thiopalmitoylation on MHC class II-restricted altered peptide ligands (APLs), which are normally protective in experimental autoimmune encephalomyelitis, the animal model of multiple sclerosis. We hypothesized that if thiopalmitoylation of a peptide merely enhances its innate potential, then thiopalmitoylated APLs (S-palmAPLs) should show enhanced protective effects. Alternatively, if thiopalmitoylation itself can make a peptide pathogenic, then S-palmAPLs should have decreased therapeutic potential. We synthesized APLs and corresponding S-palmAPLs and showed that the S-palmAPLs were much more effective than the nonconjugated APL at inhibiting the development of experimental autoimmune encephalomyelitis. This was due to several features of the S-palmAPL:S-palmAPL-primed cells show an enhanced ability to proliferate and produce the anti-inflammatory cytokine, IL-10, in vitro. Furthermore, the bioavailability of S-palmAPL was greatly enhanced, compared with the nonpalmitoylated APL, and S-palm APL was taken up more rapidly into dendritic cells and channeled into the MHC class II processing pathway. These results show that thiopalmitoylation of MHC class II-restricted peptides is a simple way to enhance their effects in vivo and could have wide therapeutic application.

Quantitative proteome profiling of CNS-infiltrating autoreactive CD4+ cells reveals selective changes during experimental autoimmune encephalomyelitis.

Experimental autoimmune encephalomyelitis (EAE) is a murine model of multiple sclerosis, a chronic neurodegenerative and inflammatory autoimmune condition of the central nervous system (CNS). Pathology is driven by the infiltration of autoreactive CD4(+) lymphocytes into the CNS, where they attack neuronal sheaths causing ascending paralysis. We used an isotope-coded protein labeling approach to investigate the proteome of CD4(+) cells isolated from the spinal cord and brain of mice at various stages of EAE progression in two EAE disease models: PLP139-151-induced relapsing-remitting EAE and MOG35-55-induced chronic EAE, which emulate the two forms of human multiple sclerosis. A total of 1120 proteins were quantified across disease onset, peak-disease, and remission phases of disease, and of these 13 up-regulated proteins of interest were identified with functions relating to the regulation of inflammation, leukocyte adhesion and migration, tissue repair, and the regulation of transcription/translation. Proteins implicated in processes such as inflammation (S100A4 and S100A9) and tissue repair (annexin A1), which represent key events during EAE progression, were validated by quantitative PCR. This is the first targeted analysis of autoreactive cells purified from the CNS during EAE, highlighting fundamental CD4(+) cell-driven processes that occur during the initiation of relapse and remission stages of disease.

Correlation of adrenomedullin gene expression in peripheral blood leukocytes with severity of ischemic stroke.

Human adrenomedullin (ADM), a 52-amino acid peptide, belongs to the calcitonin/calcitonin gene-related peptide (CGRP)/amylin peptide family. ADM acts as a multifunctional regulatory peptide and is upregulated in response to hypoxia. Previous microarray studies have found increased ADM gene (ADM) expression in peripheral blood cells of patients with stroke, however, it is unknown if an increased ADM level is correlated with severity of human ischemic stroke. This study investigated ADM expression in peripheral blood leukocytes (PBL) of healthy controls and subjects at day 1, week 1 and week 3 postacute ischemic stroke using rtPCR methodology. We found that ADM expression was significantly upregulated on the first day of stroke compared to the healthy subjects and the disease controls; the levels remained elevated for up to week 3. Further, ADM expression at day 1 was correlated with stroke severity measured by the National Institute of Healthy Stroke Scale (NIHSS), the modified Barthel Index (mBI) and the modified Rankin Scale (mRS). This could indicate that ADM expression level is related to the severity of tissue damage. We suggest that increased ADM expression in PBL after acute ischemic stroke is most likely to indicate that these cells have been subjected to hypoxia and that the magnitude of expression is likely to be related to the volume of hypoxic tissue. Hypoxia can affect lymphocytes function and could affect the immune response to stroke. The correlation of ADM expression level with the measures of stroke severity implicates ADM--a potential blood bio-marker in studies of ischemic stroke.

