STING controls T cell memory fitness during infection through T cell-intrinsic and IDO-dependent mechanisms.

Stimulator of interferon genes (STING) signaling has been extensively studied in inflammatory diseases and cancer, while its role in T cell responses to infection is unclear. Using Listeria monocytogenes strains engineered to induce different levels of c-di-AMP, we found that high STING signals impaired T cell memory upon infection via increased Bim levels and apoptosis. Unexpectedly, reduction of TCR signal strength or T cell-STING expression decreased Bim expression, T cell apoptosis, and recovered T cell memory. We found that TCR signal intensity coupled STING signal strength to the unfolded protein response (UPR) and T cell survival. Under strong STING signaling, Indoleamine-pyrrole 2,3-dioxygenase (IDO) inhibition also reduced apoptosis and led to a recovery of T cell memory in STING sufficient CD8 T cells. Thus, STING signaling regulates CD8 T cell memory fitness through both cell-intrinsic and extrinsic mechanisms. These studies provide insight into how IDO and STING therapies could improve long-term T cell protective immunity.

Rapid intestinal and systemic metabolic reprogramming in an immunosuppressed environment.

Intrinsic metabolism shapes the immune environment associated with immune suppression and tolerance in settings such as organ transplantation and cancer. However, little is known about the metabolic activities in an immunosuppressive environment. In this study, we employed metagenomic, metabolomic, and immunological approaches to profile the early effects of the immunosuppressant drug tacrolimus, antibiotics, or both in gut lumen and circulation using a murine model. Tacrolimus induced rapid and profound alterations in metabolic activities within two days of treatment, prior to alterations in gut microbiota composition and structure. The metabolic profile and gut microbiome after seven days of treatment was distinct from that after two days of treatment, indicating continuous drug effects on both gut microbial ecosystem and host metabolism. The most affected taxonomic groups are Clostriales and Verrucomicrobiae (i.e., Akkermansia muciniphila), and the most affected metabolic pathways included a group of interconnected amino acids, bile acid conjugation, glucose homeostasis, and energy production. Highly correlated metabolic changes were observed between lumen and serum metabolism, supporting their significant interactions. Despite a small sample size, this study explored the largely uncharacterized microbial and metabolic events in an immunosuppressed environment and demonstrated that early changes in metabolic activities can have significant implications that may serve as antecedent biomarkers of immune activation or quiescence. To understand the intricate relationships among gut microbiome, metabolic activities, and immune cells in an immune suppressed environment is a prerequisite for developing strategies to monitor and optimize alloimmune responses that determine transplant outcomes.

Artificial intelligence-based prediction of lycopene content in raw tomatoes using physicochemical attributes.

INTRODUCTION: Lycopene consumption reduces risk and incidence of cancer and cardiovascular diseases. Tomatoes are a rich source of phytochemical compounds including lycopene as a major constituent. Lycopene estimation using high-performance liquid chromatography is time-consuming and expensive. OBJECTIVE: To develop artificial intelligence models for prediction of lycopene in raw tomatoes using 14 different physicochemical parameters including salinity, total dissolved solids (TDS), electrical conductivity (EC), firmness, pH, total soluble solids (TSS), titratable acidity (TA), colour values on Hunter scale (L, a, b), total phenolic content (TPC), total flavonoid content (TFC) and antioxidant activity (AOA). MATERIAL AND METHODS: The post-harvest data acquisition was collected through investigation for more than 100 raw tomatoes stored for 15 days. Linear multivariate regression (LMVR), principal component regression (PCR) and partial least squares regression (PLSR) models were developed by splitting data set into train and test datasets. The training of models was performed using 10-fold cross validation (CV). RESULTS: Principal component analysis showed strong positive association between lycopene, colour value 'a', TPC, TFC and AOA. The R2 (CV), root mean square error (RMSE) (CV) and RMSE (Test) for best LMVR model was observed to be at 0.70, 8.48 and 9.69 respectively. The PCR model revealed R2 (CV) at 0.59, RMSE (CV) at 8.91 and RMSE (Test) at 10.17 while PLSR model revealed R2 (CV) at 0.60, RMSE (CV) at 9.10 and RMSE (Test) at 10.11. CONCLUSION: Results of the present study show that epidemiological studies suggest fully ripened tomatoes are most beneficial for consumption to ensure recommended daily intake of lycopene content.

