Negative impact of bone-marrow-derived mesenchymal stem cells on dextran sulfate sodium-induced colitis

AIM:To investigate the effects of mesenchymal stem cells(MSCs)on dextran sulfate sodium-induced inflammatory bowel disease(IBD).METHODS:C57BL/6 mice were fed 3.5%(g/L)dextran sulfate sodium.On day seven,the mice received intraperitoneal injections of 1×106 MSCs.The survival rate,disease activity index values,and body weight,were monitored daily.On day ten,colon lengths and histopathologic changes were assessed.In addition,immunoregulatory changes following MSC administration were evaluated by determining the levels of effector T cell responses in the spleen and mesenteric lymph nodes,and the expression levels of inflammatory cytokines in homogenized colons.RESULTS:Intraperitoneal administration of MSCs did not prevent development of colitis and did not reduce the clinicopathologic severity of IBD.No significant difference was evident in either survival rate or disease activity index score between the control and MSCtreated group.Day ten-sacrificed mice exhibited no significant difference in either colon length or histopathologic findings.Indeed,the MSC-treated group exhibited elevated levels of interleukin(IL)-6 and transforming growth factor-β,and a reduced level of IL-10,in spleens,mesenteric lymph nodes,and homogenized colons.The IL-17 level was lower in the mesenteric lymph nodes of the MSC-treated group(P=0.0126).In homogenized colons,the IL-17 and tumor necrosis factor-α(P=0.0092)expression levels were also lower in the treated group.CONCLUSION:MSC infusion provided no significanthistopathologic or clinical improvement,thus representing a limited therapeutic approach for IBD.Functional enhancement of MSCs is needed in further study.

Human mesenchymal stem cells derived from umbilical cord and bone marrow exert immunomodulatory effects in different mechanisms

BACKGROUND Mesenchymal stem cells(MSCs)are an attractive tool to treat graft-versus-host disease because of their unique immunoregulatory properties.Although human bone marrow-derived MSCs(BM-MSCs)were the most widely used MSCs in cell therapy until recently,MSCs derived from human umbilical cords(UC-MSCs)have gained popularity as cell therapy material for their ethical and noninvasive collection.AIM To investigate the difference in mechanisms of the immunosuppressive effects of UC-MSCs and BM-MSCs.METHODS To analyze soluble factors expressed by MSCs,such as indolamine 2,3-dioxygenase,cyclooxygenase-2,prostaglandin E2 and interleukin(IL)-6,inflammatory environments in vitro were reconstituted with combinations of interferon-gamma(IFN-γ),tumor necrosis factor alpha and IL-1βor with IFN-γalone.Activated T cells were cocultured with MSCs treated with indomethacin and/or anti-IL-10.To assess the ability of MSCs to inhibit T helper 17 cells and induce regulatory T cells,induced T helper 17 cells were cocultured with MSCs treated with indomethacin or anti-IL-10.Xenogeneic graft-versus-host disease was induced in NOG mice(NOD/Shi-scid/IL-2Rγnull)and UC-MSCs or BM-MSCs were treated as cell therapies.RESULTS Our data demonstrated that BM-MSCs and UC-MSCs shared similar phenotypic characteristics and immunomodulation abilities.BM-MSCs expressed more indolamine 2,3-dioxygenase after cytokine stimulation with different combinations of IFN-γ,tumor necrosis factor alpha-αand IL-1βor IFN-γalone.UC-MSCs expressed more prostaglandin E2,IL-6,programmed death-ligand 1 and 2 in the in vitro inflammatory environment.Cyclooxygenase-2 and IL-10 were key factors in the immunomodulatory mechanisms of both MSCs.In addition,UC-MSCs inhibited more T helper 17 cells and induced more regulatory T cells than BM-MSCs.UC-MSCs and BM-MSCs exhibited similar effects on attenuating graft-versus-host disease.CONCLUSION UC-MSCs and BM-MSCs exert similar immunosuppressive effects with different mechanisms involved.These findings suggest that UC-MSCs have distinct immunoregulatory functions and may substitute BM-MBSCs in the field of cell therapy.