Female reproductive issues in multiple sclerosis.

Multiple sclerosis (MS) is more common in females than males and frequently affects women during their reproductive years. Thus, issues surrounding pregnancy and reproduction are of concern to women with MS. This review documents studies that shed light on reproductive issues in women with MS. The available literature was searched for papers relating to pregnancy and MS. Pregnancy is protective in MS in the short term, perhaps due to modulation of the immune system in pregnancy. It also possible that changes in the brain in pregnancy could protect against the effects of inflammation. The long-term effects of pregnancy also seem to be beneficial to MS, perhaps due to long-term epigenetic changes or possibly due to the effects of fetal microchimerism. Obstetric outcomes in women with MS are similar to those in the general population. In addition, there have been no reports of severe fetal abnormalities in babies exposed to first-line MS therapies. There is no good evidence that breast-feeding is protective in MS. There is no evidence that oral contraceptive pill use predisposes to MS, nor influences the clinical course of MS. After menopause, there is possible deterioration of MS, but it's difficult to disentangle this from the effects of aging and the natural progressive history of MS. The strong biological effect of pregnancy on MS deserves further study, so that these mechanisms can possibly be replicated as therapies for MS.

An appraisal of emerging therapeutic targets for multiple sclerosis derived from current preclinical models.

INTRODUCTION: Multiple sclerosis (MS) is a chronic inflammatory, demyelinating, and neurodegenerative condition affecting the central nervous system (CNS). Although therapeutic approaches have become available over the last 20 years that markedly slow the progression of disease, there is no cure for MS. Furthermore, the capacity to repair existing CNS damage caused by MS remains very limited. AREAS COVERED: Several animal models are widely used in MS research to identify potential druggable targets for new treatment of MS. In this review, we look at targets identified since 2019 in studies using these models, and their potential for effecting a cure for MS. EXPERT OPINION: Refinement of therapeutic strategies targeting key molecules involved in the activation of immune cells, cytokine, and chemokine signaling, and the polarization of the immune response have dominated recent publications. While some progress has been made in identifying effective targets to combat chronic demyelination and neurodegeneration, much more work is required. Progress is largely limited by the gaps in knowledge of how the immune system and the nervous system interact in MS and its animal models, and whether the numerous targets present in both systems respond in the same way in each system to the same therapeutic manipulation.

Interleukin-6 gene promoter-572 C allele may play a role in rate of disease progression in multiple sclerosis.

Multiple sclerosis (MS) is an inflammatory demyelinating disease affecting the central nervous system. Although the exact pathogenesis of MS is unknown, it is generally considered to be an autoimmune disease, with numerous genetic and environmental factors determining disease susceptibility and severity. One important mediator of immune responses and inflammation is interleukin-6 (IL-6). Previously, elevated levels of IL-6 in mononuclear cells in blood and in brain tissue from MS patients have been reported. Various polymorphisms in the promoter region of the IL6 gene have also been linked with IL-6 protein levels. In MS, several small studies have investigated whether two IL6 promoter polymorphisms (-597 G>A and -174 G>C) correlate with MS susceptibility, but with varying results. In the present study, we analyzed these polymorphisms, together with an additional polymorphism (-572 G>C) in 279 healthy controls and 509 patients with MS. We found no significant differences between MS patients and healthy controls for the different -597 or -174 IL6 promoter alleles or genotypes. There was a slight reduction in the percentage of individuals with MS who carried a C allele at position -572, although this was not significant after correction for multiple comparisons. Interestingly, however, the -572 C allele showed a significant correlation with the MS severity score, suggesting a possible role in disease progression.

Effects of biological sex and pregnancy in experimental autoimmune encephalomyelitis: It's complicated.

