Expanding IPEX: Inborn Errors of Regulatory T Cells.

Regulatory T cells (Tregs) are critical for enforcing peripheral tolerance. Monogenic "Tregopathies" affecting Treg development, stability, and/or function commonly present with polyautoimmunity, atopic disease, and infection. While autoimmune manifestations may present in early childhood, as more disorders are characterized, conditions with later onset have been identified. Treg numbers in the blood may be decreased in Tregopathies, but this is not always the case, and genetic testing should be pursued when there is high clinical suspicion. Currently, hematopoietic cell transplantation is the only curative treatment, but gene therapies are in development, and small molecule inhibitors/biologics may also be used.

Type I interferon signature and cycling lymphocytes in macrophage activation syndrome.

BACKGROUNDMacrophage activation syndrome (MAS) is a life-threatening complication of Still's disease (SD) characterized by overt immune cell activation and cytokine storm. We aimed to further understand the immunologic landscape of SD and MAS.METHODWe profiled PBMCs from people in a healthy control group and patients with SD with or without MAS using bulk RNA-Seq and single-cell RNA-Seq (scRNA-Seq). We validated and expanded the findings by mass cytometry, flow cytometry, and in vitro studies.RESULTSBulk RNA-Seq of PBMCs from patients with SD-associated MAS revealed strong expression of genes associated with type I interferon (IFN-I) signaling and cell proliferation, in addition to the expected IFN-γ signal, compared with people in the healthy control group and patients with SD without MAS. scRNA-Seq analysis of more than 65,000 total PBMCs confirmed IFN-I and IFN-γ signatures and localized the cell proliferation signature to cycling CD38+HLA-DR+ cells within CD4+ T cell, CD8+ T cell, and NK cell populations. CD38+HLA-DR+ lymphocytes exhibited prominent IFN-γ production, glycolysis, and mTOR signaling. Cell-cell interaction modeling suggested a network linking CD38+HLA-DR+ lymphocytes with monocytes through IFN-γ signaling. Notably, the expansion of CD38+HLA-DR+ lymphocytes in MAS was greater than in other systemic inflammatory conditions in children. In vitro stimulation of PBMCs demonstrated that IFN-I and IL-15 - both elevated in MAS patients - synergistically augmented the generation of CD38+HLA-DR+ lymphocytes, while Janus kinase inhibition mitigated this response.CONCLUSIONMAS associated with SD is characterized by overproduction of IFN-I, which may act in synergy with IL-15 to generate CD38+HLA-DR+ cycling lymphocytes that produce IFN-γ.

Development of a Screening Algorithm for Lung Disease in Systemic Juvenile Idiopathic Arthritis.

OBJECTIVE: Lung disease (LD) is an increasingly recognized complication of systemic juvenile idiopathic arthritis (sJIA). As there are no currently available guidelines for pulmonary screening in sJIA, we sought to develop such an algorithm at our institution. METHODS: A multidisciplinary workgroup was convened, including members representing rheumatology, pulmonary, stem cell transplantation, and patient families. The workgroup leaders drafted an initial algorithm based on published literature and experience at our center. A modified Delphi approach was used to achieve agreement through three rounds of anonymous, asynchronous voting and a consensus meeting. Statements approved by the workgroup were rated as appropriate with moderate or high levels of consensus. These statements were organized into the final approved screening algorithm for LD in sJIA. RESULTS: The workgroup ultimately rated 20 statements as appropriate with a moderate or high level of consensus. The approved algorithm recommends pulmonary screening for newly diagnosed patients with sJIA with clinical features that the workgroup agreed may confer increased risk for LD. These "red flag features" include baseline characteristics (young age of sJIA onset, human leukocyte antigen type, trisomy 21), high disease activity (macrophage activation syndrome [MAS], sJIA-related ICU admission, elevated MAS biomarkers), respiratory symptoms or abnormal pulmonary examination findings, and features of drug hypersensitivity-like reactions (eosinophilia, atypical rash, anaphylaxis). The workgroup achieved consensus on the recommended pulmonary work-up and monitoring guidelines. CONCLUSION: We developed a pulmonary screening algorithm for sJIA-LD through a multidisciplinary consensus-building process, which will be revised as our understanding of sJIA-LD continues to evolve.