Generation and characterization of novel stromal specific antibodies

Rheumatoid synovial fibroblasts were used as an immunogen to produce monoclonal antibodies selected for their reactivity with stromal cell antigens.Mice were immunised with low passage whole cell preparations and the subsequent hybridomas were screened by immunohistochemistry on rheumatoid synovium and tonsil sections.The aim was to identify those antibodies that recognised antigens that were restricted to stromal cells and were not expressed on CD45 positive leucocytes.A significant number of antibodies detected antigen that identified endothelial cells.These antibodies were further characterised to determine whether the vessels identified by these antibodies were vascular or lymphatic.From five fusions clones were identified with predominant reactivity with:1)fibroblasts and endothelial cells;or 2)broad stromal elements(fibroblast,endothelium,epithelium,follicular dendritic cells).A fibroblast-specific antibody that did not also identify vessels was not generated.Examples of each reactivity pattern are discussed.

Vaccine therapy for dysbiosis-related diseases

Progress in genomic analysis has resulted in the proposal that the intestinal microbiota is a crucial environmental factor in the development of multifactorial diseases,such as obesity,diabetes,rheumatoid arthritis,and inflammatory bowel diseases represented by Crohn’s disease and ulcerative colitis.Dysregulated gut microbiome contributes to the pathogenesis of such disorders;however,there are few effective treatments for controlling only disease-mediating bacteria.Here,we review current knowledge about the intestinal microbiome in health and disease,and discuss a regulatory strategy using a parenteral vaccine with emulsified curdlan and CpG oligodeoxynucleotides,which we have recently developed.Unlike other conventional injectable immunizations,our vaccine contributes to the induction of antigen-specific systemic and mucosal immunity.This vaccine strategy can prevent infectious diseases such as Streptococcus pneumoniae infection,and control metabolic symptoms mediated by intestinal bacteria(e.g.Clostridium ramosum)by induction of high titers of antigen-specific IgA at target mucosal sites.In the future,our vaccination approach could be an effective therapy for common infectious diseases and dysbiosis-related disorders that have been difficult to control so far.

Comparative studies of serum-free media and detection techniques for <i>in vitro</i>drug sensitivity assessment of <i>Plasmodium falciparum</i>

Malaria continues to be a devastating disease. In a previous study, we formulated a chemically defined culture medium that is able to sustain the complete intraerythrocytic growth of Plasmodium falciparum. We tested the feasibility of using the medium (CDRPMI) as well as human serum-free media enriched with commercially available human-serum substitutes (GFSRPMI and ALBRPMI) to assess the drug sensitivity of P. falciparum, using chloroquine diphosphate (CQ) and dihydroartemisinin (DHART) as conventional antimalarial drugs. Growth inhibition was measured by four different methods: flow cytometry with SYBR Green I (FCM), microscopy (Giemsa method), enzymatic estimation of parasite lactate dehydrogenase (pLDH), and histidine-rich protein 2 (HRPII) determination. In drug sensitivity tests on asynchronous parasites cultured for 96 h in the presence of drugs, the dose-response curves were similar and differences in the 50% growth inhibition concentrations for the drugs, which were estimated by the four methods, were not statistically significant for the three culture media. The effect of the drugs on the growth of synchronous parasites at the ring stage was also assessed in micro-volume tests by three different methods of FCM: tracking fluorescent erythrocytes, schizont test, and merozoite test. Dose-response curves for the drugs were similar, and differences in the 50% growth inhibition concentrations were not statistically significant for CDRPMI and GFSRPMI. Thus CDRPMI as well as GFSRPMI and ALBRPMI can be similarly useful media for drug sensitivity testing of P. falciparum. The FCM, pLDH and HRPII estimations were fast and reliable detection methods, with FCM allowing schizont and merozoite tests to be performed with shorter periods of culture.

