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Since the start of the COVID-19 pandemic, in a short span of 3 years, vaccination against SARS-CoV-2 has resulted in the end of the pandemic. Patients with inborn errors of immunity (IEI) are at an increased risk for SARS-CoV-2 infection; however, serious illnesses and mortality, especially in primary antibody deficiencies (PADs), have been lower than expected and lower than other high-risk groups. This suggests that PAD patients may mount a reasonable effective response to the SARS-CoV-2 vaccine. Several studies have been published regarding antibody responses, with contradictory reports. The current study is, perhaps, the most comprehensive study of phenotypically defined various lymphocyte populations in PAD patients following the SARS-CoV-2 vaccine. In this study, we examined, following two vaccinations and, in a few cases, prior to and following the 1st and 2nd vaccinations, subsets of CD4 and CD8 T cells (Naïve, TCM, TEM, TEMRA), T follicular helper cells (TFH1, TFH2, TFH17, TFH1/17), B cells (naïve, transitional, marginal zone, germinal center, IgM memory, switched memory, plasmablasts, CD21low), regulatory lymphocytes (CD4Treg, CD8Treg, TFR, Breg), and SARS-CoV-2-specific activation of CD4 T cells and CD8 T cells (CD69, CD137), SARS-CoV-2 tetramer-positive CD8 T cells, and CD8 CTL. Our data show significant alterations in various B cell subsets including Breg, whereas only a few subsets of various T cells revealed alterations. These data suggest that large proportions of PAD patients may mount significant responses to the vaccine.

Corona Virus Disease 19 (COVID-19) has been a major pandemic impacting a huge population worldwide, and it continues to present serious health threats, necessitating the development of novel protective nutraceuticals. Biobran/MGN-3, an arabinoxylan rice bran, is a potent immunomodulator for both humans and animals that has recently been demonstrated to protect against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in vitro. We here investigate Biobran/MGN-3′s potential to enhance an antiviral immune response in humans. Peripheral blood mononuclear cells (PBMCs) derived from eight subjects taking Biobran/MGN-3 (age 55–65 years) and eight age-matched control subjects were stimulated with irradiated SARS-CoV-2 virus and then subjected to immuno-phenotyping and multiplex cytokine/chemokine assays. Results showed that PBMCs from subjects supplemented with Biobran/MGN-3 had significantly increased activation of plasmacytoid dendritic cells (pDCs) coupled with increased IFN-α secretion. We also observed higher baseline expression of HLA-DR (human leukocyte antigen-DR isotype) on dendritic cells (DCs) and increased secretion of chemokines and cytokines, as well as a substantial increase in cytotoxic T cell generation for subjects taking Biobran/MGN-3. Our results suggest that Biobran/MGN-3 primes immunity and therefore may be used for boosting immune responses against SARS-CoV-2 infections and other diseases, particularly in high-risk populations such as the elderly.

BackgroundAdvancing age is a major risk factor for respiratory viral infections. The infections are often prolonged and difficult to resolve resulting hospitalizations and mortality. The recent COVID-19 pandemic has highlighted this as elderly subjects have emerged as vulnerable populations that display increased susceptibility and severity to SARS-CoV-2. There is an urgent need to identify the probable mechanisms underlying this to protect against future outbreaks of such nature. Innate immunity is the first line of defense against viruses and its decline impacts downstream immune responses. This is because dendritic cells (DCs) and macrophages are key cellular elements of the innate immune system that can sense and respond to viruses by producing inflammatory mediators and priming CD4 and CD8 T-cell responses.ResultsWe investigated the changes in innate immune responses to SARS-CoV-2 as a function of age. Our results using human PBMCs from aged, middle-aged, and young subjects indicate that the activation of DCs and monocytes in response to SARS-CoV-2 is compromised with age. The impairment is most apparent in pDCs where both aged and middle-aged display reduced responses. The secretion of IL-29 that confers protection against respiratory viruses is also decreased in both aged and middle-aged subjects. In contrast, inflammatory mediators associated with severe COVID-19 including CXCL-8, TREM-1 are increased with age. This is also apparent in the gene expression data where pathways related host defense display an age dependent decrease with a concomitant increase in inflammatory pathways. Not only are the inflammatory pathways and mediators increased after stimulation with SARS-CoV-2 but also at homeostasis. In keeping with reduced DC activation, the induction of cytotoxic CD8 T cells is also impaired in aged subjects. However, the CD8 T cells from aged subjects display increased baseline activation in accordance with the enhanced baseline inflammation.ConclusionsOur results demonstrate a decline in protective anti-viral immune responses and increase in damaging inflammatory responses with age indicating that dysregulated innate immune responses play a significant role in the increased susceptibility of aged subjects to COVID-19. Furthermore, the dysregulation in immune responses develops early on as middle-aged demonstrate several of these changes.

In this study, efficient T cell activation is demonstrated using cell-sized artificial antigen-presenting cells (aAPCs) with protein-conjugated bilayer lipid membranes that mimic biological cell membranes. The highly uniform aAPCs are generated by a facile method based on standard droplet microfluidic devices. These aAPCs are able to activate the T cells in peripheral blood mononuclear cells, showing a 28-fold increase in interferon gamma (IFNγ) secretion, a 233-fold increase in antigen-specific CD8 T cells expansion, and a 16-fold increase of CD4 T cell expansion. The aAPCs do not require repetitive boosting or additional stimulants and can function at a relatively low aAPC-to-T cell ratio (1:17). The research presents strong evidence that the surface fluidity and size of the aAPCs are critical to the effective formation of immune synapses essential for T cell activation. The findings demonstrate that the microfluidic-generated aAPCs can be instrumental in investigating the physiological conditions and mechanisms for T cell activation. Finally, this method demonstrates the feasibility of customizable aAPCs for a cost-effective off-the-shelf approach to immunotherapy.

The authors regret the incorrect publication of the name of the co-author ‘Veedamali S Subramanian’ in the original article. The correct name ‘Veedamali S Subramanian’ is updated in the authors list. The authors would like to apologise for any inconvenience caused.

Macrophages play a role in preventing inflammation in the respiratory tract. To investigate the mechanisms that lead to tolerance in macrophages, we examined the crosstalk between airway epithelial cells (AECs) and macrophages using a 2D coculture model. Culture of macrophages with AECs led to a significant inhibition of LPS induced pro-inflammatory responses. More importantly, AECs induced the secretion of TGF-β and IL-10 from macrophages even in the absence of LPS stimulation. In addition, the expression of inhibitory molecule, CD200R was also upregulated on AEC exposed macrophages. Furthermore, the AECs exposed macrophages induced significantly increased level of T regulatory cells. Investigation into the possible mechanisms indicated that a combination of growth factor, G-CSF, and metabolites, Kynurenine and lactic acid produced by AECs is responsible for inducing tolerance in macrophages. Interestingly, all these molecules had differential effect on macrophages with G-CSF inducing TGF-β, Kynurenine elevating IL-10, and lactic acid upregulating CD200R. Furthermore, a cocktail of these factors/metabolites induced similar changes in macrophages as AEC exposure. Altogether, these data identify factors secreted by AECs that enhance tolerance in the respiratory tract. These mediators thus have the potential to be used for therapeutic purposes to modulate respiratory inflammation following local viral infections and lung diseases.