Real-world experience with low-dose IL-2 for children and young adults with refractory chronic graft-versus-host disease.

The majority of patients with chronic graft-versus-host disease (cGVHD) are steroid refractory (SR), creating a need for safe and effective therapies. Subcutaneous low-dose interleukin-2 (LD IL-2), which preferentially expands CD4+ regulatory T cells (Tregs), has been evaluated in 5 clinical trials at our center with partial responses (PR) in ∼50% of adults and 82% of children by week 8. We now report additional real-world experience with LD IL-2 in 15 children and young adults. We conducted a retrospective chart review of patients with SR-cGVHD at our center who received LD IL-2 from August 2016 to July 2022 not on a research trial. The median age at start of LD IL-2 was 10.4 years (range, 1.2-23.2 years) at a median of 234 days from cGVHD diagnosis (range, 11-542 days). Patients had a median of 2.5 (range, 1-3) active organs at LD IL-2 start and received a median of 3 (range, 1-5) prior therapies. The median duration of LD IL-2 therapy was 462 days (range, 8-1489 days). Most patients received 1 × 106 IU/m2 per day. There were no serious adverse effects. The overall response rate in 13 patients who received >4 weeks of therapy was 85% (complete response, n = 5; PR, n = 6) with responses in diverse organs. Most patients significantly weaned corticosteroids. Tregs preferentially expanded with a median peak fold increase of 2.8 in the ratio of Tregs to CD4+ conventional T cells (range, 2.0-19.8) by 8 weeks on therapy. LD IL-2 is a well-tolerated, steroid-sparing agent with a high response rate in children and young adults with SR-cGVHD.

Incidence and Risk Factors for Eosinophilia and Lung Disease in Biologic-Exposed Children With Systemic Juvenile Idiopathic Arthritis.

OBJECTIVE: Although interleukin-1 (IL-1)/IL-6 inhibitors are effective therapies for systemic juvenile idiopathic arthritis (JIA), some patients develop eosinophilia and lung disease during treatment. This study was undertaken to retrospectively evaluate incidence and risk factors for eosinophilia and describe lung disease outcomes in IL-1/IL-6 inhibitor-exposed patients with systemic JIA. METHODS: Among JIA patients at our institution exposed to interleukin-1 (IL-1)/IL-6 inhibitors (1995-2022), we compared incidence rate of eosinophilia in systemic JIA compared to other JIA, stratified by medication class (IL-1/IL-6 inhibitors, other cytokine inhibitors, methotrexate). We used Cox models to identify predictors of eosinophilia during IL-1/IL-6 inhibitor use and summarized treatment changes and outcomes after eosinophilia, including lung disease. HLA typing was performed on a clinical or research basis. RESULTS: There were 264 new medication exposures in 75 patients with systemic JIA and 41 patients with other JIA. A total of 49% of patients with systemic JIA with HLA typing (n = 45) were positive for HLA-DRB1*15 alleles. Eosinophilia was common during IL-1/IL-6 inhibitor use and did not differ by systemic JIA compared to other JIA (0.08 and 0.07 per person-year, respectively; P = 0.30). Among systemic JIA patients, pretreatment macrophage activation syndrome (MAS) was associated with a higher rate of subsequent eosinophilia on biologic therapy (unadjusted hazard ratio 3.2 [95% confidence interval 1.2-8.3]). A total of 4 of 5 patients who switched therapy within 10 weeks of eosinophilia experienced disease flare compared to none of the patients who continued the original therapy. A total of 8 of 25 patients with pulmonary evaluations had lung disease, and all had severe manifestations of systemic JIA (MAS, intensive care unit stay). One death was attributed to systemic JIA-lung disease. CONCLUSION: Eosinophilia is common in JIA patients using IL-1/IL-6 inhibitors. Severe disease may be associated with eosinophilia and lung disease in systemic JIA.