IMPAIRED AUTOLOGOUS MIXED LYMPHOCYTE REACTION CORRELATED WITH DECREASED EXPRESSION OF HLA-Ⅱ ANTIGENS ON MONOCYTES IN PATIENTS WITH MYELOID LEUKEMIA

The proliferative response of T-cells to autolo-gous non-T-cells is referred to as the autologous mixed lymphocyte reaction (AMLR). Recent studies have suggested that AMLR represents a mechanism of immune regulation in vivo. We investigated AMLR in patients with acute- and chronic myeloid leukemia (AML and CML). AMLR was found to be significantly depressed (P<0.001) in AML patients (n=17, cpm=532±95) and CML patients (n=13, cpm=688±99) when compared with that of their healthy HLA-identical siblings serving as controls (n=17, cpm=4152±619 and n=13 cpm=4086±421, respectively). In order to understand the cellular basis of the defective AMLR in patients with AML end CML, we performed mitogen-treated T-cell cultures analysis of T-cell subsets and HLA-Ⅱ antigen detection on monocytes. The results indicated that the defect of AMLR in patients resided at the stimulator monocyte level rather than at the responder T-cell level. Enumeration of monocytes reactive with monoclonal antibody Tu22, which recognizes determinants of HLA-DQ, demonstrated that ML patients had a significantly decreased (P<0.091) number of circulating Tu22+ monocytes when compared with normal controls. These studies suggest that a deficiency of HLA-DQ+ monocytes contributes to the depression of AMLR in ML and possibly underlies the abnormalities of immune response present in this disease.

A critical regulation of Th2 cell responses by RORa in allergic asthma

Allergic asthma is a chronic inflammatory disease of the lung and the airway,which is characterized by aberrant type 2 immune responses to otherwise unharmful aeroallergens.While the central role of Th2 cells and type 2 cytokines in the pathogenesis of allergic asthma is well documented,the regulation and plasticity of Th2 cells remain incompletely understood.By using an animal model of allergic asthma in IL-4-reporter mice,we found that Th2 cells in the lung expressed higher levels of Rora than those in the lymph nodes,and that treatment with an RORa agonist SRI 078 resulted in diminished Th2 cell responses in vivo.To determine the T cell-intrinsic role of RORa in allergic asthma in vivo,we established T cell-specific RORa-deficient^(f/f)fyf(Cd4creRora^(f/f))mice.Upon intranasal allergen challenges,Cd4creRora^(f/f)mice exhibited a significantly increased Th2 cells in the lungs and the airway and showed an enhanced eosinophilic inflammation compared to littermate control mice.Studies with Foxp3^(YFP-cre)Rora^(f/f)mice and CD^(8+)T cell depletion showed that the increased Th2 cell responses in the Cd4creRora^(f/f)mice were independent of Treg cells and CD^(8+)T cells.Our findings demonstrate a critical regulatory role of RORa in Th2 cells,which suggest that RORa agonists could be effective for the treatment of allergic diseases.

MCMV-based vaccine vectors expressing full-length viral proteins provide long-term humoral immune protection upon a single-shot vaccination

Global pandemics caused by influenza or coronaviruses cause severe disruptions to public health and lead to high morbidity and mortality.There remains a medical need for vaccines against these pathogens.CMV(cytomegalovirus)is aβ-herpesvirus that induces uniquely robust immune responses in which remarkably large populations of antigen-specific CD8+T cells are maintained for a lifetime.Hence,CMV has been proposed and investigated as a novel vaccine vector for expressing antigenic peptides or proteins to elicit protective cellular immune responses against numerous pathogens.We generated two recombinant murine CMV(MCMV)vaccine vectors expressing hemagglutinin(HA)of influenza A virus(MCMV^(HA))or the spike protein of severe acute respiratory syndrome coronavirus 2(MCMV^(S)).A single injection of MCMVs expressing either viral protein induced potent neutralizing antibody responses,which strengthened over time.Importantly,MCMV^(HA)-vaccinated mice were protected from illness following challenge with the influenza virus,and we excluded that this protection was due to the effects of memory T cells.Conclusively,we show here that MCMV vectors induce not only long-term cellular immunity but also humoral responses that provide long-term immune protection against clinically relevant respiratory pathogens.