Role of lymph node stroma and microenvironment in T cell tolerance.

Lymph nodes (LNs) are at the cross roads of immunity and tolerance. These tissues are compartmentalized into specialized niche areas by lymph node stromal cells (LN SCs). LN SCs shape the LN microenvironment and guide immunological cells into different zones through establishment of a CCL19 and CCL21 gradient. Following local immunological cues, LN SCs modulate activity to support immune cell priming, activation, and fate. This review will present our current understanding of LN SC subsets roles in regulating T cell tolerance. Three major types of LN SC subsets, namely fibroblastic reticular cells, lymphatic endothelial cells, and blood endothelial cells, are discussed. These subsets serve as scaffolds to support and regulate T cell homeostasis. They contribute to tolerance by presenting peripheral tissue antigens to both CD4 and CD8 T cells. The role of LN SCs in regulating T cell migration and tolerance induction is discussed. Looking forward, recent advances in bioengineered materials and approaches to leverage LN SCs to induce T cell tolerance are highlighted, as are current clinical practices that allow for manipulation of the LN microenvironment to induce tolerance. Increased understanding of LN architecture, how different LN SCs integrate immunological cues and shape immune responses, and approaches to induce T cell tolerance will help further combat autoimmune diseases and graft rejection.

Mechanisms of exTreg induction.

CD4+ CD25+ Foxp3+ Tregs play an important role in the maintenance of the immune system by regulating immune responses and resolving inflammation. Tregs exert their function by suppressing other immune cells and mediating peripheral self-tolerance. Under homeostatic conditions, Tregs are stable T-cell populations. However, under inflammatory environments, Tregs are converted to CD4+ CD25low Foxp3low cells. These cells are termed "exTreg" or "exFoxp3" cells. The molecular mechanism of Treg transition to exTregs remains incompletely understood. Uncertainties might be explained by a lack of consensus of biological markers to define Treg subsets in general and exTregs in particular. In this review, we summarize known markers of Tregs and factors responsible for exTreg generation including cytokines, signaling pathways, transcription factors, and epigenetic mechanisms. We also identify studies demonstrating the presence of exTregs in various diseases and sources of exTregs. Understanding the biology of Treg transition to exTregs will help in designing Treg-based therapeutic approaches.

G-CSF promotes alloregulatory function of MDSCs through a c-Kit dependent mechanism.

Myeloid-derived suppressor cells (MDSC) are a heterogeneous population of immature myeloid cells that expand in inflammatory conditions including transplantation. MDSCs may be capable of controlling rejection. The critical mechanisms underlying MDSC mediated alloregulation remain unexplored. G-CSF potently stimulates MDSC expansion. We hypothesized that G-CSF-induced MDSCs use a novel mechanism to suppress T cell responses. G-CSF promoted expansion of MDSCs and enhanced their suppressive function against T cell proliferation. Gene expression analysis revealed MDSCs expanded with G-CSF upregulated immune-related genes, but downregulated proliferation-related genes when compared to naïve control MDSCs. The KIT oncogene, encoding the c-Kit (CD117) transmembrane tyrosine kinase receptor, was the most significantly increased in MDSCs expanded with G-CSF. c-Kit inhibition with both imatinib and monoclonal blocking antibody reduced expression of ARG-1, iNOS, PD-L1, and SAA3. Further, imatinib also reduced MDSC-mediated T cell suppression in vitro. Modulation of c-Kit activity may represent a therapeutic target for alloregulatory MDSCs.

Chronic rejection as a persisting phantom menace in organ transplantation: a new hope in the microbiota?

PURPOSE OF REVIEW: The microbiota plays an important role in health and disease. During organ transplantation, perturbations in microbiota influence transplant outcome. We review recent advances in characterizing microbiota and studies on regulation of intestinal epithelial barrier function and mucosal and systemic immunity by microbiota and their metabolites. We discuss implications of these interactions on transplant outcomes. RECENT FINDINGS: Metagenomic approaches have helped the research community identify beneficial and harmful organisms. Microbiota regulates intestinal epithelial functions. Signals released by epithelial cells or microbiota trigger pro-inflammatory or anti-inflammatory effects on innate and adaptive immune cells, influencing the structure and function of the immune system. Assessment and manipulation of microbiota can be used for biomarkers for diagnosis, prognosis, and therapy. SUMMARY: The bidirectional dialogue between the microbiota and immune system is a major influence on immunity. It can be targeted for biomarkers or therapy. Recent studies highlight a close association of transplant outcomes with microbiota, suggesting exciting potential avenues for management of host physiology and organ transplantation.