Long-Term Organ Function After HCT for SCD: A Report From the Sickle Cell Transplant Advocacy and Research Alliance.

Hematopoietic cell transplantation (HCT) is an established cure for sickle cell disease (SCD) supported by long-term survival, but long-term organ function data are lacking. We sought to describe organ function and assess predictors for dysfunction in a retrospective cohort (n = 247) through the Sickle cell Transplant Advocacy and Research alliance. Patients with <1-year follow-up or graft rejection/second HCT were excluded. Organ function data were collected from last follow-up. Primary measures were organ function, comparing pre- and post-HCT. Bivariable and multivariable analyses were performed for predictors of dysfunction. Median age at HCT was 9.4 years; the majority had HbSS (88.2%) and severe clinical phenotype (65.4%). Most received matched related (76.9%) bone marrow (83.3%) with myeloablative conditioning (MAC; 57.1%). Acute and chronic graft-versus-host disease (GVHD) developed in 24.0% and 24.8%. Thirteen patients (5.3%) died ≥1 year after HCT, primarily from GVHD or infection. More post-HCT patients had low ejection or shortening fractions than pre-HCT (0.6% â†’ 6.0%, P = .007 and 0% â†’ 4.6%, P = .003). The proportion with lung disease remained stable. Eight patients (3.2%) had overt stroke; most had normal (28.3%) or stable (50.3%) brain magnetic resonance imaging. On multivariable analysis, cardiac dysfunction was associated with MAC (odds ratio [OR] = 2.71; 95% confidence interval [CI], 1.09-6.77; P = .033) and severe acute GVHD (OR = 2.41; 95% CI, 1.04-5.62; P = .041). Neurologic events were associated with central nervous system indication (OR = 2.88; 95% CI, 2.00-4.12; P < .001). Overall organ dysfunction was associated with age ≥16 years (OR = 2.26; 95% CI, 1.35-3.78; P = .002) and clinically severe disease (OR = 1.64; 95% CI, 1.02-2.63; P = .043). In conclusion, our results support consideration of HCT at younger age and use of less intense conditioning.

Dysregulation of the cGAS-STING Pathway in Monogenic Autoinflammation and Lupus.

One of the oldest mechanisms of immune defense against pathogens is through detection of foreign DNA. Since human DNA is compartmentalized into the nucleus, its presence in the cytosol heralds a potential threat. The cGAS-STING pathway is one of the most important cytosolic DNA sensing pathways and leads to interferon signaling, inflammasome activation, autophagy, and cell death. While STING signaling is protective at physiologic levels, chronic activation of this pathway can instead drive autoinflammation and autoimmunity. Here we discuss several monogenic disorders of the STING pathway that highlight its impact on both innate and adaptive immunity in the progressive loss of tolerance. The potential relevance of STING signaling in systemic lupus erythematosus is then discussed with a focus on future avenues for monitoring and targeting this pathway.

Genetic diagnosis of immune dysregulation can lead to targeted therapy for interstitial lung disease: A case series and single center approach.

In recent years, a growing number of monogenic disorders have been described that are characterized by immune dysregulation. A subset of these "primary immune regulatory disorders" can cause severe interstitial lung disease, often recognized in late childhood or adolescence. Patients presenting to pulmonary clinic may have long and complex medical histories, but lack a unifying genetic diagnosis. It is crucial for pulmonologists to recognize features suggestive of multisystem immune dysregulation and to initiate genetic workup, since targeted therapies based on underlying genetics may halt or even reverse pulmonary disease progression. Through such an approach, our center has been able to diagnose and treat a cohort of patients with interstitial lung disease from gene defects that affect immune regulation. Here we present representative cases related to pathogenic variants in three distinct pathways and summarize disease manifestations and treatment approaches. We conclude with a discussion of our perspective on the outstanding challenges for diagnosing and managing these complex life-threatening and chronic disorders.