Single cell whole-genome sequencing of brain cells:age-and cell-type specific mutational profiles

In a recent study published in Cell,Ganz et al.shed new light on the mutational landscape of brain cells,particularly neurons and oligodendrocytes(OLs).1 Utilizing a combination of optimized single-cell whole-genome sequencing with single-nucleus chromatin accessibility and gene expression analysis,they profiled somatic mutations in 86 OLs from 13 neurotypical individuals,spanning in age from infants to elderly.Neurons investigated were 56,derived from 19(including 12 overlapping)individuals(Fig.1).

Heme oxygenase 1-mediated ferroptosis in Kupffer cells initiates liver injury during heat stroke

With the escalating prevalence of global heat waves,heat stroke has become a prominent health concern,leading to substantial liver damage.Unlike other forms of liver injury,heat strokeinduced damage is characterized by heat cytotoxicity and heightened inflammation,directly contributing to elevated mortality rates.While clinical assessments have identified elevated bilirubin levels as indicative of Kupffer cell dysfunction,their specific correlation with heat stroke liver injury remains unclear.Our hypothesis proposes the involvement of Kupffer cell ferroptosis during heat stroke,initiating IL-1bmediated inflammation.Using single-cell RNA sequencing of murine macrophages,a distinct and highly susceptible Kupffer cell subtype,Clec4Ft/CD206t,emerged,with heme oxygenase 1(HMOX-1)playing a pivotal role.Mechanistically,heat-induced HMOX-1,regulated by early growth response factor 1,mediated ferroptosis in Kupffer cells,specifically in the Clec4F t/CD206 t subtype(KC2),activating phosphatidylinositol 4-kinase beta and promoting PI4P production.This cascade triggered NLRP3 inflammasome activation and maturation of IL-1b.These findings underscore the critical role of targeted therapy against HMOX-1 in ferroptosis within Kupffer cells,particularly in Clec4F t/CD206 t KCs.Such an approach has the potential to mitigate inflammation and alleviate acute liver injury in the context of heat stroke,offering a promising avenue for future therapeutic interventions.

Antigen Processing by Autoreactive B Cells Promotes Determinant Spreading

Acute primary immune responses tend to focus on few immunodominant determinants using a very limited number of T cell clones for expansion, whereas chronic inflammatory responses generally recruit a large number of different T cell clones to attack a broader range of determinants of the invading pathogens or the inflamed tissues. In T cell-mediated organ-specific autoimmune disease, a transition from the acute to the chronic phase contributes to pathogenesis, and the broadening process is called determinant spreading. The cellular components catalyzing the spreading reaction are not identified. It has been suggested that autoreactive B cells may play a central role in diversifying autoreactive T cell responses, possibly through affecting antigen processing and presentation. The clonal identity and diversity of the B cells and antibodies seem critical in regulating T cell activity and subsequent tissue damage or repair. Here, we use two autoimmune animal models, experimental autoimmune thyroiditis （EAT） and type 1 diabetes （T1D）, to discuss how autoreactive B cells or antibodies alter the processing and presentation of autoantigens to regulate specific T cell response. Cellular ＆ Molecular Immunology. 2005;2（3）：169-175.

