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What is the distinction between chemokines, cytokines, interferons and interleukins?

What is the distinction between chemokines, cytokines, interferons and interleukins?


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They all seem to describe molecules of similar function and many people seem to use them interchangeably.

Also please include any other similar molecules if I've forgotten any in the list above.


Cytokines is the general class of molecules to which chemokines, interferons, interleukins and others belong. Biologists dispute whether something is a hormone or a cytokine, but generally the consensus goes with if it's to do with immunology it's a cytokine or if the resting concentration is in the picomolar range, but that's a very rough distinction.

Chemokines are molecules that drive cellular chemotaxis. That means they make cells move towards a desired place. Generally chemokines refer to immune cells and there's loads and loads of them.

Interleukins are anything which are messenger molecules between immune cells (inter- means between and -leukins means leukocytes/white blood cells). They're typically denoted by IL + number. However the interferon and tumour necrosis families come under interleukins too in most people's opinion. The interferons are a special group that typically inhibit viruses by making cells non-permissible to viral replication. They also do a few other things like activate macrophages or promote Th1 response, which both also interfere with viruses but there is bacterial overlap. The TNF family is a bit weirder as some are interleukin-like but others aren't as much. TNF-alpha (the classic one) is involved in macrophage maturation. However not all interleukins are strictly between white blood cells, as IL1 acts on the hypothalamus among other leukocytes.

See the problem is everything is a bit grey because most cytokines (if not all) have more than one role and there's never consensus in the world of immunology when it comes to cytokines because more and more keeps getting discovered.


In short, this is my understanding following my immunology course (which is mostly identical to AndroidPenguin's answer):

  • Chemokines: Produce cell movement, i.e. act as chemoattractants. Typically these are used to recruit more immune cells to a site of infection. (Name origin says it all: "chemotactic cytokine = chemokine… )

  • Cytokines: Produce an inflammatory response by altering transcription (via surface receptor signal cascades). These often act on non-immune cells.

    • Interferons: A type of cytokine. Secreted by virtually every cell in response to being infected by a virus. Produce an 'anti-viral' state in other cells, through MANY mechanisms. Causes the typical flu-like symptoms.

    • Tumor necrosis factors (specifically TNF alpha): A type of cytokine. Prepares endothelium (blood vessel walls) to support an inflammatory response by vasodilation, increased adhesion and increased permeability.

  • Interleukins: Some are chemokines (e.g. secreted by macrophages and dendritic cells upon activation to recruit further phagocytes and adaptive immune cells), others are cytokines (e.g. activating B cells to differentiate into plasma and memory cells after T cell contact).


Pharmacology cross reference: Recombinant type I interferons are injected as therapeutics. Viral infections would seem like logical indications, but interferons are both expensive and have considerable adverse effects, e. g., flu-like symptoms on injection, anemia and depression. Their application is therefore limited to life-threatening viral diseases, e. g. hepatitis C.


11.3C: Cytokines Important in Innate Immunity

  • Contributed by Gary Kaiser
  • Professor (Microbiology) at Community College of Baltimore Country (Cantonsville)

Cytokines are low molecular weight, soluble proteins that are produced in response to an antigen and function as chemical messengers for regulating the innate and adaptive immune systems. They are produced by virtually all cells involved in innate and adaptive immunity, but especially by T- helper (Th) lymphocytes. The activation of cytokine-producing cells triggers them to synthesize and secrete their cytokines. The cytokines, in turn, are then able to bind to specific cytokine receptors on other cells of the immune system and influence their activity in some manner.

Cytokines are pleiotropic, redundant, and multifunctional.

  • Pleiotropic means that a particular cytokine can act on a number of different types of cells rather than a single cell type.
  • Redundant refers to to the ability of a number of different cytokines to carry out the same function.
  • Multifunctional means the same cytokine is able to regulate a number of different functions.

Some cytokines are antagonistic in that one cytokine stimulates a particular defense function while another cytokine inhibits that function. Other cytokines are synergistic wherein two different cytokines have a greater effect in combination than either of the two would by themselves. There are three functional categories of cytokines:

1. cytokines that regulate innate immune responses,
2. cytokines that regulate adaptive Immune responses, and
3. cytokines that stimulate hematopoiesis.