Features predicting colchicine efficacy in treatment of children with undefined systemic autoinflammatory disease: A retrospective cohort study.

OBJECTIVE: Patients with undefined systemic autoinflammatory diseases (uSAIDs) are challenging to manage, as there are no guidelines or recommendations for targeted therapy. We aimed to evaluate the efficacy of empiric treatment with colchicine in our single-center uSAID population in the United States, as well as the patient characteristics associated with the most robust colchicine response. METHODS: Children with uSAID 18 years old at initial evaluation during 2000-2019 were included if they received 3 months of colchicine therapy. Data on demographics, clinical features, laboratory/ genetic studies, and treatment responses were collected. Most statistics were based on chi-square analyses for categorical data. Complete response to colchicine was defined as resolution of episodes or the presence of minor residual symptoms that did not require any further therapy. A partial response was defined as a decrease in the frequency, severity, or length of episodes but still necessitating additional therapy. Patients were considered nonresponders if they did not experience any improvement with colchicine at target therapeutic dosing. RESULTS: We identified 133 children diagnosed with uSAID who met our inclusion criteria. The median time to starting empiric colchicine was 5 months from the diagnosis of autoinflammatory disease. 92.5% (n = 123) of patients had a beneficial response to colchicine, including 46.6% (n = 62) partial responders and 45.9% (n = 61) complete responders. The presence of a nonurticarial rash was associated with an incomplete colchicine response (29.2% (n = 21) vs 13.1% (n = 8), P = .025). The presence of a heterozygous MEFV mutation in patients who did not fit Familial Mediterranean Fever diagnostic criteria (n = 25) appeared to be associated with a greater likelihood of complete colchicine response, although this was not statistically significant (62.5% (n = 14) vs 42.6% (n =11), P = .08). In MEFV mutation-negative patients, a nonurticarial rash was even more strongly associated with incomplete colchicine response, with an OR of 27.53 (CI [1.59-477], P = .023). The presence of oral ulcers also corresponded to incomplete colchicine response, although this did not reach clinical significance (38.9% (n = 28) vs 24.6% (n = 15), P = .08). There was no significant association between episode duration or frequency and colchicine response. CONCLUSION: Colchicine leads to clinical benefits in most children with uSAID. We, thus, recommend an early trial of colchicine in newly diagnosed patients with uSAID.

Disordered T cell-B cell interactions in autoantibody-positive inflammatory arthritis.

T peripheral helper (Tph) cells, identified in the synovium of adults with seropositive rheumatoid arthritis, drive B cell maturation and antibody production in non-lymphoid tissues. We sought to determine if similarly dysregulated T cell-B cell interactions underlie another form of inflammatory arthritis, juvenile oligoarthritis (oligo JIA). Clonally expanded Tph cells able to promote B cell antibody production preferentially accumulated in the synovial fluid (SF) of oligo JIA patients with antinuclear antibodies (ANA) compared to autoantibody-negative patients. Single-cell transcriptomics enabled further definition of the Tph gene signature in inflamed tissues and showed that Tph cells from ANA-positive patients upregulated genes associated with B cell help to a greater extent than patients without autoantibodies. T cells that co-expressed regulatory T and B cell-help factors were identified. The phenotype of these Tph-like Treg cells suggests an ability to restrain T cell-B cell interactions in tissues. Our findings support the central role of disordered T cell-help to B cells in autoantibody-positive arthritides.

Low CD4 count may be a risk factor for non-tuberculous mycobacteria infection in pediatric hematopoietic cell transplant recipients.