Experimental autoimmune encephalomyelitis (EAE) can be induced in many animal strains by inoculation with central nervous system antigens and adjuvant or by the passive transfer of lymphocytes reactive with these antigens and is widely used as an animal model for multiple sclerosis (MS). There are reports that female sex and pregnancy affect EAE. Here we review the effects of biological sex and the effects of pregnancy on the clinical features (including disease susceptibility) and pathophysiology of EAE. We also review reports of the possible mechanisms underlying these differences. These include sex-related differences in the immune system and in the central nervous system, the effects of hormones and the sex chromosomes and molecules unique to pregnancy. We also review sex differences in the response to factors that can modify the course of EAE. Our conclusion is that the effects of biological sex in EAE vary amongst animal models and should not be widely extrapolated. In EAE, it is therefore essential that studies looking at the effects of biological sex or pregnancy give full information about the model that is used (i.e. animal strain, sex, the inducing antigen, timing of EAE induction in relation to pregnancy, etc.). In addition, it would be preferable if more than one EAE model were used, to show if any observed effects are generalizable. This is clearly a field that requires further work. However, understanding of the mechanisms of sex differences could lead to greater understanding of EAE, and suggest possible therapies for MS.

Sex differences in Guillain Barré syndrome, chronic inflammatory demyelinating polyradiculoneuropathy and experimental autoimmune neuritis.

Guillain Barré syndrome (GBS) and its variants, and chronic inflammatory demyelinating polyradiculoneuropathy (CIDP and its variants, are regarded as immune mediated neuropathies. Unlike in many autoimmune disorders, GBS and CIDP are more common in males than females. Sex is not a clear predictor of outcome. Experimental autoimmune neuritis (EAN) is an animal model of these diseases, but there are no studies of the effects of sex in EAN. The pathogenesis of GBS and CIDP involves immune response to non-protein antigens, antigen presentation through non-conventional T cells and, in CIDP with nodopathy, IgG4 antibody responses to antigens. There are some reported sex differences in some of these elements of the immune system and we speculate that these sex differences could contribute to the male predominance of these diseases, and suggest that sex differences in peripheral nerves is a topic worthy of further study.

The effect of ageing on human lymphocyte subsets: comparison of males and females.

BACKGROUND: There is reported to be a decline in immune function and an alteration in the frequency of circulating lymphocytes with advancing age. There are also differences in ageing and lifespan between males and females. We performed this study to see if there were differences between males and females in the frequency of the different lymphocyte subsets with age. RESULTS: Using flow cytometry we have examined different populations of peripheral blood leukocytes purified from healthy subjects with age ranging from the third to the tenth decade. We used linear regression analysis to determine if there is a linear relationship between age and cell frequencies. For the whole group, we find that with age there is a significant decline in the percentage of naïve T cells and CD8(+) T cells, and an increase in the percentage of effector memory cells, CD4(+)foxp3(+) T cells and NK cells. For all cells where there was an effect of ageing, the slope of the curve was greater for men than for women and this was statistically significant for CD8(+)alphabeta(+) T cells and CD3(+)CD45RA(-)CCR7(-) effector memory cells. There was also a difference for naïve cells but this was not significant. CONCLUSION: The cause of the change in percentage of lymphocyte subsets with age, and the different effects on males and females is not fully understood but warrants further study.

Correlation Between Anti-Myelin Proteolipid Protein (PLP) Antibodies and Disease Severity in Multiple Sclerosis Patients With PLP Response-Permissive HLA Types.