Cytokines that regulate innate immunity are produced primarily by mononuclear phagocytes such as macrophages and dendritic cells, although they can also be produced by T-lymphocytes, NK cells, endothelial cells, and mucosal epithelial cells. They are produced primarily in response to pathogen-associated molecular patterns (PAMPs) such as LPS, peptidoglycan monomers, teichoic acids, unmethylated cytosine-guanine dinucleotide or CpG sequences in bacterial and viral genomes, and double-stranded viral RNA. Cytokines produced in response to PRRs on cell surfaces, such as the inflammatory cytokines IL-1, IL-6, IL-8, and TNF-alpha, mainly act on leukocytes and the endothelial cells that form blood vessels in order to promote and control early inflammatory re sponses (Figure (PageIndex<1>)). C ytokines produced in response to PRRs that recognize viral nucleic acids, such as type I interferons, primarily block viral replication within infected host cells (see Figure (PageIndex<2>)A and Figure (PageIndex<2>)B).

Figure (PageIndex<1>): Integrins on the surface of the leukocyte bind to adhesion molecules on the inner surface of the vascular endothelial cells. The leukocytes flatten out and squeeze between the endothelial cells to leave the blood vessels and enter the tissue. The increased capillary permeability also allows plasma to enter the tissue.


A genomic analysis of chicken cytokines and chemokines

As most mechanisms of adaptive immunity evolved during the divergence of vertebrates, the immune systems of extant vertebrates represent different successful variations on the themes initiated in their earliest common ancestors. The genes involved in elaborating these mechanisms have been subject to exceptional selective pressures in an arms race with highly adaptable pathogens, resulting in highly divergent sequences of orthologous genes and the gain and loss of members of gene families as different species find different solutions to the challenge of infection. Consequently, it has been difficult to transfer to the chicken detailed knowledge of the molecular mechanisms of the mammalian immune system and, thus, to enhance the already significant contribution of chickens toward understanding the evolution of immunity. The availability of the chicken genome sequence provides the opportunity to resolve outstanding questions concerning which molecular components of the immune system are shared between mammals and birds and which represent their unique evolutionary solutions. We have integrated genome data with existing knowledge to make a new comparative census of members of cytokine and chemokine gene families, distinguishing the core set of molecules likely to be common to all higher vertebrates from those particular to these 300 million-year-old lineages. Some differences can be explained by the different architectures of the mammalian and avian immune systems. Chickens lack lymph nodes and also the genes for the lymphotoxins and lymphotoxin receptors. The lack of functional eosinophils correlates with the absence of the eotaxin genes and our previously reported observation that interleukin- 5 (IL-5) is a pseudogene. To summarize, in the chicken genome, we can identify the genes for 23 ILs, 8 type I interferons (IFNs), IFN-gamma, 1 colony-stimulating factor (GM-CSF), 2 of the 3 known transforming growth factors (TGFs), 24 chemokines (1 XCL, 14 CCL, 8 CXCL, and 1 CX3CL), and 10 tumor necrosis factor superfamily (TNFSF) members. Receptor genes present in the genome suggest the likely presence of 2 other ILs, 1 other CSF, and 2 other TNFSF members.


Overview

Cytokines are important mediators of immune responses and produced by almost every cell in the body. Growth stimulatory or inhibitory cytokines could be subclassified as interleukins (ILs), lymphokines, monokines, chemokines, and hematopoietic growth factors. In cancer, certain cytokines act directly on the growth, differentiation, or survival of endothelial cells, whereas others act by attracting inflammatory cell types affecting angiogenesis or by inducing secondary cytokines or other mediators regulating angiogenesis. Proinflammatory and chemotactic cytokines influence the tumor environment and control the quantity and nature of infiltrating hematopoietic effector cells, with inhibiting or enhancing effects on tumor growth. The important role of cytokines in regulating immune responses may permit an effective immune response against the tumors or suppress the function of antigen-presenting cells (APC).

The understanding of cytokines has now emerged as complex picture of interacting stimulatory and inhibitory factors. Many of the molecules that govern this process have been cloned and have entered clinical trials. It is now clear that regulatory cytokines are characteristically pleiotropic and, at the same time, exhibit significant functional redundancy.

The biologic characterization of the known clinically relevant ILs, interferons and selected growth factors, the rationale for their use in therapy for patients with cancer, and the accumulated clinical experience represent the subjects of this chapter.