BACKGROUND: HCT leaves patients in a relative state of immune deficiency both during their initial transplant admission and for several years following discharge. NTM are generally harmless colonizers of the outside environment, but for immunocompromised patients, they can cause significant disease due to a paucity of T-cell defense. While routine prophylaxis against NTM is recommended for patients with low CD4 counts in certain clinical settings (eg, AIDS), this is not yet established for HCT patients despite their higher risk. METHODS: Here we build upon our prior work to determine risk factors for NTM in pediatric HCT patients by comparing NTM patient characteristics to matched HCT controls. RESULTS: We followed 272 patients across a 13-year time period, with 11 cases of NTM. Patients with NTM had a significantly lower CD4 count at Day 365 than matched HCT controls (105.5 ± 97.0 cells/µl vs. 856.2 ± 446.1 cells/µl, respectively; p = .001). No other potential risk factors (eg, CMV, GvHD, disease type) were found to be statistically significant, including use of T-cell depleting agents. This is consistent with an average diagnosis of NTM at Day +323 (ie, outside immediate post-transplant period). All-cause mortality was similar between NTM and control HCT groups, with an NTM attributable mortality of <10%. CONCLUSION: Since reduced CD4 counts are associated with NTM, and cost and morbidity are high, azithromycin prophylaxis for CD4 count <200 cells/µl in high-risk patients should be considered.

Mesenchymal stromal cells: Getting ready for clinical primetime.

Mesenchymal stromal cells (MSC) are multipotent precursor cells that can be derived from a variety of tissue sources, with a working definition based on immunophenotyping and cell differentiation capacity. Despite historical roots in the field of tissue engineering, they have generated great interest as cell therapies for their immune regulatory function, which has led to numerous clinical trials for a range of inflammatory and autoimmune conditions. Importantly, due to the lack of traditional MHC expression and their expression of other immune regulatory proteins, they can be used from third party donors without generating a dangerous alloreactivity. After 20 years of clinical trials, they have earned themselves an excellent safety record but are currently only approved for use in Canada, New Zealand, Japan, South Korea and Europe due to a lack of consistent efficacy data. In the United States, the indication that has seen the most progress is steroid refractory acute graft-versus-host disease (SR-aGVHD). Issues with early clinical trials can be attributed to both challenges with defining optimal patient populations and trial design as well as limitations related to commercial manufacturing. Earlier this year, the encouraging data for a repeat Phase III trial in pediatric patients with SR-aGVHD was published. This review provides information on the proposed mechanism of action of MSCs, clinical utilization of MSCs with focus on SR-aGVHD and potential modalities that can improve the efficacy of MSCs.

Risk Factors, Clinical Outcomes, and Cost-of-Care Related to Graft Failure in Pediatric Allogeneic Hematopoietic Cell Transplant Recipients.

Allogeneic hematopoietic cell transplantation (HCT) has the capacity to cure numerous malignant and nonmalignant disorders. A dreaded complication is graft failure (GF), as it puts patients at high risk of infection and disease relapse. There are few contemporary data on the risks, outcomes, and economic burden of GF in pediatric patients. In this study, we address this gap by focusing on 14 years of transplant at our single center, for which data are compared in 2 time periods: 2005 to 2010 (n = 146) and 2011 to 2018 (n = 144). In the 290 patients studied, the median age was 9.33 years, and 50.3% had malignant versus nonmalignant disease. Cell source included bone marrow (51%), cord blood (19.7%), unmanipulated peripheral blood stem cells (PBSCs; 12.1%), and CD34-selected PBSCs (17.2%). Twenty-one percent of patients had reduced-intensity conditioning (RIC), and 54.8% of transplants were fully HLA matched. Most patients received serotherapy with rabbit anti-thymocyte globulin (39.3%) or alemtuzumab (42.8%). The incidence of neutropenic and non-neutropenic GF (NGF and NNGF) was 6.6% and 3.8%, respectively. Multivariate analysis demonstrated alemtuzumab (odds ratio [OR], 6.256, P < .001) was the main variable associated with a higher rate of GF in both time periods, whereas RIC (OR, 11.8, P < .001) and cell source (CD34-selected PBSCs; OR, 4.22, P = .04) showed period-specific effects. Specifically, from time periods 1 to 2, cord blood transplants were discontinued at our center, with a concomitant increase in CD34-selected grafts and a shift from more episodes of NGF to NNGF. Overall survival was 69% in the entire HCT cohort and 50% among patients with GF. Survival among GF patients improved from time periods 1 to 2 (20% versus 80%, P = .001), potentially due to a higher incidence of NNGF and increased ability to perform stem cell boosts from the same donor once cord blood transplants were phased out. Inpatient length of stay was consistently higher for patients with GF. Similar trends were seen for inpatient costs, although improvements were seen in our entire HCT population over time. In summary, GF remains a significant challenge in pediatric HCT and poses an economic burden on the health care system.