Myeloid-derived suppressor cells expand after transplantation and their augmentation increases graft survival.

Myeloid-derived suppressor cells (MDSCs) expand in an inflammatory microenvironment such as cancer and autoimmunity. To study if transplantation induces MDSCs and these cells regulate allograft survival, C57BL/6 donor hearts were transplanted into BALB/c recipients and endogenous MDSCs were characterized. The effects of adoptive transfer of transplant (tx), tumor (tm), and granulocyte-colony stimulating factor (g-csf)-expanded MDSCs or depletion of MDSC were assessed. MDSCs expanded after transplantation (1.7-4.6-fold) in the absence of immunosuppression, homed to allografts, and suppressed proliferation of CD4 T cells in vitro. Tx-MDSCs differed phenotypically from tm-MDSCs and g-csf-MDSCs. Among various surface markers, Rae-1 expression was notably low and TGF-ß receptor II was high in tx-MDSCs when compared to tm-MDSCs and g-csf-MDSCs. Adoptive transfer of these three MDSCs led to differential graft survival: control (6 days), tx-MDSCs (7.5 days), tm-MDSCs (9.5 days), and g-csf-MDSCs (19.5 days). In combination with anti-CD154 mAb, MDSCs synergistically extended graft survival from 40 days (anti-CD154 alone) to 86 days with tm-MDSCs and 132 days with g-csf-MDSCs. Early MDSC depletion (day 0 or 20), however, abrogated graft survival, but late depletion (day 25) did not. In conclusion, MDSCs expanded following transplantation, migrated to cardiac allografts, prolonged graft survival, and were synergistic with anti-CD154 mAb.

NFκB-Pim-1-Eomesodermin axis is critical for maintaining CD8 T-cell memory quality.

T-cell memory is critical for long-term immunity. However, the factors involved in maintaining the persistence, function, and phenotype of the memory pool are undefined. Eomesodermin (Eomes) is required for the establishment of the memory pool. Here, we show that in T cells transitioning to memory, the expression of high levels of Eomes is not constitutive but rather requires a continuum of cell-intrinsic NFκB signaling. Failure to maintain NFκB signals after the peak of the response led to impaired Eomes expression and a defect in the maintenance of CD8 T-cell memory. Strikingly, we found that antigen receptor [T-cell receptor (TCR)] signaling regulates this process through expression of the NFκB-dependent kinase proviral integration site for Moloney murine leukemia virus-1 (PIM-1), which in turn regulates NFκB and Eomes. T cells defective in TCR-dependent NFκB signaling were impaired in late expression of Pim-1, Eomes, and CD8 memory. These defects were rescued when TCR-dependent NFκB signaling was restored. We also found that NFκB-Pim-1 signals were required at memory to maintain memory CD8 T-cell longevity, effector function, and Eomes expression. Hence, an NFκB-Pim-1-Eomes axis regulates Eomes levels to maintain memory fitness.

IL-12 Signals through the TCR To Support CD8 Innate Immune Responses.

CD8 T cells must integrate antigenic and inflammatory signals to differentiate into efficient effector and memory T cells able to protect us from infections. The mechanisms by which TCR signaling and proinflammatory cytokine receptor signaling cooperate in these processes are poorly defined. In this study, we show that IL-12 and other proinflammatory cytokines transduce signals through the TCR signalosome in a manner that requires Fyn activity and self-peptide-MHC (self-pMHC) interactions. This mechanism is crucial for CD8 innate T cell functions. Loss of Fyn activity or blockade of self-pMHC interactions severely impaired CD8 T cell IFN-γ and NKG2D expression, proliferation, and cytotoxicity upon cytokine-mediated bystander activation. Most importantly, in the absence of self-pMHC interactions, CD8 memory T cells fail to undergo bystander activation upon an unrelated infection. Thus, CD8 T cell bystander activation, although independent of cognate Ag, still requires self-pMHC and TCR signaling.

Affinity chromatography: A versatile technique for antibody purification.