Cytokines, a diverse family of signaling molecules, are important mediators of immune responses and produced by almost every cell in the body, including various cancer cells. In general, some cytokines are growth stimulatory and others are inhibitory. Cytokines with clinical relevance to cancer include those subclassified further as interleukins (ILs), monokines, chemokines, and hematopoietic growth factors. IL designates any soluble protein or glycoprotein product of leukocytes that regulates the responses of other leukocytes. ILs produce their effects primarily through paracrine interactions. In cancer, certain cytokines act directly on the growth, differentiation, or survival of endothelial cells, whereas others act by attracting inflammatory cell types affecting angiogenesis or by inducing secondary cytokines or other mediators regulating angiogenesis. Proinflammatory and chemotactic cytokines influence the tumor environment and control the quantity and nature of infiltrating hematopoietic effector cells, with inhibiting or enhancing effects on tumor growth. The important role of cytokines in regulating immune responses may permit an effective immune response against the tumors or suppress the function of APC. Presuming antigens exist on tumor cells, various immunostimulatory cytokines, and particularly ILs, are now administered to patients in an attempt to initiate, augment, or otherwise stimulate a weak or previously nonexistent antitumor immune response. In addition to immune response stimulation, some ILs have been used to stimulate the growth and differentiation of various subpopulations of blood cells after chemotherapy or bone marrow transplantation (BMT) in a restorative role.

It is now clear that the pleiotropic nature of many cytokines allows them to influence virtually all organ systems (Figure 1). Cytokines may have their own private receptor but may also share a “public” receptor with other cytokines (Tables 1 and 2).


Figure 1 In addition to their effects on hematopoiesis and immunocompetence, “hematopoietic” growth factors influence multiple organ systems, including (but not limited to) bone remodeling, cardiorespiratory function, hepatic function, and the gastrointestinal tract.

Table 1 Types of hematopoietic growth factor receptors



























Type Characteristics Receptor examples
Type I cytokine receptor Does not possess intrinsic kinase activity. Receptor acts as docking site for adaptor molecules, which leads to phosphorylation of cellular substrates IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-9, IL-13, IL-18, IL-21, GM-CSF, G-CSF, EPO, TPO, and leukemia inhibitory factor
Type II cytokine receptor Contains extracellular fibronectin III type domain Interferon and IL-10
Receptors with tyrosine kinase domains (type III) Large extracellular immunoglobulin-like domain, single transmembrane spinning region, and a cytoplasmic tyrosine kinase domain(s) fms (M-CSF receptor), FLT-3, c-kit (SCF receptor), and PDGFR
Chemokine receptor Seven transmembrane spanning G protein-linked regions IL-8
Tumor necrosis factor family Cysteine-rich repeats in the extracellular domain, and cytoplasmic 80 amino acid “death domain” Tumor necrosis factor and Fas

Abbreviations: EPO, erythropoietin G-CSF, granulocyte colony-stimulating factor GM-CSF, granulocyte macrophage colony-stimulating factor IL, interleukin M-CSF, macrophage colony-stimulating factor SCF, stem cell factor TPO, thrombopoietin.


































































































































































































































































































































































































