The most prominent pathological features of multiple sclerosis (MS) are demyelination and neurodegeneration. The exact pathogenesis of MS is unknown, but it is generally regarded as a T cell-mediated autoimmune disease. Increasing evidence, however, suggests that other components of the immune system, particularly B cells and antibodies, contribute to the cumulative CNS damage and worsening disability that characterize the disease course in many patients. We have previously described strongly elevated T cell reactivity to an extracellular domain of the most abundant CNS myelin protein, myelin proteolipid protein (PLP) in people with MS. The current paper addresses the question of whether this region of PLP is also a target of autoantibodies in MS. Here we show that serum levels of isotype-switched anti-PLP181-230 specific antibodies are significantly elevated in patients with MS compared to healthy individuals and patients with other neurological diseases. These anti-PLP181-230 antibodies can also live-label PLP-transfected cells, confirming that they can recognize native PLP expressed at the cell surface. Importantly, the antibodies are only elevated in patients who carry HLA molecules that allow strong T cell responses to PLP. In that subgroup of patients, there is a positive correlation between the levels of anti-PLP181-230 antibodies and the severity of MS. These results demonstrate that anti-PLP antibodies have potentially important roles to play in the pathogenesis of MS.

Reduced IκB-α Protein Levels in Peripheral Blood Cells of Patients with Multiple Sclerosis-A Possible Cause of Constitutive NF-κB Activation.

NF-κB signaling pathways are dysregulated in both the central nervous system (CNS) and peripheral blood cells in multiple sclerosis (MS), but the cause of this is unknown. We have recently reported that peripheral blood mononuclear cells (PBMC) of patients with MS have increased constitutive activation and translocation of the transcription factor NF-κB to the nucleus compared to healthy subjects. NF-κB can be activated through either canonical or non-canonical pathways. In the canonical pathway, activation of NF-κB is normally negatively regulated by the inhibitor IκB. We therefore hypothesized that the increased activation of NF-κB could be caused by reduced IκB-α in the cells of patients with MS, possibly due to increased activity of the IκB kinase (IKK) complex, which regulates IκB-α. Alternatively, changes to the activity of key molecules in the non-canonical pathway, such as IKKα, could also lead to increased NF-κB activation. We therefore used Western blotting to detect IκB-α levels and ELISA to investigate NF-κB DNA binding activity and phosphorylation of IKKα and IKKß in samples from PBMC of MS patients and controls. The level of full-length IκB-α protein in the cytosolic fraction of PBMC of MS patients was significantly reduced compared to healthy subjects, with significantly more evidence of multiple low molecular weight putative degradation products of IκB-α present in MS patients compared to healthy subjects. Conversely, the level of NF-κB DNA binding activity was increased in whole cell lysates from MS patients. Both IKKα and IKKß showed increased overall activity in MS compared to healthy subjects, although not all of the MS patients showed increased activity compared to the healthy subjects, suggesting that there may be several different mechanisms underlying the constitutive activation of NF-κB in MS. Taken together, these findings suggest that there may be multiple points at which the NF-κB pathway is dysregulated in MS and that decreased levels of the full-length IκB-α protein are a major component in this.

Immune activation in the peripheral blood of patients with acute ischemic stroke.

Lymphocytes, neutrophils and macrophages are found in the brain in areas of acute ischaemic stroke. There is also evidence of modulation of systemic immune function after stroke, with post-stroke immunosuppression being observed. Because lymphocytes are activated in the peripheral immune compartment, before entry to the target organ, we reasoned that activated lymphocytes would be present in the circulation, prior to entering the brain, in patients after stroke. Because immune responses are controlled by regulatory mechanisms, we also reasoned that the post-stroke immunosuppression would involve T regulatory cells. The aim of the study was to look for evidence of immune activation and alterations in regulatory T cells in the peripheral blood of patients after acute ischaemic stroke, in comparison to age-matched healthy controls and patients with other neurological diseases (OND), and to determine the phenotype of the activated cells. The percentages of total and activated T cells, B cells, monocyte/ macrophages, and NK/NK-T cells were determined by labelling peripheral blood leukocytes with specific cell surface markers and analysis with 4-colour flow cytometry. The percentages of activated T cells and regulatory T cells were significantly increased in patients with ischemic stroke compared to healthy subjects and patients with OND. There was also an increase in the percentage of CCR7+ T cells. There were no significant differences in the activation of other cell types. In conclusion, there is evidence of immune activation and Treg cells in acute ischaemic stroke.