PurposeCD8 cytotoxic T cells (CTLs) play a critical role in the clearance of virally infected cells. SARS-CoV-2-specific CD8 T cells and functional CTLs in natural infections and following COVID-19 vaccine in primary antibody deficiency (PAD) have not been reported. In this study, we evaluated T cell response following COVID-19 or COVID-19 mRNA vaccination in patients with PADs by assessing SARS-CoV-2 tetramer-positive CD8 T cells and functional CTLs.MethodsSARS-CoV-2-specific CD8 and functional CTLs were examined in a patient with X-linked agammaglobulinemia (XLA) and a patient with common variable immunodeficiency (CVID) following COVID-19 infection, and in 5 patients with CVID and 5 healthy controls 1 month following 2nd dose of COVID-19 mRNA vaccine (Pfizer-BioNTech). Cells were stained with SARS-CoV-2 spike protein-specific tetramers, and for functional CTLs (CD8+ CD107a+ granzyme B+ perforin+), with monoclonal antibodies and isotype controls and analyzed by flow cytometry.ResultsSARS-CoV-2-specific tetramer + CD8 T cells and functional CTLs in the patient with XLA following COVID-19 infection were higher, as compared to healthy control subject following COVID-19 infection. On the other hand, SARS-CoV2-tetramer + CD8 T cells and functional CTLs were lower in CVID patient following COVID19 infection as compared to healthy control following COVID-19 infection. SARS-CoV2-tetramer + CD8 T cells and functional CTLs were significantly lower in SARS-CoV2-naive CVID patients (n = 10) following vaccination when compared to SARS-CoV-2-naive healthy vaccinated controls (n = 10).ConclusionsCVID is associated with reduced SARS-CoV-2-specific CD8 T cells and functional CTLs in both natural SARS-CoV-2 infection and in response to SARS-CoV-2 mRNA vaccine, whereas natural infection in XLA is associated with a robust SARS-CoV-2-specific CD8 and functional CTL responses.

Alzheimer's disease (AD) is associated with dysregulated immune and inflammatory responses. Emerging evidence indicates that peripheral immune activation is linked to neuroinflammation and AD pathogenesis. The present study focuses on determining the role of IL-21 in the pathogenesis of AD using human samples and the 5xFAD mice model. We find that the levels of IL-21 are increased in the periphery of both humans and mice in AD. In addition, the proportions of IL-21 target cells, Tfh and B plasma cells as well as activation of monocytes is increased in PBMCs from AD and mild cognitively impaired (MCI) subjects as compared to age-matched controls, indicating immune activation. In contrast, the percentage of B1 cells that control inflammation is decreased. These changes are due to IL-21 as the expression of IL-21 receptor (IL-21R) is higher on all these cells in AD. Furthermore, treatment with recombinant IL-21 in AD mice also leads to similar alterations in Tfh, B, B1, and macrophages. The effect of IL-21 is not confined to the periphery since increased expression of IL-21R is also observed in both humans and mice hippocampus derived from the AD brains. In addition, mice injected with IL-21 display increased deposition of amyloid beta (Aβ) plaques in the brain which is reduced following anti-IL-21R antibody that blocks the IL-21 signaling. Moreover, activation of microglia was enhanced in IL-21-injected mice. In keeping with enhanced microglial activation, we also observed increased production of pro-inflammatory cytokines, IL-18 and IL-6 in IL-21-injected mice. The microglial activation and cytokines were both inhibited following IL-21R blockage. Altogether, IL-21 escalates AD pathology by enhancing peripheral and brain immune and inflammatory responses leading to increased Aβ plaque deposition. IL-21 impacts AD neuropathology by enhancing peripheral and neuronal immune activation, inflammation, and Aβ plaque deposition. Increased levels of IL-21 in the circulation of AD and MCI subjects enhances the proportions of Tfh and B plasma cells indicative of peripheral immune activation. On the other hand, the proportions of B1 cells that help reduce inflammation and clear Aβ are reduced. In addition to the periphery, IL-21 also acts on the brain via IL-21 receptor, IL-21R that displays increased expression in the hippocampi of AD and MCI subjects. IL-21 enhances the activation of microglia, induces the secretion of pro-inflammatory cytokines and deposition of Aβ plaques in the brain in AD.

Abstract: COVID-19 exhibits a sex-bias with males showing signs of more severe disease and hospitalizations compared to females. The mechanisms are not clear but differential immune responses, particularly the initial innate immune response, between sexes may be playing a role. The early innate immune responses to SARS-CoV2 have not been studied because of the gap in timing between the patient getting infected, showing symptoms, and getting the treatment. The primary objective of the present study was to compare the response of DCs and monocytes from males and females to SARS-CoV2 24h after infection. To investigated this, PBMCs from healthy young individuals were stimulated in vitro with the virus. Our results indicate that DCs and monocytes from females displayed increased activation compared to males. In addition, females secreted significantly higher levels of IFN-α and IL-29 early while the secretion of IFN-α was delayed and prolonged in males. Further investigations revealed that the secretion of CXCL-10, a chemokine associated with lung complications, was higher in males than females. The PBMCs from females also displayed increased induction of CTLs. Bulk RNA-seq analysis indicates that IFN related pathways may be upregulated in females at baseline. Altogether, our results suggest that decreased activation of pDCs, mDCs and monocytes, the delayed and prolonged IFN-α secretion along with increased CXCL-10 secretion may be responsible for the severity of COVID-19 in males. The study is supported by grants R00RG2352 and R01RG3735 to AA from University of California office of the President (UCOP).

The disruption of methionine (L-MET) metabolism has been linked with neurodevelopmental disorders such as autism and schizophrenia and neurodegenerative disorders such as Alzheimer's disorder. We previously showed that repeated administration to adult mice of methionine produced impairments of cognitive deficits. Considering the decreased neurogenesis and increased molecular inflammation hypotheses of cognitive deficits in Alzheimer's, we aimed to explore whether the methionine regimen that produced cognitive deficits is associated with altered neuroinflammation, neurogenesis, or neurodegeneration. We found that repeated administration of L-MET at a dose equivalent to two-fold of daily dietary intake for seven days enhanced the activation of microglia and inflammation in the brain, and decreased neurogenesis in the hippocampus without affecting degeneration. Furthermore, sub-chronic and chronic L-MET treatment of human neuroblastoma (SH-SY5Y) inhibited cell cycle progression, an effect that was reversed by decreasing removing L-MET from the medium. These results support a role for neuroinflammation and neurogenesis in mediating the mechanism through which L-MET induces cognitive deficits. The results also uncover L-MET restriction, neuroinflammation, and neurogenesis as potential preventive and/or therapeutic targets for mental disorders associated with cognitive disorders, including schizophrenia and Alzheimer's disease.

IntroductionIn the trials of corona virus vaccines, detailed analyses of subsets of lymphocytes were not carried out. We present perhaps the most comprehensive immunological analysis of 29 subsets of B and T cells in 2 healthy subjects receiving 2 doses of the Pfizer SARS-CoV-2 (COVID-19) vaccine.MethodsAnalyses were performed prior to vaccination, 3 weeks following the 1st dose, and 4 weeks following the 2nd dose. Total, naïve (TN), and different memory and effector subsets (TCM, TEM, and TEMRA) of CD4+ and CD8+ T cells; SARS-CoV-2 spike protein-specific tetramer+, and cytotoxic CD8+ T; subsets of T follicular cells (TFH, TFH1, TFH2, TFH1/TFH17, and TFH17); B-cell subsets (mature B cells, naive B cells, transitional B cells, marginal zone B cells, class-switched memory B cells, germinal center B cells, and CD21low B cells), and plasmablasts; and regulatory lymphocytes (CD4+ Treg, CD8+ Treg, Breg, and TFR cells) were evaluated with specific monoclonal antibodies by flow cytometry.ResultsA lack of COVID-19 IgG antibodies after the 1st dose in one of 2 subjects was associated with increased regulatory lymphocytes and decreased plasmablasts. Seroconversion after the 2nd dose in this subject was associated with decreased TFR cells and increased plasmablasts. In both subjects, CD4 TEM and CD8 TCM were markedly increased following the 2nd dose. TFH1 and regulatory lymphocytes were increased (except Breg) following the 1st dose. A striking increase in SARS-CoV-2-specific CD8+ T cells was observed following the 2nd dose.ConclusionOur data support the need for 2nd dose of vaccine to induce strong SARS-CoV-2 CD8 T-cell specific response and generation of memory subsets of CD4+ and CD8+ T cells. Regulatory lymphocytes appear to play a role in the magnitude of response.

