Neonatal Immune Tolerance Research Articles

Zfp335 establishes eTreg lineage and neonatal immune tolerance by targeting Hadha-mediated fatty acid oxidation.

Regulatory T cells (Tregs) are instrumental in maintaining immune tolerance and preventing destructive autoimmunity, but how heterogeneous Treg populations are established remains largely unknown. Here, we show that Zfp335 deletion in Tregs failed to differentiate into effector Tregs (eTregs) and lose Treg-suppressive function and that KO mice exhibited early-onset lethal autoimmune inflammation with unrestricted activation of conventional T cells. Single-cell RNA-Seq analyses revealed that Zfp335-deficient Tregs lacked a eTreg population and showed dramatic accumulation of a dysfunctional Treg subset. Mechanistically, Zfp335-deficient Tregs displayed reduced oxidative phosphorylation and dysfunctional mitochondrial activity. Further studies revealed that Zfp335 controlled eTreg differentiation by regulating fatty acid oxidation (FAO) through direct targeting of the FAO enzyme Hadha. Importantly, we demonstrate a positive correlation between ZNF335 and HADHA expression in human eTregs. Our findings reveal that Zfp335 controls FAO-driven eTreg differentiation to establish immune tolerance.

Single-cell transcriptional diversity of neonatal umbilical cord blood immune cells reveals neonatal immune tolerance

The characteristics of neonatal immune cells display intrinsic differences compared with adult immune cells. Therefore, a comprehensive analysis of key gene expression regulation is required to understand the response of the human fetal immune system to infections. Here, we applied single-cell RNA sequencing (scRNA-seq) and single-cell sequencing assay for transposase-accessible chromatin (scATAC-seq) to systematically profile umbilical cord blood (UCB) nucleated cells and peripheral blood mononuclear cells (PBMCs) to identify their composition and differentially expressed genes. The immune cells in neonatal UCB demonstrated the expression of key genes, such as HBG2, NFKBIA, JUN, FOS, and TNFAIP3. In contrast, natural killer and T cells, which are constituents of adult PBMCs, exhibited high cytotoxic gene expression. Furthermore, we obtained similar results from the data of scATAC-seq by identifying the status of chromatin accessibility of key genes. Therefore, scRNA-seq and scATAC-seq of neonatal UCB nucleated cells and adult PBMCs could serve as an invaluable resource for elucidating the regulatory mechanisms of responses of distinct immune cell types and further identifying the differences between neonatal and adult immune responses to predict the potential underlying mechanism for neonatal immune tolerance.

Maternal Type 1 Diabetes Reduces Autoantigen-Responsive CD4+ T Cells in Offspring

Autoimmunity against pancreatic β-cell autoantigens is a characteristic of childhood type 1 diabetes (T1D). Autoimmunity usually appears in genetically susceptible children with the development of autoantibodies against (pro)insulin in early childhood. The offspring of mothers with T1D are protected from this process. The aim of this study was to determine whether the protection conferred by maternal T1D is associated with improved neonatal tolerance against (pro)insulin. Consistent with improved neonatal tolerance, the offspring of mothers with T1D had reduced cord blood CD4+ T-cell responses to proinsulin and insulin, a reduction in the inflammatory profile of their proinsulin-responsive CD4+ T cells, and improved regulation of CD4+ T cell responses to proinsulin at 9 months of age, as compared with offspring with a father or sibling with T1D. Maternal T1D was also associated with a modest reduction in CpG methylation of the INS gene in cord blood mononuclear cells from offspring with a susceptible INS genotype. Our findings support the concept that a maternal T1D environment improves neonatal immune tolerance against the autoantigen (pro)insulin.

Immunological microenvironment in the testis.