CTLA-4 single-nucleotide polymorphisms in a Caucasian population with schizophrenia.

Associations between a single-nucleotide polymorphism (SNP) in exon 1 of the cytotoxic T-lymphocyte antigen-4 (CTLA4) gene and schizophrenia in a Korean population have been previously described. The current study investigated whether a similar link occurs in a Caucasian population with schizophrenia. One hundred and twenty-two age- and sex-matched pairs of people with DSM-III-R diagnosis of schizophrenia and healthy controls were included in this study. Three previously described SNPs (from the promoter, exon 1 and 3' UTR) of the CTLA4 gene were analysed. In the entire sample, we detected no allelic or genotypic association for any of the three SNPs. Given documented gender differences in incidence of schizophrenia, we conducted separate analyses of male and female participants. In males, both the promoter region SNP (-318C/T) and the 3' UTR SNP demonstrated nominally significant association with schizophrenia. The 3' UTR SNP remained significant following correction for multiple testing (permuted P=0.046). In addition, all possible haplogenotypes showed significant association with disease in males with two--both containing the 3' UTR SNP--remaining significant following correction for the genotypic tests of all SNPs and haplogenotypes in males. These results suggest a role for the 3' UTR SNP and/or variants in high linkage disequilibrium with this SNP in the pathogenesis of schizophrenia.

PLP1 Mutations in Patients with Multiple Sclerosis: Identification of a New Mutation and Potential Pathogenicity of the Mutations.

PLP1 is located on the X-chromosome and encodes myelin proteolipid protein (PLP), the most abundant protein in central nervous system myelin. Generally, point mutations in PLP1 result in X-linked dysmyelinating disorders, such as Pelizaeus-Merzbacher disease (PMD) or spastic paraplegia type 2 (SPG2). However, several case studies have identified patients with missense point mutations in PLP1 and clinical symptoms and signs compatible with a diagnosis of multiple sclerosis (MS). To investigate if PLP1 mutations occur relatively frequently in MS, we sequenced the coding regions of PLP1 in 22 female MS patients who had developed disease after the age of 40 and in 42 healthy women, and identified a missense mutation in exon 2 of PLP1 resulting in a Leu30Val mutation in the protein in one of the MS patients. mCherry-tagged plasmids containing wild type or mutant PLP1 sequences of PLP, including two known PMD/SPG2-related mutations as positive controls, were constructed and transfected into Cos-7 cells. In comparison with cells transfected with wild type PLP1, all mutations caused significant accumulation of PLP in the endoplasmic reticulum of the cells and induction of the unfolded protein response-a mechanism that leads to apoptosis of cells expressing mutant proteins. Additionally, in silico analysis of the binding of peptides containing the Leu30Val mutation to the human leukocyte antigen (HLA) molecules carried by the patient harboring this mutation suggested that the mutation could produce several novel immunogenic epitopes in this patient. These results support the idea that mutations in myelin-related genes could contribute to the development of MS in a small proportion of patients.

Increased constitutive activation of NF-κB p65 (RelA) in peripheral blood cells of patients with progressive multiple sclerosis.

The NF-κB signalling pathway plays an important role in controlling cellular immune responses, inflammation and apoptosis. In multiple sclerosis (MS), there is evidence of dysregulation of NF-κB signalling in patients with a relapsing-remitting disease course, but thus far there is little information on whether it is also dysregulated in patients with progressive disease. We hypothesised that patients with progressive MS would have more activation of NF-κB than relapsing-remitting MS patients. Using several different methods, we showed that there was more nuclear translocation of p65 in cells from progressive MS patients, particularly in T cells and monocytes. In addition, the amount of p65 translocated to the nucleus in cells of patients with progressive MS was not increased upon non-specific activation of the cells with the mitogen Con A. These results suggest that NF-κB dysregulation occurs in patients with progressive MS patients, as well as those with relapsing-remitting MS.