Vitamin C is well documented to have antiviral functions; however, there is limited information about its effect on airway epithelial cells-the first cells to encounter infections. Here, we examined the effect of vitamin C on human bronchial epithelium transformed with Ad12-SV40 2B (BEAS-2B) cells, and observed that sodium-dependent vitamin C transporter 2 (SVCT2) was the primary vitamin C transporter. Transcriptomic analysis revealed that treating BEAS-2B cells with vitamin C led to a significant upregulation of several metabolic pathways and interferon-stimulated genes (ISGs) along with a downregulation of pathways involved in lung injury and inflammation. Remarkably, vitamin C also enhanced the expression of the viral-sensing receptors retinoic acid-inducible gene 1 (RIG-1) and melanoma differentiation-associated protein 5 (MDA-5), which was confirmed at the protein and functional levels. In addition, the lungs of l-gulono-γ-lactone oxidase knockout (GULO-KO) mice also displayed a marked decrease in these genes compared to wild-type controls. Collectively, our findings indicate that vitamin C acts at multiple levels to exert its antiviral and protective functions in the lungs.

We demonstrate a label free and high-throughput microbubble-based acoustic microstreaming technique to isolate rare circulating cells such as circulating cancer associated fibroblasts (cCAFs) in addition to circulating tumor cells (CTCs) and immune cells (i.e. leukocytes) from clinically diagnosed patients with a capture efficiency of 94% while preserving cell functional integrity within 8 minutes. The microfluidic device is self-pumping and was optimized to increase flow rate and achieve near perfect capturing of rare cells enabled by having a trapping capacity above the acoustic vortex saturation concentration threshold. Our approach enables rapid isolation of CTCs, cCAFs and their associated clusters from blood samples of cancer patients at different stages. By examining the combined role of cCAFs and CTCs in early cancer onset and metastasis progression, the device accurately diagnoses both cancer and the metastatic propensity of breast cancer patients. This was confirmed by flow cytometry where we observed that metastatic breast cancer blood samples had significantly higher percentage of exhausted CD8+ T cells expressing programmed cell death protein 1 (PD1), higher number of CD4+ T regulatory cells and T helper cells. We show for the first time that our lateral cavity acoustic transducers (LCATs)-based approach can thus be developed into a metastatic propensity assay for clinical usage by elucidating cancer immunological responses and the complex relationships between CTCs and its companion tumor microenvironment.

We report perhaps the most comprehensive study of subsets of CD4+ and CD8+ and subsets of B cells in a mild symptomatic SARS-CoV-2+ immunocompetent patient and a common variable immunodeficiency disease (CVID) patient who had normal absolute lymphocyte counts and remained negative for SARS-CoV-2 IgG antibodies. Naïve (TN), central memory (TCM), effector memory (TEM), and terminally differentiated effector memory (TEMRA) subsets of CD4+ and CD8+ T cells, subsets of T follicular helper cells (cTFH, TFH1, TFH2, TFH17, TFH1/TFH17, and TFR), CD4 Treg, CD8 Treg, mature B cells, transitional B cells, marginal zone B cells, germinal center (GC) B cells, CD21low B cells, antibody-secreting cells (plasmablasts), and Breg cells were examined in patients and age-matched controls with appropriate monoclonal antibodies and isotype controls using multicolor flow cytometry. Different patterns of abnormalities (often contrasting) were observed in the subsets of CD4+ T, CD8+ T, B-cell subsets, and regulatory lymphocytes among the immunocompetent patient and CVID patient as compared to corresponding healthy controls. Furthermore, when data were analyzed between the 2 patients, the immunocompetent patient demonstrated greater changes in various subsets as compared to the CVID patient. These data demonstrate different immunological responses to SARS-CoV-2 infection in an immunocompetent patient and the CVID patient. A marked decrease in GC B cells and plasmablasts may be responsible for failure to make SARS-CoV-2 antibodies. The lack of SARS-CoV-2 antibodies with mild clinical disease suggests an important role of T-cell response in defense against SARS-CoV-2 infection.

Coronavirus disease 2019 (COVID-19) exhibits a sex bias with males showing signs of more severe disease and hospitalizations compared with females. The mechanisms are not clear but differential immune responses, particularly the initial innate immune response, between sexes may be playing a role. The early innate immune responses to SARS-CoV-2 have not been studied because of the gap in timing between the patient becoming infected, showing symptoms, and getting the treatment. The primary objective of the present study was to compare the response of dendritic cells (DCs) and monocytes from males and females to SARS-CoV-2, 24 h after infection. To investigate this, peripheral blood mononuclear cells (PBMCs) from healthy young individuals were stimulated in vitro with the virus. Our results indicate that PBMCs from females upregulated the expression of HLA-DR and CD86 on pDCs and mDCs after stimulation with the virus, while the activation of these cells was not significant in males. Monocytes from females also displayed increased activation than males. In addition, females secreted significantly higher levels of IFN-α and IL-29 compared with males at 24 h. However, the situation was reversed at 1 week post stimulation and males displayed high levels of IFN-α production compared with females. Further investigations revealed that the secretion of CXCL-10, a chemokine associated with lung complications, was higher in males than females at 24 h. The PBMCs from females also displayed increased induction of CTLs. Altogether, our results suggest that decreased activation of pDCs, mDCs, and monocytes and the delayed and prolonged IFN-α secretion along with increased CXCL-10 secretion may be responsible for the increased morbidity and mortality of males to COVID-19.

During the last 2 years, the entire world has been severely devastated by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic (COVID-19) as it resulted in several million deaths across the globe. While the virus infects people indiscriminately, the casualty risk is higher mainly in old, and middle-aged COVID-19 patients. The incidences of COVID-19 associated co-morbidity and mortality have a great deal of correlation with the weakened and malfunctioning immune systems of elderly people. Presumably, due to the physiological changes associated with aging and because of possible comorbidities such as diabetes, hypertension, obesity, cardiovascular, and lung diseases, which are more common in elderly people, may be considered as the reason making the elderly vulnerable to the infection on one hand, and COVID-19 associated complications on the other. The accretion of senescent immune cells not only contributes to the deterioration of host defense, but also results in elevated inflammatory phenotype persuaded immune dysfunction. In the present review, we envisage to correlate functioning of the immune defense of older COVID-19 patients with secondary/super infection, increased susceptibility or aggravation against already existing cancer, infectious, autoimmune, and other chronic inflammatory diseases. Moreover, we have discussed how age-linked modulations in the immune system affect therapeutic response against administered drugs as well as immunological response to various prophylactic measures including vaccination in the elderly host. The present review also provides an insight into the intricate pathophysiology of the aging and the overall immune response of the host to SARS-CoV-2 infection. A better understanding of age-related immune dysfunction is likely to help us in the development of targeted preemptive strategies for deadly COVID-19 in elderly patients.