Chromosomal location Receptors Selected biologic activities
IL-1 2q13 IL-1RI and IL-1RII Promotes acute-phase response. IL-1 acts on nearly every organ system. Induces production of multiple cytokines
Upregulates cell-surface cytokine expression
Synergizes with other cytokines to stimulate hematopoietic progenitor proliferation
Influences immune regulation
Modulates endocrine function
Affects bone formation
IL-1R acts as a cofactor in neural transmission
IL-2 4q26-q27 αβγ heterotrimeric complex Induces proliferation and activation of T cells, B cells, and NK cells
IL-3 5q31 IL-3 receptor (heterodimer of IL-3-specific α subunit and β subunit) Stimulation of multilineage hematopoietic progenitors, especially when used in combination with other cytokines (SCF, IL-1, IL-6, G-CSF, GM-CSF, EPO, and TPO)
IL-4 and IL-13 5q31 Type I IL-4 receptor (IL-4Rα and IL-2 receptor γ c chain subunits) transduces IL-4 IL-4 and IL-13 are involved in allergic reaction (induce switch to IgE)
Type II IL-4 receptor (IL-4Rα and the IL-13 Rα1 subunits) transduces IL-4 and IL-13
IL-4Rα and IL-13 Rα2 complex or two IL-13 Rα transduce IL-13
IL-5 5q31 Consists of IL-5Rα (IL-5-specific) and a β subunit Regulates production, function, survival, and migration of eosinophils
β subunit is common to IL-3 and GM-CSF complexes Enhances basophil number and function
IL-6 7p21 IL-6Rα together with gp130 B- and T-cell development and function
Thrombopoiesis
Acute-phase protein synthesis
Inhibition of hepatic albumin excretion
Osteoclastic bone resorption
Neural differentiation
IL-7 8q12-q13 Composed of IL-7Rα (CD127) and the common γc chain subunits Critical for T- and B-cell development
IL-8 4q12-q13 IL-8Rα and IL-8Rβ exist Potent chemoattractant agent for a variety of leukocytes, especially neutrophils
Suppresses colony formation of immature myeloid progenitors
Increases keratinocyte and endothelial cell proliferation
IL-9 5q31.1 IL-9 receptor Supports clonogenic maturation of erythroid progenitors
Acts as a mast cell differentiation factor
Protects lymphomas from apoptosis
Cooperates with IL-4 in B-cell responses
Enhances neuronal differentiation
IL-10 1q31-q32 IL-10 receptor interferon receptors Inhibits cytokine synthesis by Th1 cells and monocytes/macrophages
Stimulates B-cell proliferation
Involved in transformation of B cells by Epstein–Barr virus and tumor necrosis factor (TNF) receptors
IL-11 19q13.3-q13.4 IL-11Rα and gp 130 subunits gp 130 = CD130 on 5q11 IL-6, oncostatin M, and leukemia inhibitory factor also use gp130 subunit Best known as a thrombopoietic factor
Stimulates multilineage progenitors, erythropoiesis, myelopoiesis, and lymphopoiesis
Decreases mucositis in animal models
Stimulates osteoclast development
Inhibits adipogenesis
Stimulates proliferation of neuronal cells
IL-12 IL-12A:3p12-q13.2 IL-12Rβ1 and IL-12Rβ2 chains are related to gp 130 Proinflammatory cytokine important in resistance to infections
IL-12B:5q31.1-q33.1 Th1 development
Stimulatory and inhibitory effects on hematopoiesis
IL-15 4q31 High-affinity receptor requires IL-2Rβ and γ chains and IL-15 Rα chain Triggers proliferation and immunoglobulin production in preactivated B cells
Number of CD8 + memory T cells may be controlled by balance of IL-15 (stimulatory) and IL-12 (inhibitory)
Stimulates proliferation of NK cells and activated CD4 + or CD8 + T cells
Facilitates the induction of LAK cells and CTLs
Stimulates mast cell proliferation
Promotes proliferation of hairy-cell leukemia and chronic lymphocytic leukemia cells
IL-16 15q26.1 Requires CD4 for biologic activities Tetraspanin CD9 Chemoattractant for CD4 + cells (T cells, monocytes, and eosinophils)
May be involved in asthma and in granulomatous inflammation
Has antiviral effects on HIV-1
IL-17 2q31 IL-17 receptor May mediate, in part, T-cell contribution to inflammation
Stimulates epithelial, endothelial, fibroblastic, and macrophage cells to express a variety of inflammatory cytokines
Promotes the capacity of fibroblasts to sustain hematopoietic progenitor growth
Promotes differentiation of dendritic cell progenitors
May be involved in the pathogenesis of rheumatoid arthritis and graft rejection
IL-18 11q22.2-q22.3 IL-18 receptor Promotes production of IFN-γ and TNF
Targets are T cells, NK cells, and macrophages
Promotes Th1 responses to virus
IL-19 1q32 IL-20Rl and IL-20R2 Induces IL-6 and TNF-α
IL-20 1q32 IL-20R1 and IL-20R2 Induction of genes involved in inflammation such as TNF-α, MRP14, and MCP-1
IL-21 4q26–27 IL-21 receptor Mainly regulates T-cell proliferation and differentiation
Regulates cell-mediated immunity and the clearance of tumors
IL-22 12q14 IL-22R1 and IL-10R2 Upregulates the production of acute-phase reactants
Induces the production of ROS in resting B cells
IL-23 12q13 IL-12Rb1 and IL-23R A unique function of IL-23 is the preferential induction of proliferation of the memory subset of T cells
IL-24 1q32 IL-20R1 and IL-20R2 Induces IL-6, TNF-a, IL-1b, IL-12, and GM-CSF
IL-22R1 and IL-20R2 Functionally it has opposite effects with IL-10
Infection with Ad-IL24 results in downregulation of Bcl-2 and Bcl-XL (antiapoptotic proteins) and upregulation of Bax and Bak (proapoptotic proteins) in cancer cells
IL-25 14q11 IL-17BR IL-25 induces IL-4, IL-5, and IL-13 gene expression and protein production
IL-26 12q14 IL-20R1 and IL-10R2 Immune-protective role against viral infection
IL-27 12q13 TCCR/WSX-1 and GP130 Early Th1 initiation
Synergizes with IL-12 in inducing IFN-γ production by T cells and NK cells
IL-28A, 28B, and 29 19q13 IL-28R1 and IL-10R2 Antiviral activities
IL-31 12q24 IL-31 receptor A and oncostatin M receptor Responsible for promoting the dermatitis and epithelial responses that characterize allergic and nonallergic diseases
IL-32 16p13.3 Proteinase 3 Induces other proinflammatory cytokines and chemokines such as TNF-α, IL-1β, IL-6, and IL-8
Induces IκB degradation
Phosphorylates p38 MAPK signaling pathway
IL-33 9p24.1 ST2 Activates NF-κB and MAP kinases
Drives production of Th2-associated cytokines from in vitro polarized Th2 cells
Induces the expression of IL-4, IL-5, and IL-13
Leads to severe pathologic changes in mucosal organs
IL-35 19p13.3 IL-12Rβ2 and gp130 Contributes Treg suppressor activity
Induces IL-10 and IFN-g serum levels
Reduces induction of IL-17
IL-36 IL36A2q12-q14.1 IL-1Rrp2 and IL-1RAcP Activates NF-κB and MAP kinases
IL36B2q14 Plays important role in skin biology
IL36G:2q12-q21 Involved in the initiation and regulation of immune responses
IL36RN:2q14
IL-37 2q12-q14.1 IL-18R Regulates inflammatory responses
IL-38 2q13 IL36R Reduces IL-36g-induced IL-8 production