Antibodies continue to be extremely utilized entities in myriad applications including basic research, imaging, targeted delivery, chromatography, diagnostics, and therapeutics. At production stage, antibodies are generally present in complex matrices and most of their intended applications necessitate purification. Antibody purification has always been a major bottleneck in downstream processing of antibodies, due to the need of high quality products and associated high costs. Over the years, extensive research has focused on finding better purification methodologies to overcome this holdup. Among a plethora of different techniques, affinity chromatography is one of the most selective, rapid and easy method for antibody purification. This review aims to provide a detailed overview on affinity chromatography and the components involved in purification. An array of support matrices along with various classes of affinity ligands detailing their underlying working principles, together with the advantages and limitations of each system in purifying different types of antibodies, accompanying recent developments and important practical methodological considerations to optimize purification procedure are discussed.

Evolution of a cell culture-derived genotype 1a hepatitis C virus (H77S.2) during persistent infection with chronic hepatitis in a chimpanzee.

UNLABELLED: Persistent infection is a key feature of hepatitis C virus (HCV). However, chimpanzee infections with cell culture-derived viruses (JFH1 or related chimeric viruses that replicate efficiently in cell culture) have been limited to acute-transient infections with no pathogenicity. Here, we report persistent infection with chronic hepatitis in a chimpanzee challenged with cell culture-derived genotype 1a virus (H77S.2) containing 6 cell culture-adaptive mutations. Following acute-transient infection with a chimeric H77/JFH1 virus (HJ3-5), intravenous (i.v.) challenge with 10(6) FFU H77S.2 virus resulted in immediate seroconversion and, following an unusual 4- to 6-week delay, persistent viremia accompanied by alanine aminotransferase (ALT) elevation, intrahepatic innate immune responses, and diffuse hepatopathy. This first persistent infection with cell culture-produced HCV provided a unique opportunity to assess evolution of cell culture-adapted virus in vivo. Synonymous and nonsynonymous nucleotide substitution rates were greatest during the first 8 weeks of infection. Of 6 cell culture-adaptive mutations in H77S.2, Q1067R (NS3) had reverted to Q1067 and S2204I (NS5A) was replaced by T2204 within 8 weeks of infection. By 62 weeks, 4 of 6 mutations had reverted to the wild-type sequence, and all reverted to the wild-type sequence by 194 weeks. The data suggest H77S.2 virus has greater potential for persistence and pathogenicity than JFH1 and demonstrate both the capacity of a nonfit virus to persist for weeks in the liver in the absence of detectable viremia as well as strong selective pressure against cell culture-adaptive mutations in vivo. IMPORTANCE: This study shows that mutations promoting the production of infectious genotype 1a HCV in cell culture have the opposite effect and attenuate replication in the liver of the only fully permissive animal species other than humans. It provides the only example to date of persistent infection in a chimpanzee challenged with cell culture-produced virus and provides novel insight into the forces shaping molecular evolution of that virus during 5 years of persistent infection. It demonstrates that a poorly fit virus can replicate for weeks within the liver in the absence of detectable viremia, an observation that expands current concepts of HCV pathogenesis and that is relevant to relapses observed with direct-acting antiviral therapies.

Early Immunomodulatory Program Triggered by Protolerogenic Bifidobacterium pseudolongum Drives Cardiac Transplant Outcomes.

BACKGROUND: Despite ongoing improvements to regimens preventing allograft rejection, most cardiac and other organ grafts eventually succumb to chronic vasculopathy, interstitial fibrosis, or endothelial changes, and eventually graft failure. The events leading to chronic rejection are still poorly understood and the gut microbiota is a known driving force in immune dysfunction. We previously showed that gut microbiota dysbiosis profoundly influences the outcome of vascularized cardiac allografts and subsequently identified biomarker species associated with these differential graft outcomes. METHODS: In this study, we further detailed the multifaceted immunomodulatory properties of protolerogenic and proinflammatory bacterial species over time, using our clinically relevant model of allogenic heart transplantation. RESULTS: In addition to tracing longitudinal changes in the recipient gut microbiome over time, we observed that Bifidobacterium pseudolongum induced an early anti-inflammatory phenotype within 7 d, whereas Desulfovibrio desulfuricans resulted in a proinflammatory phenotype, defined by alterations in leukocyte distribution and lymph node (LN) structure. Indeed, in vitro results showed that B pseudolongum and D desulfuricans acted directly on primary innate immune cells. However, by 40 d after treatment, these 2 bacterial strains were associated with mixed effects in their impact on LN architecture and immune cell composition and loss of colonization within gut microbiota, despite protection of allografts from inflammation with B pseudolongum treatment. CONCLUSIONS: These dynamic effects suggest a critical role for early microbiota-triggered immunologic events such as innate immune cell engagement, T-cell differentiation, and LN architectural changes in the subsequent modulation of protolerant versus proinflammatory immune responses in organ transplant recipients.