Influenza-like illness (ILI) remains a major cause of severe mortality and morbidity in the elderly. Aging is associated with a decreased ability to sense pathogens and mount effective innate and adaptive immune responses, thus mandating the development of protective nutraceuticals. Biobran/MGN-3, an arabinoxylan from rice bran, has potent anti-aging and immunomodulatory effects, suggesting that it may be effective against ILI. The objective of the current study was to investigate the effect of Biobran/MGN-3 on ILI incidence, natural killer (NK) cell activity, and the expressions of RIG-1 (retinoic acid-inducible gene 1), MDA5 (melanoma differentiation-associated protein 5), and their downstream signaling genes ISG-15 (interferon-stimulated genes 15) and MX1 (myxovirus (influenza) resistance 1, interferon-inducible). A double-blind, placebo-controlled clinical trial included eighty healthy older adults over 55 years old, 40 males and 40 females, who received either a placebo or Biobran/MGN-3 (500 mg/day) for 3 months during known ILI seasonality (peak incidence) in Egypt. The incidence of ILI was confirmed clinically according to the WHO case definition criteria. Hematological, hepatic, and renal parameters were assessed in all subjects, while the activity of NK and NKT (natural killer T) cells was assessed in six randomly chosen subjects in each group by the degranulation assay. The effect of Biobran/MGN-3 on RIG-1 and MDA5, as well as downstream ISG15 and MX1, was assessed in BEAS-2B pulmonary epithelial cells using flow cytometry. The incidence rate and incidence density of ILI in the Biobran/MGN-3 group were 5.0% and 0.57 cases per 1000 person-days, respectively, compared to 22.5% and 2.95 cases per 1000 person-days in the placebo group. Furthermore, Biobran/MGN-3 ingestion significantly enhanced NK activity compared to the basal levels and to the placebo group. In addition, Biobran/MGN-3 significantly upregulated the expression levels of RIG-1, MDA5, ISG15, and MX1 in the human pulmonary epithelial BEAS-2B cell lines. No side effects were observed. Taken together, Biobran/MGN-3 supplementation enhanced the innate immune response of elderly subjects by upregulating the NK activity associated with reduction of ILI incidence. It also upregulated the intracellular RIG-1, MDA5, ISG15, and MX1 expression in pulmonary epithelial tissue cultures. Biobran/MGN-3 could be a novel agent with prophylactic effects against a wide spectrum of respiratory viral infections that warrants further investigation.

This study examines the link between peripheral immune changes in perpetuation of the Alzheimer's disease (AD) neuropathology and cognitive deficits. Our research design using human AD patients and rodent model is supported by past evidence from genomic studies. We observed an active immune response against Aβ as indicated by the increased Aβ specific IgG antibody in the serum of AD and patients with mild cognitive impairments as compared to healthy controls. A similar increase in IgG and decrease in IgM antibody against Aβ was also confirmed in the 5xFAD mouse model of AD. More importantly, we observed a negative correlation between reduced IgM levels and cognitive dysfunction that manifested as impaired memory consolidation. Strong peripheral immune activation was supported by increased activation of microglia in the brain and macrophages in the spleen of AD mice compared to wild type control littermates. Furthermore, inflammatory cytokine IL-21 that is involved in antibody class switching was elevated in the plasma of AD patients and correlated positively with the IgG antibody levels. Concurrently, an increase in IL-21 and IL-17 was observed in spleen cells from AD mice. Further investigation revealed that proportions of T follicular helper (Tfh) cells that secrete IL-21 are increased in the spleen of AD mice. In contrast to Tfh, the frequency of B1 cells that produce IgM antibodies was reduced in AD mice. Altogether, these data indicate that in AD the immune tolerance to Aβ is compromised leading to chronic immune/inflammatory responses against Aβ that are detrimental and cause neuropathology. Graphical Abstract Healthy subjects are tolerant to Aβ and usually react weakly to it resulting the in the production of IgM class of antibodies that are efficient at clearing up self-antigens such as Aβ without causing inflammation. In contrast, Alzheimer's disease patients mount a strong immune response against Aβ probably in an effort to clear up excessive Aβ. There is enhanced production of inflammatory cytokines such as IL-21 as well as an increase in Tfh cells that cause antibody class switching form IgM to IgG. The strong immune response is inefficient at clearing up Aβ and instead exacerbates inflammation that causes AD neuropathology and cognitive dysfunction.

Smoking is a major risk factor for pulmonary diseases that include chronic obstructive pulmonary diseases (COPD) and cancer. Nicotine is the toxic and addictive component of tobacco products, like cigarettes, that negatively affects the immune system. Here, we examined the effect of nicotine on the IL-22 pathway that protects lung function by increasing transepithelial resistance and epithelial cell regeneration and repair. Our results indicate that exposure to nicotine impairs the regenerative capacity of primary bronchial epithelial cells in scratch assays. IL-22 at 100 ng/ml significantly improved wound healing in epithelial cells; however, the exposure to nicotine hampered the IL-22-mediated effect of wound healing. Investigation into the mechanisms showed that IL-22 receptor, IL-22Rα1, was downregulated in the presence of nicotine as determined by q-PCR and flow cytometry. We also investigated the effect of nicotine on IL-22 production by T cells. Results indicate that nicotine inhibited the secretion of IL-22 from T cells in response to aryl hydrocarbon receptor (AHR) ligand, FICZ. Altogether, the data suggests that nicotine negatively influences the IL-22-IL-22R axis. This impairment may contribute to the nicotine-mediated detrimental effects on lung function.

BACKGROUND:CD8+ regulatory T cells (CD8+ Tregs) are relatively recently described T cell subsets that have been shown to regulate various T cell responses and appear to play a role in autoimmunity. However, their effects on B cells have not been explored. OBJECTIVES:In this investigation we examine the effect of CD8+ Tregs on various subsets of peripheral B cells include naïve B cells, transitional B cells, marginal zone B cells, IgM memory B cells, class switched memory B cells, and plasmablasts, and on the expression of B cell-activating factor receptor (BAFF-R). METHODS:CD8+ T cells were first purified and then activated with anti-CD3/CD28 beads to generate CD8+ Tregs. Purified CD19+ B cells were cultured alone or with sorted CD8+ Tregs (CD8+CD183+CCR7+CD45RA-) and activated with anti-CD40 monoclonal antibody and CpG. B cell subsets and the expression of BAFF-R on naïve and memory B cells were analyzed using various monoclonal antibodies and corresponding control isotypes. Ten thousand cells were acquired and analyzed by FACSCalibur using the FlowJo software. RESULTS:CD8+ Tregs selectively and significantly suppressed plasmablasts without any significant effect on other B cell subsets or on the expression of BAFF-R. CONCLUSION:CD8+ Tregs may play a role in autoimmunity by regulating antibody production via suppression of plasmablasts.

AimThe role of CD4+ Treg in immune responses has been well established. More recently, a role of CD8+ T regulatory cells (CD8 Treg) in the regulation of immune responses in health and autoimmune diseases has been investigated. Furthermore, different investigators have used different markers to define CD8 Treg. Finally, regulatory effects of CD8 Treg have been studied against T-cell responses; however, their role in regulating B-cell proliferation and immunoglobulin production has not been evaluated. Therefore, in this study we examined the effect of two types of CD8 Treg on B-cell proliferation and immunoglobulin production.MethodsPurified CD8+ T cells were activated with anti-CD3/CD28 for 48 h and then sorted into two different types of CD8 Treg as defined by two different sets of markers, CD8+CD183+CD197+CD45RA- and CD8+CD183+CD25highCD278+. Purified B cells were cocultured with sorted CD8 Treg at 1:1, 1:1/2, and 1:1/4 ratios and activated with anti-CD40 and CpG. B-cell proliferation was assessed by the CFSE dye dilution assay and immunoglobulin production by the ELISA assay.ResultsOur data show CD183+CD197+CD45RA-CD8 Treg significantly inhibited B-cell proliferation and inhibited IgM and IgG production but not IgA production at 1:1 ratio only. However, CD183+CD25highCD278+CD8 Treg inhibited significantly B-cell proliferation at 1:1 and 1:1/2 ratios and IgM, IgG, and IgA production at all ratios.ConclusionCD8 Treg regulate B-cell responses, and CD183+CD25highCD278+CD8 Treg are more powerful regulators of B-cell proliferation and immunoglobulin production than CD183+CD197+CD45RA-CD8 Treg and, therefore, may be used as preferred markers for CD8 Treg.