When Local Bone Pain Is Just the Tip of the Iceberg-A Case Report of Three Patients With Chronic Multifocal Recurrent Osteomyelitis and Some Red Flags to Help Make the Diagnosis.

Chronic recurrent multifocal osteomyelitis (CRMO) is an uncommon cause of chronic inflammatory bone pain in children that can be disabling. Often, this diagnosis is considered only after a prolonged workup, leading to frustration for families and unnecessary interventions for patients. Here we describe three cases of CRMO to increase awareness of how it may present. The first patient had a typical presentation of focal bone pain (knee), for which she underwent bone scan (hint of >1 lesion), had a bone biopsy to rule out malignancy, received empiric antibiotics for presumed infection, and finally had whole-body imaging confirming CRMO when symptoms persisted. The second patient had a similar workup, but initially presented with clavicular pain. This location should raise suspicion for CRMO, as it is an uncommon location for infectious osteomyelitis. The third patient presented with delayed growth and right hip pain, and simultaneously developed palmoplantar pustulosis. These secondary findings can also serve as red flags for CRMO, as it has been linked to this skin condition and inflammatory bowel disease. All patients improved on non-steroidal anti-inflammatory (NSAID) medications, methotrexate, and/or tumor necrosis factor (TNF)-α antagonists. By raising awareness of clinical findings suggestive of CRMO, this report may help expedite diagnosis, so patients can be started on anti-inflammatory therapy.

Engineering of human cardiac muscle electromechanically matured to an adult-like phenotype.

The application of tissue-engineering approaches to human induced pluripotent stem (hiPS) cells enables the development of physiologically relevant human tissue models for in vitro studies of development, regeneration, and disease. However, the immature phenotype of hiPS-derived cardiomyocytes (hiPS-CMs) limits their utility. We have developed a protocol to generate engineered cardiac tissues from hiPS cells and electromechanically mature them toward an adult-like phenotype. This protocol also provides optimized methods for analyzing these tissues' functionality, ultrastructure, and cellular properties. The approach relies on biological adaptation of cultured tissues subjected to biomimetic cues, applied at an increasing intensity, to drive accelerated maturation. hiPS cells are differentiated into cardiomyocytes and used immediately after the first contractions are observed, when they still have developmental plasticity. This starting cell population is combined with human dermal fibroblasts, encapsulated in a fibrin hydrogel and allowed to compact under passive tension in a custom-designed bioreactor. After 7 d of tissue formation, the engineered tissues are matured for an additional 21 d by increasingly intense electromechanical stimulation. Tissue properties can be evaluated by measuring contractile function, responsiveness to electrical stimuli, ultrastructure properties (sarcomere length, mitochondrial density, networks of transverse tubules), force-frequency and force-length relationships, calcium handling, and responses to ß-adrenergic agonists. Cell properties can be evaluated by monitoring gene/protein expression, oxidative metabolism, and electrophysiology. The protocol takes 4 weeks and requires experience in advanced cell culture and machining methods for bioreactor fabrication. We anticipate that this protocol will improve modeling of cardiac diseases and testing of drugs.

Regeneration of severely damaged lungs using an interventional cross-circulation platform.