Annexin A2 is involved in the formation of hepatitis C virus replication complex on the lipid raft.

The hepatitis C virus (HCV) RNA replicates in hepatic cells by forming a replication complex on the lipid raft (detergent-resistant membrane [DRM]). Replication complex formation requires various viral nonstructural (NS) proteins as well as host cellular proteins. In our previous study (C. K. Lai, K. S. Jeng, K. Machida, and M. M. Lai, J. Virol. 82:8838-8848, 2008), we found that a cellular protein, annexin A2 (Anxa2), interacts with NS3/NS4A. Since NS3/NS4A is a membranous protein and Anxa2 is known as a lipid raft-associated scaffold protein, we postulate that Anxa2 helps in the formation of the HCV replication complex on the lipid raft. Further studies showed that Anxa2 was localized at the HCV-induced membranous web and interacted with NS4B, NS5A, and NS5B and colocalized with them in the perinuclear region. The silencing of Anxa2 decreased the formation of membranous web-like structures and viral RNA replication. Subcellular fractionation and bimolecular fluorescence complementation analysis revealed that Anxa2 was partially associated with HCV at the lipid raft enriched with phosphatidylinositol-4-phosphate (PI4P) and caveolin-2. Further, the overexpression of Anxa2 in HCV-nonsusceptible HEK293 cells caused the enrichment of HCV NS proteins in the DRM fraction and increased the colony-forming ability of the HCV replicon. Since Anxa2 is known to induce the formation of the lipid raft microdomain, we propose that Anxa2 recruits HCV NS proteins and enriches them on the lipid raft to form the HCV replication complex.

Management of Non-union Distal Femur Fractures With Augmentation Nail Plate Construct.

Background and objective A non-union distal femur fracture is a challenging fracture to treat. Common treatment modalities for non-union distal femur fractures include dual plating, intramedullary nails, ilizarov, and hybrid fixators. Despite the availability of a wide armamentarium of constructs, the clinical and functional outcome of these modalities is often complicated by significant morbidity, joint stiffness, and delayed union. The augmentation of the intramedullary nail with a locking plate results in a robust architecture, improving the likelihood of union. The use of this nail plate construct improves biomechanical stability and restores limb alignment, which enables early rehabilitation and weight bearing and lowers the likelihood of fixation failure. Methodology A prospective study was conducted at the Government Institute of Medical Science, Greater Noida, from January 2021 to January 2022 on 10 patients with non-union of the distal femur. All the patients were operated on with nail plate construct. The minimum follow-up period was 12 months. Results A total of 10 patients with a mean age of 55 years were included. Six were earlier treated with an intramedullary nail and four with extramedullary implants. All patients were managed with implant removal and fixation with nail plate construct and bone grafting. The average duration of the union was 10.3 months. The International Knee Documentation Committee (IKDC) score improved from 30.6 preoperatively to 67.3 postoperatively. Only one patient developed a superficial infection, which was managed by wound debridement and targeted antibiotic therapy. Conclusion In our experience, this relatively novel technique of combining nail plate constructs offers encouraging outcomes in the management of non-union of distal femur fractures, especially in elderly and osteopenic patients.

Strain-specific alterations in gut microbiome and host immune responses elicited by tolerogenic Bifidobacterium pseudolongum.