BACKGROUND: Macrophages and fibroblasts are 2 major cell types involved in healing after myocardial infarction (MI), contributing to myocardial remodeling and fibrosis. Post-MI fibrosis progression is characterized by a decrease in cardiac macrophage content. OBJECTIVES: This study explores the potential of macrophages to express fibroblast genes and the direct role of these cells in post-MI cardiac fibrosis. METHODS: Prolonged in vitro culture of human macrophages was used to evaluate the capacity to express fibroblast markers. Infiltrating cardiac macrophages was tracked in vivo after experimental MI of LysM(Cre/+);ROSA26(EYFP/+) transgenic mice. The expression of Yellow Fluorescent Protein (YFP) in these animals is restricted to myeloid lineage allowing the identification of macrophage-derived fibroblasts. The expression in YFP-positive cells of fibroblast markers was determined in myocardial tissue sections of hearts from these mice after MI. RESULTS: Expression of the fibroblast markers type I collagen, prolyl-4-hydroxylase, fibroblast specific protein-1, and fibroblast activation protein was evidenced in YFP-positive cells in the heart after MI. The presence of fibroblasts after MI was evaluated in the hearts of animals after depletion of macrophages with clodronate liposomes. This macrophage depletion significantly reduced the number of Mac3+Col1A1+ cells in the heart after MI. CONCLUSIONS: The data provide both in vitro and in vivo evidence for the ability of macrophages to transition and adopt a fibroblast-like phenotype. Therapeutic manipulation of this macrophage-fibroblast transition may hold promise for favorably modulating the fibrotic response after MI and after other cardiovascular pathological processes.

The original version of this Article contained an error in the spelling of the author Hannah Nguyen, which was incorrectly given as Hannah Ngyuen. This has now been corrected in both the PDF and HTML versions of the Article.

Plasmacytoid dendritic cells (PDCs) are critical for defense against respiratory viruses because of their propensity to secrete high levels of type I interferons (IFN). The functions of PDCs in the lung can be influenced by airway epithelial cells. We examined the effect of human primary bronchial epithelial cells (PBECs) on PDC functions by performing RNA-sequencing of PDCs after co-culture with air liquid interface differentiated PBECs. Functional analysis revealed that PDCs co-cultured with PBECs displayed upregulation of type I IFN production and response genes. Upregulated transcripts included those encoding cytosolic sensors of DNA, ZBP-1,IRF-3, and NFkB as well as genes involved in amplification of the IFN response, such as IFNAR1, JAK/STAT, ISG15. In keeping with the RNA-seq data, we observe increased secretion of type I IFN and other cytokines in response to influenza in PDCs co-cultured with PBECs. The PDCs also primed Th1 responses in T cells. The enhanced response of PDCs co-cultured with PBECs was due to the action of growth factors, GMCSF, GCSF, and VEGF, which were secreted by PBECs on differentiation. These data highlight possible mechanisms to enhance the production of type-I IFN in the airways, which is critical for host defense against respiratory infections.

CMV colitis has been reported in immunocompromized patients with severe deficiency of CD4+ T cells and T cell functions. In this study we present an extensive immunological analysis in a patient with primary hypogammaglobulinemia and CMV colitis who had normal numbers of CD3+T, CD4+T and CD8+T cells, and normal T cell proliferative responses to mitogens and recall antigens. Naïve (TN), central (TCM), and effector (TEM) memory subsets of CD4+ and CD8+ T cells, Granzyme+ and Perforin+ CD8+ T cells, PD-1+ T cells, CD4 Treg, CD8 Treg, and CMV tetramer specific CD8+ T cells were analyzed with specific antibodies and isotype controls using multicolor flow cytometry. CD8 TEM, Granzyme+ and Perforin+, and PD-1 CD8+T cells were increased, whereas CD8 TN and CD8 TCM cells were decreased in the patient as compared to controls. CMV tetramer+ CD8+ T cells were decreased in the patient. These data demonstrate that a deficiency of CMV-specific CD8+ T cells even in the presence of normal CD4+ T cell numbers and normal T cell functions may predispose patients with primary hypogammaglobulinemia to CMV colitis.

Immune dysfunction is a hallmark of aging and is thought to be responsible for the age-associated diseases. Dendritic cells (DCs) of the immune system function as initiators and regulators of the immune responses. Recent studies have highlighted the division of labor between various DC subsets. CD1c+ DC subset has emerged as a major inducer of CD4 T cell response. There is a scarcity of information regarding the age-associated changes in the functions of DC subsets in the elderly. Here, we investigated the changes in transcriptional profile of CD1c+ DC subset from healthy aged and young individuals using RNA sequencing. Our results suggest that majority of the genes in DCs are upregulated with age. Glucose transport, GPCR, and potassium channel genes are all upregulated in DCs from aged as compared to young indicating an enhanced activation state of DCs from aged individuals. The expression of histones, small nucleolar RNA H/ACA box (SNORA) and small nucleolar RNA C/D/box (SNORD), and long non-coding RNA (lncRNA) is also substantially upregulated in the DCs from aged. In contrast, the antigen-presenting and energy generating pathways are downregulated. In summary, DCs from aged subjects display an activated state coupled with reduced antigen presentation which may be responsible for age-associate immune dysfunction.

IntroductionRegulatory lymphocytes (CD4+ T regulatory cells [Treg], CD8+ Treg, and B regulatory cells [Breg]) play a critical role in immune homeostasis and tolerance. Common variable immunodeficiency (CVID) is associated with increased susceptibility to infections and increased frequency of inflammatory and autoimmune diseases. CD4+ Treg cell abnormalities have been reported in CVID; however, CD8+ Treg cells have not been reported in CVID. The objective of this study was to evaluate CD4+ Treg and CD8+ Treg cells in CVID patients.MethodsIn 25 patients with CVID and age-matched healthy controls, Treg cells, evaluated in freshly isolated peripheral blood mononuclear cells (natural; nCD4+ Treg and nCD8+ Treg) and following in vitro activation with anti-CD3/CD28 monoclonal antibodies (induced; iCD4+ Treg and iCD8+ Treg) as well as Breg cells were analyzed with specific monoclonal antibodies and isotype controls using flow cytometry.ResultsThe proportions of nCD4+ Treg (CD4+ CD127low CD25high FoxP3+), iCD4+ Treg (CD4+ CD127low CD25high FoxP3+), iCD8+ Treg (CD8+ CD25high CD183+ FoxP3+), and Breg (CD19+ CD24high CD38high) lymphocytes were significantly lower in patients with CVID than in controls.ConclusionsAltered regulatory lymphocytes may play a role in the pathogenesis and autoimmunity and inflammation associated with CVID.