The biologic characterization of selected ILs (those for which we discuss a role in cancer), interferons (IFNs) and selected growth factors, the rationale for their use in therapy for patients with cancer, and the accumulated clinical experience represent the subjects of this chapter.

Interleukins

Interleukin-1

IL-1 (IL-1α and IL-1β) is the prototypic pleiotropic cytokine and influences nearly every cell type. 1, 2 Because IL-1 is a highly inflammatory cytokine, the margin between salutary effects and serious toxicity in humans is exceedingly narrow. Compounds that attenuate the production and/or activity of IL-1 are therefore being explored in clinical trials.

Biologic effects of IL-1

IL-1 can increase the expression of itself as well as many other cytokines (including IL-1RA), cytokine receptors (including IL-2, IL-3, IL-5, granulocyte macrophage-colony-stimulating factor [GM-CSF], and c-kit), inflammatory mediators (such as cyclooxygenase and inducible nitric oxide synthase), hepatic acute-phase reactants, growth factors, clotting factors, neuropeptides, lipid-related genes, extracellular matrix molecules, and oncogenes (e.g., c-jun, cabl , c-fms , c-myc , and c-fos ). 1 Data suggest that an inflammatory component is present in the microenvironment of most neoplastic tissues, including those not causally related to an obvious inflammatory process. Thus, as a proinflammatory cytokine, IL-1 may also be a major proangiogenic stimulus of both physiological and pathological angiogeneses.

The IL-1 family has been implicated in the function and the dysfunction of virtually every human organ system. Indeed, increased IL-1 production has been reported in patients with infections (viral, bacterial, fungal, and parasitic), intravascular coagulation, cancer (both solid tumors and hematologic malignancies), Alzheimer’s disease, autoimmune disorders, trauma, ischemic diseases, pancreatitis, graft-versus-host disease, transplant rejection, and in healthy subjects after exercise. 1

It has been suggested that the balance between IL-1 and its naturally occurring antagonists is most relevant to illness. 3 This balance may be altered in different ways, depending on the disease. In AML, IL-10 is spontaneously expressed, but IL-1RA gene expression is suppressed even when stimulated with GM-CSF. 4, 5 CML patients with advanced disease and poor survival have suppressed IL-1RA accompanied by high IL-1β. 6 In AML and CML patients, IL-1β acts as an autocrine growth factor exposure to molecules that decrease the activity of IL-1 suppresses leukemic proliferation. 7, 8 Constitutive production of IL-lα, IL-1β, and/or IL-1RA in solid tumors (melanomas, hepatoblastoma, sarcomas, squamous cell carcinomas, transitional cell cancers, and ovarian carcinomas) has been described and may, in some cases, contribute to metastatic potential. However, the relationship between IL-1 and tumor growth is complex.