BackgroundThe testis is specific in that it produces haploid germ cells of which autoantigens newly appear long after the neonatal immune tolerance. Under normal condition, these autoantigens are protected by the blood‐testis barrier formed by Sertoli cells. Thus, the testis is an immunologically privileged site where haploid cells are protected from autoimmune attack.MethodsThe immunological microenvironment in the testis was experimentally investigated using mice and rats.Main findingsNot only the blood‐testis barrier but also various immuno‐suppressive factors are involved in the immune‐privileged testis. Indeed, germ cells transplanted into the xenogeneic seminiferous tubules could proliferate and differentiate with no aid of artificial immunosuppression. On the other hand, autoimmune orchitis could be experimentally produced by various methods of immunization with syngeneic or xenogeneic germ cell antigens.ConclusionOur results indicate that the testis is immunologically privileged but also immunologically fragile organ. Therefore, the dual nature is critical for immunoregulation of testicular function.

Neonatal Immune Tolerance Induction to Allow Long-Term Studies With an Immunogenic Therapeutic Monoclonal Antibody in Mice.

The purpose of this study is to test the feasibility of neonatal immune tolerance induction in mice to enable long-term pharmacokinetic studies with immunogenic therapeutic monoclonal antibodies (mAb). Neonatal immune tolerance was induced by transfer of a mAb to neonatal mice via colostrum from nursing mother mice treated with two subcutaneous doses of a tolerogen starting within the first 24 h after delivery. Adalimumab and efalizumab were administered as tolerogens at various dose levels. Tolerance induction was evaluated in the offspring after reaching adulthood at 8 weeks of age. After a single intravenous injection of the same mAb as used for tolerance induction, the pharmacokinetics of the mAb and formation of anti-drug antibodies (ADA) in plasma were assessed using ELISA. Tolerance induction to adalimumab was achieved in a maternal dose-dependent manner. Adalimumab immune-tolerant offspring showed a slower adalimumab clearance (4.24 ± 0.32 mL/day/kg) as compared to the control group (12.09 ± 3.81 mL/day/kg). In the control group, accelerated clearance started 7 days after adalimumab dosing, whereas immune-tolerant offspring showed a log-linear terminal concentration-time course. In the offspring, the absence of predose ADA levels was indicative of successful tolerance induction. The second test compound efalizumab was not immunogenic in mice under our experimental conditions. Overall, the present study demonstrated the suitability of neonatal immune tolerance induction for a 4-week single dose study in adult mice with a human therapeutic mAb that is otherwise immunogenic in laboratory animals.

Neonatal immune-tolerance in mice does not prevent xenograft rejection

Assessing the efficacy of human stem cell transplantation in rodent models is complicated by the significant immune rejection that occurs. Two recent reports have shown conflicting results using neonatal tolerance to xenografts in rats. Here we extend this approach to mice and assess whether neonatal tolerance can prevent the rapid rejection of xenografts. In three strains of neonatal immune-intact mice, using two different brain transplant regimes and three independent stem cell types, we conclusively show that there is rapid rejection of the implanted cells. We also address specific challenges associated with the generation of humanized mouse models of disease.

Is MC1 dispensable for regulation of cutaneous inflammatory and immune responses?

The melanocortin-1 receptor (MC1 ) - being most abundantly expressed in the skin by melanocytes - has a physiological role for melanin pigmentation in many vertebrate species. MC1 has also been implicated in regulation of skin inflammation as this receptor is detectable in the majority of non-melanocytic cell types and its ligand α-melanocyte-stimulating hormone (α-MSH) exerts immunoregulatory and anti-inflammatory effects. However, in vivo studies on mice with targeted disruption of MC1 have been missing in the context of skin inflammation until recently. Wolnicka-Glubisz et al. now reported that the course of ultraviolet (UV)-induced inflammation, contact hypersensitivity, neonatal immune tolerance and UV-induced immunosuppression is similar in MC1 signal-deficient (C57BL/6-Mc1r(e/e)) and wild-type mice. These unexpected findings are supported by own observations in experimentally induced immune-complex-mediated vasculitis: Mc1r(e/e) mice exhibited a similar extent of the reverse passive cutaneous Arthus reaction compared with wild-type animals. Future studies are thus needed to clarify whether these findings are due to limitations in the chosen mouse model and/or point to additional MC subtypes that may regulate inflammatory and immune responses in the skin.