The hydroferrate fluid MRN-100, an iron-based compound with potent antioxidant characteristics, was examined to identify its possible anti-inflammatory effects on human dendritic cells (DCs) in vitro. Human monocyte-derived DCs were treated with MRN-100 at two concentrations (50 and 100 μL/mL) for 24 h and then stimulated with or without lipopolysaccharides (LPS). The expression of DC maturation markers was assessed by flow cytometry and the production of cytokines was determined by enzyme-linked immunosorbent assay (ELISA). Functional assay was performed by co-culturing MRN-100-treated and untreated DCs with allogeneic naïve CD4+ T cells and assaying the T cells' cytokine production. Results show that treatment with MRN-100 significantly upregulated the co-stimulatory molecules CD80 and CD86 and increased human leukocyte antigen-DR (HLA-DR) though not significantly. MRN-100 treatment also significantly increased the production of the anti-inflammatory cytokine interleukin (IL)-10. On the other hand, MRN-100 significantly induced the secretion of pro-inflammatory cytokines such as IL-6 only at high concentrations. Furthermore, DCs pretreated with MRN-100 and either stimulated or not with LPS were able to prime CD4+ T cells to secrete significant amounts of IL-10 while inhibiting the secretion of pro-inflammatory cytokine tumor necrosis factor (TNF)-α. These results indicate that MRN-100 is a powerful anti-inflammatory agent that promotes the generation of an anti-inflammatory immune response in vitro. MRN-100 could be beneficial for treating patients with inflammatory diseases, including arthritis and type 1 diabetes, and its potential benefits should be examined in clinical trials.

BackgroundProgressive T cell decline in aged humans is associated with a deficiency of naïve (TN) and central memory (TCM) T cells. We have previously reported increased Tumor necrosis factor-α (TNF-α)-induced apoptosis in TN and TCM T cells in aged humans; however, the molecular basis of increased apoptosis remains to be defined. Since expression of TNF receptors (TNFRs) was reported to be comparable in young and aged, we investigated signaling events downstream of TNFRs to understand the molecular basis of increased TNF-α-induced apoptosis in aged TN and TCM CD8+ cells.ResultsThe expression of TRAF-2 and RIP, phosphorylation of JNK, IKKα/β, and IκBα, and activation of NF-κB activation were significantly decreased in TN and TCM CD8+ cells from aged subjects as compared to young controls. Furthermore, expression of A20, Bcl-xL, cIAP1, and FLIP-L and FLIP-S was significantly decreased in TN and TCM CD8+ cells from aged subjects.ConclusionsThese data demonstrate that an impaired expression/function of molecules downstream TNFR signaling pathway that confer survival signals contribute to increased apoptosis of TN and TCM CD8+ cells in aged humans.

BackgroundThe immune system plays a major role in the pathogenesis of age-related dementia, including Alzheimer's disease (AD). An insight into age-associated changes in the immune response to amyloid-beta (Aβ) in individuals without AD may be beneficial in identifying mechanisms preventing accumulation of Aβ.MethodsWe examined the response of human monocyte-derived dendritic cells (DCs), T cells, and peripheral blood mononuclear cells (PBMCs) from healthy aged and young subjects to Aβ peptide 1-42, Aβ fibrils, and recombinant, nonaggregated tau-4 protein with a view to understand the role of peripheral immunity in AD.ResultsOur studies revealed that DCs from healthy aged subjects display weak reactivity towards the Aβ peptide and no reactivity towards Aβ fibrils and tau compared with their young counterparts. An analysis of old and young PBMCs revealed that there is no significant T-cell memory against Aβ peptide, fibrils, or tau. Remarkably, the plasma levels of IgM antibodies specific to Aβ peptide 1-42 were significantly increased in aged subjects compared with young subjects, while IgG levels were comparable. Aβ peptide-specific IgM and IgG levels were also determined in the plasma of AD subjects compared with age-matched controls to demonstrate that the immune response against Aβ is stronger in AD patients. A decline in Aβ peptide-specific IgM antibodies was observed in AD patients compared with age-matched controls. In contrast, the levels of IgG as well as interleukin-21, the major cytokine involved in class switching, were increased in AD and patients with mild cognitive impairment, indicating a strong immune response against Aβ.ConclusionsCollectively, low immunogenicity of Aβ in healthy controls may prevent inflammation while the generation of specific IgM antibodies may help in the clearance of Aβ in healthy subjects.

Biotin (vitamin B7) is essential for human health because of its involvement, as a cofactor, in a variety of critical cellular metabolic reactions. Previous studies have shown that biotin deficiency enhances inflammation, and certain chronic inflammatory diseases are associated with biotin deficiency; however, the mechanisms that mediate the association between biotin status and inflammation are not well understood. In this study, we examined the effect of biotin deficiency on human CD4+ T cell responses to determine their role in biotin deficiency-associated inflammation. Our investigations revealed that anti-CD3/CD28-stimulated CD4+ T cells cultured in biotin-deficient medium secreted significantly enhanced levels of the proinflammatory cytokines IFN-γ, TNF, and IL-17. Expression of the transcription factors T-bet and RORγt was increased, whereas Foxp3 expression was decreased, in biotin-deficient CD4+ T cells. The percentage of T regulatory cells was also decreased under biotin-deficient condition. A similar increase in T-bet, RORγt, and proinflammatory cytokine levels, as well as a decrease in Foxp3, was observed in inguinal lymph nodes of mice fed a biotin-deficient diet relative to pair-fed controls. Furthermore, differentiation of CD4+ T cells toward Th1 and Th17 cells was also enhanced. In vitro and in vivo investigations indicated that the increased inflammatory response was due to enhanced activation of the mammalian target of rapamycin signaling pathway in biotin-deficient CD4+ T cells. In summary, these results demonstrate that biotin deficiency enhances the inflammatory responses in CD4+ T cells, which may contribute to inflammation associated with biotin deficiency.

Marina crystal minerals (MCM) are a mixture that contains crystallized minerals along with trace elements extracted from seawater. It is a nutritional supplement that is capable of enhancing natural killer (NK) cell activity and increasing T and B cell proliferation in humans post ingestion. However, its effect on dendritic cells (DCs), the cells that bridge innate and adaptive immunity, is not yet known. In this study, we examine the stimulatory effects of MCM on DCs' maturation and function in vitro. Human monocyte-derived DCs were treated with MCM at two different concentrations (10 and 20 µg/mL) for 24 h. Results showed that MCM treatment activated DCs in a dose-dependent fashion. It caused the upregulation of costimulatory molecules CD80, CD86, and HLA-DR, and prompted the production of DC cytokines, including interleukin (IL)-6, IL-10, tumor necrosis factor (TNF)-α, and IL-1β, and chemokines (monocyte chemotactic protein-1 (MCP-1)) and interferon-gamma-inducible protein-10 (IP-10). In addition, activated DCs primed CD4+ T cells to secrete significant amounts of interferon gamma (IFN-γ), and they also stimulated CD8+ T cells to express higher amounts of CD107a. These results indicate that MCM is a potentially powerful adjuvant, from natural materials, that activates human DCs in vitro and therefore may suggest its possible use in immune-based therapies against cancer and viral infections.