The beneficial effects attributed to Bifidobacterium are largely attributed to their immunomodulatory capabilities, which are likely to be species- and even strain-specific. However, their strain-specificity in direct and indirect immune modulation remain largely uncharacterized. We have shown that B. pseudolongum UMB-MBP-01, a murine isolate strain, is capable of suppressing inflammation and reducing fibrosis in vivo. To ascertain the mechanism driving this activity and to determine if it is specific to UMB-MBP-01, we compared it to a porcine tropic strain B. pseudolongum ATCC25526 using a combination of cell culture and in vivo experimentation and comparative genomics approaches. Despite many shared features, we demonstrate that these two strains possess distinct genetic repertoires in carbohydrate assimilation, differential activation signatures and cytokine responses signatures in innate immune cells, and differential effects on lymph node morphology with unique local and systemic leukocyte distribution. Importantly, the administration of each B. pseudolongum strain resulted in major divergence in the structure, composition, and function of gut microbiota. This was accompanied by markedly different changes in intestinal transcriptional activities, suggesting strain-specific modulation of the endogenous gut microbiota as a key to immune modulatory host responses. Our study demonstrated a single probiotic strain can influence local, regional, and systemic immunity through both innate and adaptive pathways in a strain-specific manner. It highlights the importance to investigate both the endogenous gut microbiome and the intestinal responses in response to probiotic supplementation, which underpins the mechanisms through which the probiotic strains drive the strain-specific effect to impact health outcomes.

Valorisation of fruit peel bioactive into green synthesized silver nanoparticles to modify cellulose wrapper for shelf-life extension of packaged bread.

Fruit peels are rich source of bioactive compounds such as polyphenols, flavonoids, and antioxidants but are often discarded as waste due to limited pharmaceutical and nutraceutical applications. This study aimed to valorise pomegranate and citrus fruit peel into green synthesised silver nanoparticles (AgNPs) in order to modify cellulose-based wrapping material for prospective food packaging applications and propose an alternate and sustainable approach to replace polyethene based food packaging material. Four different concentrations of AgNO3 (0.5 mM, 1 mM, 2 mM, and 3 mM) were used for green synthesis of AgNPs from fruit peel bioactive, which were characterised followed by phytochemical analysis. Ultraviolet-Visible spectroscopy showed surface plasmon resonance at 420 nm, XRD analysis showed 2θ peak at 27.8°, 32.16°, 38.5°, 44.31°, 46.09°, 54.76°, 57.47°, 64.61° and 77.50° corresponding to (210), (122), (111), (200), (231), (142), (241), (220) and (311) plane of face centred cubic crystal structure of AgNPs. Fourier-transform infrared spectroscopy analysis of AgNPs green synthesised from pomegranate and kinnow peel extract showed a major peak at 3277, 1640 and 1250-1020 1/cm while a small peak at 2786 1/cm was observed in case of pomegranate peel extract which was negligible in AgNPs synthesized from kinnow peel extract. Particle sizes of AgNPs showed no statistically significant variance with p > 0.10 and thus, 2 mM was chosen for further experimentation and modification of cellulose based packaging material as it showed smallest average particle size. Zeta potential was observed to be nearly neutral with a partial negative strength due to presence of various phenolic compounds such as presence of gallic acid which was confirmed by ultrahigh performance liquid chromatography-photodiode array(UHPLC-PDA) detector. Thermal stability analysis of green synthesised AgNPs qualified the sterilisation conditions up to 100 °C. AgNPs green synthesized from both the peel extracts had higher polyphenolic content, antioxidant and radical scavenging activity as compared to peel extracts without treatment (p < 0.05). The cellulose based food grade packaging material was enrobed by green synthesised AgNPs. The characterisation of modified cellulose wrappers showed no significant difference in thickness of modified cellulose wrappers as compared with untreated cellulose wrapper (p > 0.42) while weight and grammage increased significantly in modified cellulose wrapper (p < 0.05). The colour values on CIE scale (L*, a* and b*) showed statistically significant increase in yellow and green colour (p < 0.05) for modified cellulose wrappers as compared to control wrapper. The oxygen permeability coefficient, water vapour permeability coefficient, water absorption capacity and water behaviour characteristics (water content, swelling degree and solubility) showed significant decrease (p < 0.05) for modified cellulose wrapper as compared to control wrapper. A uniform distribution and density of green synthesised AgNPs across cellulose wrapper matrix was observed through scanning electron microscopy (SEM) images with no significant aggregation, confirming successful enrobing and stable immobilisation of nanoparticles from cellulose matrix. A seven-day storage study of bread wrapped in modified and control cellulose wrappers showed delayed occurrence of microbial, yeast and mould count in bread packaged in modified cellulose wrappers and thus, resulting in shelf life extension of bread. The results are encouraging for the potential applications of modified cellulose wrappers to replace polyethene based food packaging.