Overall picture of an emerging neonatal infectious disease induced by a superantigenic exotoxin mainly produced by methicillin‐resistant Staphylococcus aureus

Since 1992, many neonates in neonatal intensive care units in Japan have been developing fever and systemic exanthema. Immunological analyses of neonates with these symptoms has revealed that the bacterial superantigen, toxic shock syndrome toxin-1 (TSST-1) is the cause. The name neonatal TSS-like exanthematous disease (NTED) has been applied to this condition. The most striking clinical finding has been that none of the term neonates have developed shock or died of NTED. The timing of NTED epidemics has coincided with the spread of emerging TSST-1-producing methicillin-resistant Staphylococcus aureus clones in Japan. The low frequency of pregnant women with positive anti-TSST-1 antibody titers could be one reason for the spread of NTED in Japan. Neonates have immune tolerance against TSST-1 and may actively suppress the immune response to NTED with interleukin-10. According to the T cell responses in infants or young children with diseases induced by TSST-1, the pathophysiology of TSST-1-related diseases may be age-dependent. The precise mechanism of anergy and deletion of specific T cells stimulated with TSST-1 should be investigated in neonates infected with NTED. Both NTED and TSS might provide good models for analyzing the mechanism(s) of neonatal immune tolerance and the age-dependence of human immunity. This disease has not only become representative of diseases caused by superantigens, but has also yielded a considerable amount of evidence about human immune reactions against superantigens.

Functional melanocortin 1 receptor Mc1r is not necessary for an inflammatory response to UV radiation in adult mouse skin

The G-protein-coupled receptor, Mc1r, plays a major role in pigment production and has been reported to be important in the inflammatory response. We have investigated the effect of deficiency in Mc1r on UV-induced inflammation. Mice on the same genetic background were used - C57BL/6-c (albino), C57BL/6 (black), C57BL/6-Mc1r(e/e) deficient (yellow). FACS analysis of disaggregated skin showed a similar dose-dependent increase in Ly6G(+) and CD11b(+) cells in response to UV radiation in all groups. No differences in UV-induced edema or in DNA damage were detected between groups. The contact hypersensitivity response, neonatal immune tolerance and UV immunosuppression were all similar in C57BL/6 and C57BL/6-Mc1r(e/e) mice. We conclude that the absence of Mc1r does not impair the inflammatory response to UV radiation or the generation of immunosuppression.

Putting Flesh and Polish on Autoimmune Hepatitis and Moving the Disease of Exclusion to Inclusion

Autoimmune hepatitis emerged during an era when concepts of neonatal immune tolerance, clonal selection of lymphocytes, and "forbidden clones" of activated immune cells were forming. The diagnosis had to be deduced from circumstantial evidence and by exclusion of other conditions. The goals of this review are to demonstrate how a clinician nonscientist can contribute to the maturation of autoimmune hepatitis and to illustrate the principles of clinical investigation that can be applied broadly to other projects. Autoimmune hepatitis initially had to be distinguished from other diseases, and improvements in the tests for viral and immune markers were instrumental in this regard. Diversification of the clinical phenotype to accommodate acute severe, asymptomatic, elderly, and variant forms enhanced the pertinence of the disease, and the formation of the International Autoimmune Hepatitis Group standardized the diagnosis, interconnected investigators, and promoted global acceptance of the condition. Subsequent studies refined current corticosteroid-based therapies, identified prognostic markers, assessed genetic predispositions, explored new pharmacological agents, and forecast the emergence of cellular and molecular interventions. Good fortune, stimulating mentors, career dedication, practical goal selection, protocol compliance, compulsive record keeping, personal resilience, and strong collaborations were the bases for progress. Autoimmune hepatitis exemplifies an evolutionary process in the science of autoimmunity and the people committed to its study. Lessons derived from this experience can be far-reaching.