IgG4-RD is a multisystem fibroinflammatory disease characterized by the infiltration of tissues by IgG4 plasma cells. Combined skin and biliary tract involvement in IgG4-RD has not been described. We present perhaps the most comprehensive analysis of lymphocyte subsets in the first case of IgG4-related generalized skin rash and first case of combined skin and biliary tract manifestations. A 55-year-old male presented with painful jaundice and generalized macular pigmented pruritic eruptions, and CT abdomen revealed biliary obstruction. Ampulla and skin biopsies were subjected to histology and immunostaining. Naïve, central memory (TCM), effector memory (TEM), terminally differentiated effector memory (TEMRA) subsets of CD4+ and CD8+ T cells, T follicular helper subsets, naïve, transitional, marginal zone (MZ), germinal center (GC), IgM memory, and class-switched memory (CSM) B cells, and T follicular regulatory, regulatory B cells, CD4 Treg, and CD8 Treg were analyzed. Serum IgG4 was elevated at 448 mg/dL. Ampula biopsy showed lamina propria fibrosis and increased IgG4-positive plasma cells. Skin punch biopsy showed lymphoplasmacytic infiltrates with a 67% ratio of IgG4+:IgG+ plasma cells. CD4+TN and CD4+TCM decreased, whereas CD4+TEM increased. Naïve B cells increased; transitional, MZ, CSM, GC B cells, and plasmablasts decreased compared to control. CD4 Treg increased, whereas CD8 Treg and Breg decreased. In conclusion, IgG-RD may present with combined biliary tract and generalized dermatological manifestations. Changes in regulatory lymphocytes suggest their role in the pathogenesis of IgG4-RD. [ABSTRACT FROM AUTHOR]

Nanofabrics are now being used in a wide range of products that come into direct contact with skin, including carpet, clothing, and medical fabrics. In the current study, we examined the effect of a dispersed aqueous mixture of nanodiamond (ND) and nanoplatinum (NP) (DPV576) on human primary keratinocytes with respect to transient receptor potential vanilloid (TRPV) channel expression, secretion of cytokines and production of nerve growth factor (NGF). Keratinocytes were treated with DPV576 at concentrations of 1:10 and 1:100 dilutions for 24 hours in vitro, and their activation of was determined by production of cytokines TNF-α, IL-1β, and prostaglandin (PGE2), and by production of NGF. Inhibitor experiments were carried out by incubating keratinocytes with the TRPV4-selective antagonist HC-067047. TRPV receptor expression (TRPV1, TRPV3 and TRPV4) on keratinocytes as well as changes in Ca2+ potential were examined by flow cytometry. DPV576 treatment of keratinocytes resulted in the following effects: (1) stimulation of keratinocytes as indicated by the significant secretion of cytokines IL-1β, TNF-α, and PGE2, an effect noted only at higher concentration (1:10); (2) significant decrease in the expression of TRPV4 on keratinocytes with a spike in the calcium flux, but no change in the expression of TRPV1 and TRPV3; (3) induction of cytokine secretion independent of TRPV4, as the addition of TRPV4 inhibitor had no significant effect on the cytokine production from keratinocytes; (4) induction of NGF secretion by keratinocytes. These results demonstrate that DPV576 activates keratinocytes via multiple signaling pathways which may reduce stress associated with inflammation, pain, and circadian rhythms. ND/NP-coated fabrics that target the modulation of local inflammation, pain, and circadian rhythms could potentially be of benefit to humans.

Dendritic cells (DCs) play an important role in advancing age-associated progressive decline in adaptive immune responses, loss of tolerance, and development of chronic inflammation. In aged humans, DCs secrete increased levels of pro-inflammatory cytokines and decreased levels of anti-inflammatory and immune-regulatory cytokines. This may contribute to both chronic inflammation and loss of tolerance in aging. Aged DCs also display increased immune response against self-antigens contributing further to both inflammation and loss of tolerance. The secretion of innate protective cytokines such as type I and III interferons is decreased, and the function of DCs in airway remodeling and inflammation in aged is also compromised. Furthermore, the capacity of DCs to prime T cell responses also seems to be affected. Collectively, these changes in DC functions contribute to the immune dysfunction and inflammation in the elderly. This review only focuses on age-associated changes in DC function in humans.

IgMFcR (FcμR) are expressed on B cell and B cell subsets. Mice deficient in secreted IgM and FcμR share properties of impaired specific antibody response and autoimmunity with patient with selective IgM deficiency (SIGMD). Intravenous immunoglobulin (IGIV) regulates immune response, including modulation of IgGFc receptors. However, there are no data on the expression of FcμR in patients with SIGMD, and the effects of IGIV on FcμR. In this study, we investigated FcμR expression in naïve marginal zone (MZ), IgM memory, and class-switched memory B cells in patients with selective IgM deficiency and healthy controls. Furthermore, we examined the direct effect of IGIV on FcμR expression and on the upregulation of FcμR by TLR2 agonist (Pam3). Finally, we examined the effect of IVIG on spontaneously produced IgM and natural IgM anti-phosphorylcholine (PC) antibodies by B cells and B1 cells. FcμR expression is significantly decreased in MZ B cells in patients with SIGMD as compared to control. IGIV, at immunomodulatory concentrations, inhibited FcμR upregulation by Pam3 in MZ B cells, and IgM-depleted IGIV inhibited spontaneous secretion of natural IgM anti-PC antibodies and not total IgM by B1 cells. These data suggest that decreased FcμR expression on MZ B cells may play a role in the pathogenesis of SIGMD, and an inhibition of TLR-2-induced upregulation of FcμR by IGIV may be one of the mechanisms of its anti-inflammatory action. IGIV-induced inhibition of natural IgM antibodies may be one of the mechanisms of IGIV-induced immunoregulation.

The water-soluble biotin (vitamin B7) is indispensable for normal human health. The vitamin acts as a cofactor for five carboxylases that are critical for fatty acid, glucose, and amino acid metabolism. Biotin deficiency is associated with various diseases, and mice deficient in this vitamin display enhanced inflammation. Previous studies have shown that biotin affects the functions of adaptive immune T and NK cells, but its effect(s) on innate immune cells is not known. Because of that and because vitamins such as vitamins A and D have a profound effect on dendritic cell (DC) function, we investigated the effect of biotin levels on the functions of human monocyte-derived DCs. Culture of DCs in a biotin-deficient medium (BDM) and subsequent activation with LPS resulted in enhanced secretion of the proinflammatory cytokines TNF-α, IL-12p40, IL-23, and IL-1β compared with LPS-activated DCs cultured in biotin-sufficient (control) and biotin-oversupplemented media. Furthermore, LPS-activated DCs cultured in BDM displayed a significantly higher induction of IFN-γ and IL-17 indicating Th1/Th17 bias in T cells compared with cells maintained in biotin control or biotin-oversupplemented media. Investigations into the mechanisms suggested that impaired activation of AMP kinase in DCs cultured in BDM may be responsible for the observed increase in inflammatory responses. In summary, these results demonstrate for the first time that biotin deficiency enhances the inflammatory responses of DCs. This may therefore be one of the mechanism(s) that mediates the observed inflammation that occurs in biotin deficiency.

Aged subjects display increased susceptibility to mucosal diseases. Retinoic Acid (RA) plays a major role in inducing tolerance in the mucosa. RA acts on Dendritic cells (DCs) to induce mucosal tolerance. Here we compared the response of DCs from aged and young individuals to RA with a view to understand the role of DCs in age-associated increased susceptibility to mucosal diseases. Our investigations revealed that compared to young DCs, RA stimulated DCs from aged subjects are defective in inducing IL-10 and T regulatory cells. Examinations of the underlying mechanisms indicated that RA exposure led to the upregulation of CD141 and GARP on DCs which rendered the DCs tolerogenic. CD141(hi), GARP(+) DCs displayed enhanced capacity to induce T regulatory cells compared to CD141(lo) and GARP(-) DCs. Unlike RA stimulated DCs from young, DCs from aged subjects exhibited diminished upregulation of both CD141 and GARP. The percentage of DCs expressing CD141 and GARP on RA treatment was significantly reduced in DCs from aged individuals. Furthermore, the remaining CD141(hi), GARP(+) DCs from aged individuals were also deficient in inducing T regs. In summary, reduced response of aged DCs to RA enhances mucosal inflammation in the elderly, increasing their susceptibility to mucosal diseases.