The number of available donor organs limits lung transplantation, the only lifesaving therapy for the increasing population of patients with end-stage lung disease. A prevalent etiology of injury that renders lungs unacceptable for transplantation is gastric aspiration, a deleterious insult to the pulmonary epithelium. Currently, severely damaged donor lungs cannot be salvaged with existing devices or methods. Here we report the regeneration of severely damaged lungs repaired to meet transplantation criteria by utilizing an interventional cross-circulation platform in a clinically relevant swine model of gastric aspiration injury. Enabled by cross-circulation with a living swine, prolonged extracorporeal support of damaged lungs results in significant improvements in lung function, cellular regeneration, and the development of diagnostic tools for non-invasive organ evaluation and repair. We therefore propose that the use of an interventional cross-circulation platform could enable recovery of otherwise unsalvageable lungs and thus expand the donor organ pool.

Dual IFN-γ/hypoxia priming enhances immunosuppression of mesenchymal stromal cells through regulatory proteins and metabolic mechanisms.

The immunosuppressive capacity of human mesenchymal stromal cells (MSCs) renders them promising candidates for treating diverse immune disorders. However, after hundreds of clinical trials, there are still no MSC therapies approved in the United States. MSCs require specific cues to adopt their immunosuppressive phenotype, and yet most clinical trials use cells expanded in basic culture medium and growth conditions. We propose that priming MSCs prior to administration will improve their therapeutic efficacy. Interferon-gamma (IFN-γ) priming are cues common to situations of immune escape that have individually shown promise as MSC priming cues but have not been systematically compared. Using mixed lymphocyte reactions, we show that priming MSCs with either cue alone improves T-cell inhibition. However, combining the two cues results in additive effects and markedly enhances the immunosuppressive phenotype of MSCs. We demonstrate that IFN-γ induces expression of numerous immunosuppressive proteins (IDO, PD-L1, HLA-E, HLA-G), whereas hypoxia switches MSCs to glycolysis, causing rapid glucose consumption and production of T-cell inhibitory lactate levels. Dual IFN-γ/hypoxia primed MSCs display both attributes and have even higher induction of immunosuppressive proteins over IFN-γ priming alone (IDO and HLA-G), which may reflect another benefit of metabolic reconfiguration.

The influence of hypoxia and IFN-γ on the proteome and metabolome of therapeutic mesenchymal stem cells.

Over the past 15 years, mesenchymal stem cells (MSCs) have been assessed for their capacity to suppress inflammation and promote tissue repair. Regardless of whether the cells are primed (exposed to instructive cues) before administration, their phenotype will respond to environmental signals present in the pathophysiological setting being treated. Since hypoxia and inflammation coexist in the settings of acute injury and chronic disease we sought to explore how the proteome and metabolome of MSCs changes when cells were exposed to 48 h of 1% oxygen, interferon gamma (IFN-γ), or both cues together. We specifically focused on changes in cell metabolism, immune modulation, extracellular matrix secretion and modification, and survival capacity. IFN-γ promoted expression of anti-pathogenic proteins and induced MSCs to limit inflammation and fibrosis while promoting their own survival. Hypoxia instead led to cell adaptation to low oxygen, including upregulation of proteins involved in anaerobic metabolism, autophagy, angiogenesis, and cell migration. While dual priming resulted in additive effects, we also found many instances of synergy. These data lend insight to how MSCs may behave after administration to a patient and suggest how priming cells beforehand could improve their therapeutic capacity.

Paracrine Effects of Mesenchymal Stromal Cells Cultured in Three-Dimensional Settings on Tissue Repair.

Mesenchymal stromal cells (MSCs) are a promising cell source for promoting tissue repair, due to their ability to release growth, angiogenic, and immunomodulatory factors. However, when injected as a suspension, these cells suffer from poor survival and localization, and suboptimal release of paracrine factors. While there have been attempts to overcome these limitations by modifying MSCs themselves, a more versatile solution is to grow them in three dimensions, as aggregates or embedded into biomaterials. Here we review the mechanisms by which 3D culture can influence the regenerative capacity of undifferentiated MSCs, focusing on recent examples from the literature. We further discuss how knowledge of these mechanisms can lead to strategic design of MSC therapies that overcome some of the challenges to their effective translation.