Deficient inflammatory response to UV radiation in neonatal mice

Mechanisms of juvenile susceptibility to cancer are not well understood. The immune response in neonates favors nonresponsiveness or T(H)2-dominant responses, raising the question of a role for neonatal immunity in this susceptibility. We have investigated the postulate that the inflammatory response differs in neonatal and adult skin. We found no inflammatory infiltrate into neonatal mouse skin in response to UV irradiation as a function of time, dose, or wavelength, although UV-induced DNA damage was readily detected. In contrast, UV irradiation of adult mice initiated a dose- and time-dependent influx of inflammatory cells, chiefly CD11b(+)Ly6G(+) neutrophils, into the skin, detected by immunohistochemistry and quantitated by FACS analysis. This inflammatory response was initiated by UVB (290-320 nm) but not by UVA (320-400 nm). Further, in neonates, in contrast to adults, neither topical trinitrochlorobenzene (TNCB) nor i.p. thioglycollate initiated an inflammatory infiltrate. Conversely, topical TNCB applied to neonates was tolerogenic, resulting in a subsequent antigen-specific decrease of the contact-hypersensitivity response in adults. Neonatal blood contained abundant neutrophils, which exhibited impaired chemotaxis to the chemokine growth-related oncogene-alpha but efficient chemotaxis to the bacterial product fMLP, concomitant with decreased expression of CXCR2 but normal levels of CD11b. We propose this neonatal deficiency in the inflammatory response is a significant, previously unrecognized factor in neonatal immune tolerance and may contribute to neonatal susceptibility to cancer, including melanoma and other UV-induced cancers.

Repeated DNA vaccination induced neonatal tolerance against Escherichia coli beta-galactosidase in immune-competent mice

Objectives: To induce neonatal immune tolerance in immune competent animals by repeated DNA vaccination. Materials and Methods: We used two groups of neonatal immune competent mice, C57BL/6, six in each. Mice in one study group received subcutaneous injection of 2 μg of endotoxin-free plasmid, pCBLacZ, at the posterior neck area. Concentration of the plasmid was 0.5 mg/mL. This plasmid encoded a potent immunogen, Escherichia coil beta-galactosidase. Mice in the other study group received intrarectal injection of same amount of pCBLacZ. All the animals received injection at the second day of birth and 5 consecutive injections of the same dose of pCBLacZ at an interval of 7 days. Seven days after the last shoot, the mice received an intramuscular injection of 50 μg of endotoxin-free pCBLacZ at a concentration of 2 mg/mL at the left anterior tibialis. We used 6 naive adult C57BL/6 as the control. These animals received an intramuscular injection of 50 μg of pCBLacZ at the left anterior tibialis at age of 42 days. Results: We examined the expression of beta-galactosidase in muscle fibers 6 days after the last intramuscular injection. The average numbers of fibers expressing beta-galactosidase was 168.2 in the mice receiving intrarectal immunization as compared with 0 in the mice receiving subcutaneous injection and 16.2 in their little mates. (p=0.001 by likelihood ratio test). There were two mice got 409 and 513 blue fibers respectively. Serum IgG against the beta-galactosidase was lower in the animals receiving intrarectal injection as compared with the mice in the other two groups by nonparametric Mann-Whitney test. Conclusions: Repeated DNA vaccination in the neonatal stage can induce immune tolerance against a potent immunogen in the immune competent animals. The ways of administration determine the success of tolerance induction.

UNCONTROLLED MICROSPORUM GYPSEUM INFECTION IN A NEW BORN MAMMAL

SUMMARY A newborn South American coati, Nasua nasua (Linn.), acquired an intractable and eventually fatal Microsporum gypseum infection from the soil in its cage at the Zoo. The final picture of a hepatorenal syndrome with shock developed. The disease was neither relieved nor evidently aggravated by griseofulvin therapy. Fungal resistance developed to the drug in vitro. It was thought that the infection persisted initially because of neonatal immune tolerance. The pathogenesis of M. gypseum infection was also studied in the guinea-pig while comparing the virulence of the strains isolated.