Engineering the lymph node environment promotes antigen-specific efficacy in type 1 diabetes and islet transplantation.

Antigen-specific tolerance is a key goal of experimental immunotherapies for autoimmune disease and allograft rejection. This outcome could selectively inhibit detrimental inflammatory immune responses without compromising functional protective immunity. A major challenge facing antigen-specific immunotherapies is ineffective control over immune signal targeting and integration, limiting efficacy and causing systemic non-specific suppression. Here we use intra-lymph node injection of diffusion-limited degradable microparticles that encapsulate self-antigens with the immunomodulatory small molecule, rapamycin. We show this strategy potently inhibits disease during pre-clinical type 1 diabetes and allogenic islet transplantation. Antigen and rapamycin are required for maximal efficacy, and tolerance is accompanied by expansion of antigen-specific regulatory T cells in treated and untreated lymph nodes. The antigen-specific tolerance in type 1 diabetes is systemic but avoids non-specific immune suppression. Further, microparticle treatment results in the development of tolerogenic structural microdomains in lymph nodes. Finally, these local structural and functional changes in lymph nodes promote memory markers among antigen-specific regulatory T cells, and tolerance that is durable. This work supports intra-lymph node injection of tolerogenic microparticles as a powerful platform to promote antigen-dependent efficacy in type 1 diabetes and allogenic islet transplantation.

FRC transplantation restores lymph node conduit defects in laminin α4-deficient mice.

Fibroblastic reticular cells (FRCs) play important roles in tolerance by producing laminin α4 (Lama4) and altering lymph node (LN) structure and function. The present study revealed the specific roles of extracellular matrix Lama4 in regulating LN conduits using FRC-specific KO mouse strains. FRC-derived Lama4 maintained conduit fiber integrity, as its depletion altered conduit morphology and structure and reduced homeostatic conduit flow. Lama4 regulated the lymphotoxin ß receptor (LTßR) pathway, which is critical for conduit and LN integrity. Depleting LTßR in FRCs further reduced conduits and impaired reticular fibers. Lama4 was indispensable for FRC generation and survival, as FRCs lacking Lama4 displayed reduced proliferation but upregulated senescence and apoptosis. During acute immunization, FRC Lama4 deficiency increased antigen flow through conduits. Importantly, adoptive transfer of WT FRCs to FRC Lama4-deficient mice rescued conduit structure, ameliorated Treg and chemokine distribution, and restored transplant allograft acceptance, which were all impaired by FRC Lama4 depletion. Single-cell RNA sequencing analysis of LN stromal cells indicated that the laminin and collagen signaling pathways linked crosstalk among FRC subsets and endothelial cells. This study demonstrated that FRC Lama4 is responsible for maintaining conduits by FRCs and can be harnessed to potentiate FRC-based immunomodulation.

Rapid intestinal and systemic metabolic reprogramming in an immunosuppressed environment.

Intrinsic metabolism shapes the immune environment associated with immune suppression and tolerance in settings such as organ transplantation and cancer. However, little is known about the metabolic activities in an immunosuppressive environment. In this study, we employed metagenomic, metabolomic, and immunological approaches to profile the early effects of the immunosuppressant drug tacrolimus, antibiotics, or both in gut lumen and circulation using a murine model. Tacrolimus induced rapid and profound alterations in metabolic activities within two days of treatment, prior to alterations in gut microbiota composition and structure. The metabolic profile and gut microbiome after seven days of treatment was distinct from that after two days of treatment, indicating continuous drug effects on both gut microbial ecosystem and host metabolism. The most affected taxonomic groups are Clostriales and Verrucomicrobiae (i.e., Akkermansia muciniphila), and the most affected metabolic pathways included a group of interconnected amino acids, bile acid conjugation, glucose homeostasis, and energy production. Highly correlated metabolic changes were observed between lumen and serum metabolism, supporting their significant interactions. Despite a small sample size, this study explored the largely uncharacterized microbial and metabolic events in an immunosuppressed environment and demonstrated that early changes in metabolic activities can have significant implications that may serve as antecedent biomarkers of immune activation or quiescence. To understand the intricate relationships among gut microbiome, metabolic activities, and immune cells in an immune suppressed environment is a prerequisite for developing strategies to monitor and optimize alloimmune responses that determine transplant outcomes.