The microbiota is dispensable for the early stages of peripheral regulatory T cell induction within mesenteric lymph nodes

Intestinal Foxp3+regulatory T cell(Treg)subsets are crucial players in tolerance to microbiota-derived and food-borne antigens,and compelling evidence suggests that the intestinal microbiota modulates their generation,functional specialization,and maintenance.Selected bacterial species and microbiota-derived metabolites,such as short-chain fatty acids(SCFAs),have been reported to promote Treg homeostasis in the intestinal lamina propria.Furthermore,gut-draining mesenteric lymph nodes(mLNs)are particularly efficient sites for the generation of peripherally induced Tregs(pTregs).Despite this knowledge,the direct role of the microbiota and their metabolites in the early stages of pTreg induction within mLNs is not fully elucidated.Here,using an adoptive transfer-based pTreg induction system,we demonstrate that neither transfer of a dysbiotic microbiota nor dietary SCFA supplementation modulated the pTreg induction capacity of mLNs.Even mice housed under germ-free(GF)conditions displayed equivalent pTreg induction within mLNs.Further molecular characterization of these de novo induced pTregs from mLNs by dissection of their transcriptomes and accessible chromatin regions revealed that the microbiota indeed has a limited impact and does not contribute to the initialization of the Treg-specific epigenetic landscape.Overall,our data suggest that the microbiota is dispensable for the early stages of pTreg induction within mLNs.

IL-33: a jack of all trades in the orchestration of respiratory antibacterial immunity

Influenza A virus(IAV)is the causative agent of mostly mild to moderate seasonal respiratory infections and several pandemic outbreaks,the most recent of which was reported in 2009.Previous IAV pandemics were associated with an enormous death toll;for example,the 1918 H1N1 pandemic affected hundreds of millions of people globally and resulted in~50 million deaths.1 Microbiologic analyses of patient samples revealed a strong incidence of bacterial pathogens in fatal complications of viral infection.2 To date,many pieces of epidemiologic and experimental evidence reveal pronounced susceptibility to detrimental bacterial superinfection in IAVinfected individuals.

PHF6 functions as a tumor suppressor by recruiting methyltransferase SUV39H1 to nucleolar region and offers a novel therapeutic target for PHF6-muntant leukemia

Mutations in the plant homeodomain-like finger protein 6(PHF6)gene are strongly associated with acute myeloid(AML)and T-cell acute lymphoblastic leukemia(T-ALL).In this study,we demonstrated that PHF6 can bind to H3K9me3 and H3K27me1 on the nucleolar chromatin and recruit histone methyltransferase SUV39H1 to the rDNA locus.The deletion of PHF6 caused a decrease in the recruitment of SUV39H1 to rDNA gene loci,resulting in a reduction in the level of H3K9me3 and the promotion of rDNA transcription.The knockdown of either SUV39H1 or PHF6 significantly attenuated the effects of increase in H3K9me3 and suppressed the transcription of rDNA induced by the overexpression of the other interacting partner,thereby establishing an interdependent relationship between PHF6 and SUV39H1 in their control of rRNA transcription.The PHF6 clinical mutants significantly impaired the ability to bind and recruit SUV39H1 to the rDNA loci,resulting in an increase in rDNA transcription activity,the proliferation of in vitro leukemia cells,and the growth of in vivo mouse xenografts.Importantly,significantly elevated levels of pre-rRNA were observed in clinical AML patients who possessed a mutated version of PHF6.The specific rDNA transcription inhibitor CX5461 significantly reduced the resistance of U937 AML cells deficient in PHF6 to cytarabine,the drug that is most commonly used to treat AML.Collectively,we revealed a novel molecular mechanism by which PHF6 recruits methyltransferase SUV39H1 to the nucleolar region in leukemia and provided a potential therapeutic target for PHF6-mutant leukemia.

Negative elongation factor:a key factor in the maintenance of intestinal epithelial barrier integrity

In addition to its physiological function in the uptake of dietary nutrients,the intestinal epithelium constitutes an essential mechanical barrier separating luminal gut content and mucosal microbiota from the inner body.Intestinal epithelial cells(IEC)comprise the frontline of this barrier,the maintenance of which critically depends on the expression of cell-cell junction protein structures,including tight junctions,adherence junctions,desmosomes,and gap junctions,which physically bridge and seal the intercellular niche within the IEC layer.