Primary selective IgM deficiency (SIGMD) is a rare and recently IUIS-recognized primary immunodeficiency disease with increased susceptibility to infections, allergy, and autoimmune diseases. The pathogenesis of selective IgM remains unclear. The objective of the study was to understand the pathogenesis of selective IgM deficiency via a comprehensive analysis of subsets of B cells, naïve and memory subsets of CD4+ and CD8+ T cells, and Breg, CD4Treg, and CD8Treg cells. Twenty adult patients with SIGMD (serum IgM 4 mg/dl-32 mg/dl) and age-and gender-matched healthy controls were studied. Naïve B cells, transitional B cells, marginal zone B cells, germinal center B cells, IgM memory B cells, switched memory B cells, plasmablasts, CD21(low) B cells, B1 cells, CXCR3+ naive and memory B cells; naïve, central memory, and effector memory subsets of CD4+ and CD8+ T cells, and CD4Treg, CD8Treg and Breg were phenotypically analyzed using multicolor flow cytometry. A significant increase in CD21(low), IgM memory B cells, Breg and CD8Treg, and a significant decreased in germinal center B cells, and CXCR3+ naïve and memory B cells were observed in SIGMD. These alterations in subsets of B cells, and Breg and CD8Treg cells may play a role in the pathogenesis of SIGMD.

Primary selective IgM deficiency (SIGMD) is a rare and recently IUIS-recognized primary immunodeficiency disease with increased susceptibility to infections, allergy, and autoimmune diseases. The pathogenesis of selective IgM remains unclear. The objective of the study was to understand the pathogenesis of selective IgM deficiency via a comprehensive analysis of subsets of B cells, naïve and memory subsets of CD4+ and CD8+ T cells, and Breg, CD4Treg, and CD8Treg cells. Twenty adult patients with SIGMD (serum IgM 4 mg/dl-32 mg/dl) and age-and gender-matched healthy controls were studied. Naïve B cells, transitional B cells, marginal zone B cells, germinal center B cells, IgM memory B cells, switched memory B cells, plasmablasts, CD21(low) B cells, B1 cells, CXCR3+ naive and memory B cells; naïve, central memory, and effector memory subsets of CD4+ and CD8+ T cells, and CD4Treg, CD8Treg and Breg were phenotypically analyzed using multicolor flow cytometry. A significant increase in CD21(low), IgM memory B cells, Breg and CD8Treg, and a significant decreased in germinal center B cells, and CXCR3+ naïve and memory B cells were observed in SIGMD. These alterations in subsets of B cells, and Breg and CD8Treg cells may play a role in the pathogenesis of SIGMD.

Lactobacilli have been widely studied for their probiotic effects and have been reported to function as antiviral and anticancer agents. However, the underlying mechanisms via immune modulation are poorly understood. PFT is a freeze dried compound of Lactobacillus kefiri P-IF with a unique composition and functionality. In this study, we examined the potential stimulatory effects of two concentrations (50 µg and 100 µg/mL) of PFT on human monocyte-derived dendritic cell (DC) function in vitro. Results showed that PFT upregulated the expression of DC surface co-stimulatory and maturation markers CD80, CD86, and HLADR in a concentration dependent manner. PFT at 100 µg/mL markedly increased the secretion of IL-6, IL-10, TNF-α, and IL-1β by DCs. This concentration of PFT also stimulated the production of antiviral cytokines, IFN-α and IFN-λ(IL29) in DCs. Additionally, PFT at 100 µg/mL activated moDCs prime CD4(+)T cells and significantly increased the levels of IL-10, IFN-γ, and TNF-α by 1.7, four, three-fold, respectively. Furthermore PFT-stimulated DCs were also effective in enhancing the cytotoxic potential of CD8(+)T cells via the induction of Granzyme-B and upregulation of CD107a, and CD103 expression, a marker of resident/regulatory CD8(+)T cells. These data suggest that PFT functions as a natural adjuvant for DC activation and thus may be used in DC-based vaccine strategies against viral infections and cancer.

Good Syndrome is an adult-onset combined immunodeficiency defined by hypogammaglobulinemia, low or absent number of B cells, T cell deficiency and thymic tumor. We have characterized CD8+ T cells from a patient with Good syndrome that presented with CD8+T-cell large granular lymphocytic leukemia (LGL). Characterization of peripheral blood CD8+ T cells revealed that majority of CD8+ T cells were terminally differentiated effector memory phenotype (TEMRA; CD8+CCR7-CD45RA+), and were PD-1high (CD279), ICOSlow (CD278), and granzymehigh. Almost all CD8+ T cells were IFN-γ+. CD8 Treg (CD8+CD183+CCR7+CD45RA-) were decreased. TEMRA phenotype along with CD279high, demonstrates that these are exhausted CD8+ T cells. This phenotype along with CD278low may also explain severe T cell functional deficiency in our patient. In the present patient, T-LGL appears to be a clonal expansion of CD279+granzyme+IFN-γ+CD8+TEMRA cells. To best of our knowledge this is the first case of CD8+T-cell LGL leukemia associated with Good syndrome.

Cutaneous non-disseminated, non-tuberculous mycobacterial infections have been reported in both immunocompetent and immunocompromised subjects. Systemic Mycobacterium avium intracellulaire (MAI) have been reported in non-HIV patients with Idiopathic CD4 lymphocytopenia. We report a comprehensive immunological analysis in syndrome of selective IgM deficiency and T lymphocytopenia (both CD4+ and CD8+) with disseminated cutaneous MAI infection. Naïve (TN) and Central memory (TCM) subsets of both CD4+ and CD8+ T cells were decreased, whereas terminally differentiated effector memory (TEMRA) subset of CD4+ and CD8+ T cells were markedly increased. IFN-γ producing T cells were markedly decreased. Although CD14(high)CD16- proinflammatory monocytes were modestly increased, IFN-γR+ monocytes were markedly decreased. The expression of TLR3, TLR5, TLR7, and TLR9 on monocytes was decreased. Germinal center B cells (CD19+IgD-CD38+CD27(lo)) and B1 cells (CD20+CD27+CD43+CD70-) were markedly decreased. A role of immune alterations, including B cells and antibodies in disseminated cutaneous MAI infection is discussed.

The effect of platelet derived growth factor (PDGF) on immune cells is not elucidated. Here, we demonstrate PDGF inhibited the maturation of human DCs and induced IL-10 secretion. Culture of PDGF-DCs with T cells induced the polarization of T cells towards FoxP3 expressing T regulatory cells that secreted IL-10. Gene expression studies revealed that PDGF induced the expression of C-type lectin like receptor member 2, (CLEC-2) receptor on DCs. Furthermore, DCs transfected with CLEC-2 induced T regulatory cells in DC-T cell co-culture. CLEC-2 is naturally expressed on platelets. Therefore, to confirm whether CLEC-2 is responsible for inducing the T regulatory cells, T cells were cultured with either CLEC-2 expressing platelets or soluble CLEC-2. Both conditions resulted in the induction of regulatory T cells. The generation of T regulatory cells was probably due to the binding of CLEC-2 with its ligand podoplanin on T cells, since crosslinking of podoplanin on the T cells also resulted in the induction of T regulatory cells. These data demonstrate that PDGF upregulates the expression of CLEC-2 on cells to induce T regulatory cells.