Macrophage’s little helper: vitamin A directs alternatively activated monocyte-derived macrophages to tissue-resident macrophages

Tissue-resident macrophages originate from the yolk sac in a Myb-independent manner and populate all organs during embryogenesis.These macrophages are a heterogeneous self-renewing population that adapt to the organ-specific local environment to contribute to tissue homeostasis.Depending on the age,organ and inflammatory conditions,macrophages that are derived from hematopoietic stem cells(HSCs)in a Myb-dependent manner may also infiltrate organs and develop into tissueresident macrophages.

Tumor-specific memory CD8^(+)T cells are strictly resident in draining lymph nodes during tumorigenesis

The functional exhaustion of CD8^(+)T cells represents a fundamental hallmark of chronic viral infection and cancer and,in both scenarios,is driven by prolonged exposure to persistent cognate antigens in the context of an immunoinhibitory microenvironment.Exhausted CD8^(+)T cells upregulate the expression of a wide diversity of coinhibitory immunoreceptors(also referred to as immune checkpoint receptors),such as PD-1,Tim-3,LAG-3,and TIGIT.Concomitantly,exhausted CD8^(+)T cells lose their potential to differentiate into functional memory cells and are characterized by hierarchical loss of effector function,leading to compromised tumor control and viral eradication[1,2].

The emerging role of Arid5a in cancer: A new target for tumors

AT-rich interactive domain 5a (Arid5a) is a member of the arid family of proteins, which contain a helix-turn-helix domain and an ability to bind to nucleic acids. Current evidence suggests that Arid5a performs dual functions as a transcription factor and an RNA-binding protein in immune, nonimmune, and/ or tumor cells depending on its cellular localization. The contribution of Arid5a to the development of inflammation, autoimmunity, and obesity through its transcriptional and posttranscriptional regulatory functions has broadly been reviewed. Recent studies have indeed revealed an association of Arid5a with cancers, including breast, pancreatic, colorectal, and lung cancers and glioma. Notably, Arid5a affects various aspects of cellular homeostasis, including invasion, metastasis, epithelial-to-mesenchymal transition, immune evasion, adipogenesis and M1-like tumor-associated macrophage (TAM)-to-M2-like TAM transition. This review aims to summarize current knowledge of Arid5a from a cancer perspective and highlights recent advances in Arid5a-related cancer research. This review may improve the understanding of Arid5a-mediated molecular mechanisms and their relevance to cancers.

Beware the intruder:gasodermin A as molecular guardian preventing systemic dissemination of group A streptococci following local skin infection

Pyroptosis represents a host-protective mechanism that promotes clearance of pathogens by initiating the recruitment of immune cells to the site of infection.In a study recently published in Nature,Deng et al.extensively outlined a hitherto unknown role of the group A Streptococcus(GAS)-derived cysteine protease streptococcal pyrogenic exotoxin B(SpeB)in gasodermin A(GSDMA)cleavage as the primary step in induction of pyroptosis.As clearly shown in GSDMA-deficient mice infected with SpeBproducing GAS,this cleavage event is key to preventing the systemic spread and fatal course of GAS following local skin infection[1].

Route, origin & valence matter: towards sophisticated nextgeneration vaccines to cope with the COVID-19 pandemic

In a recent study published in Cell,Afkhami and colleagues systematically compared different routes of vaccine delivery,origin of the vaccine platform as well as valence of the vaccine and demonstrated that the respiratory mucosal delivery of a trivalent chimpanzee’s adenovirus(Ad)-vectored vaccine is superior to any other of the tested conditions in inducing broadlyacting immunity and protection against current severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)and possibly future variants of concern(VOC).1 The ongoing global coronavirus disease 2019(COVID-19)pandemic caused by SARS-CoV-2 has forced an uniquely fast development of novel mRNA-and vector-based vaccines.