IL-1 in the clinic

IL-lα and IL-1β have both been administered in clinical cancer trials. 1 In general, the acute toxicities of both isoforms were greater after intravenous than subcutaneous injection. Subcutaneous injection was associated with significant local pain, erythema, and swelling. Dose-related chills and fever were observed in nearly all patients, and even a 1 ng/kg dose was pyrogenic. Nearly all patients receiving intravenous IL-1 at doses of 100 ng/kg or greater were experienced significant hypotension, probably because of induction of nitric oxide.

IL-1 infusion into humans significantly increased circulating IL-6 levels and resulted in a rise in leukocyte counts, even at doses as low as 1 or 2 ng/kg. Increases in platelets, peripheral monocyte count, and phorbol-induced superoxide production were also observed in patients with normal marrow reserves. In contrast to the results in patients with good marrow function, patients with aplastic anemia treated with five daily doses of IL-lα (30–100 ng/kg) had no increases in peripheral blood counts or bone marrow cellularity. 9 However, after chemotherapy, two doses of IL-10 significantly shortened the duration of neutropenia, 10 and IL-lα (5 days) significantly reduced thrombocytopenia. 11 Overall, the benefits of IL-1 therapy were compromised by its toxicity.

Interleukin-2

Originally described as a T-cell growth factor, the function of IL-2 extends beyond lymphocyte activation and population expansion, although T cells still appear to be its major target. 12

Biologic activities of IL-2

IL-2 primarily acts as a T-cell growth factor, but B cells, natural killer (NK) cells, and lymphokine-activated killer (LAK) cells are also responsive to this cytokine. Following binding of IL-2 with the trimeric receptor complex, internalization occurs and cell-cycle progression is induced in association with the expression of a defined series of genes. 13 A second functional response occurs through the IL-2β, dimeric receptor, also known as the intermediate affinity dimeric complex (kDa, 10 −9 ), and involves the differentiation of several subclasses of lymphocytes into LAK cells. 14 This response occurs in patients with cancer who receive IL-2 15, 16 and was originally considered to be a critical part of the anticancer effect of IL-2.

IL-2 in the clinic

IL-2 has had a profound impact on the development of cancer immunotherapy. The administration of IL-2 and the adoptive transfer of antitumor T cells grown in IL-2 represented the first effective immunotherapies for cancer in humans. 17 Since 1992, numerous clinical trials using high-dose IL-2 (HD IL-2) have delivered a remarkably consistent 7% complete response rate in two advanced cancer types, renal cell carcinoma (RCC) and malignant melanoma. 18–22 Many of these complete responses have been durable beyond 10 years. HD IL-2 likely enhances the immune response against cancer cells. Its anticancer activity is strongly related to its ability to act as a growth factor for T lymphocytes, its capacity to stimulate antigen-independent NK cells and LAK cells, and its ability to increase lymphocytes at the site of malignancy. The significant adverse effects of HD IL-2 are largely a result of severe vasodilation and capillary leak syndrome, and include hypotension, arrhythmias, and liver and renal toxicities. Its administration requires an inpatient intensive care-like setting, thus it is recommended in patients with few comorbidities and an excellent performance status. There is a 1–2% risk of mortality with IL-2, which highlights the importance of choosing a well-suited patient for this treatment modality. 23

Historically, HD IL-2 was first used in a combinational biochemotherapy (BCT) setting, usually involving cisplatin, vinblastine, and dacarbazine (CVD) or cisplatin, vinblastine, and temozolomide (CVT), plus the biologic agents IFN α and IL-2. However, a modest increase in survival came with a substantial increase in toxicity. 24 More recently, as drugs such as ipilimumab demonstrate durable responses, the role of HD IL-2 as a single agent is becoming more controversial. One rational approach is to combine the two approved immunotherapies for stage IV melanoma, ipilimumab and IL-2. No data are currently available regarding the correct sequencing of immunotherapies. Some melanoma experts believe that IL-2 is best used very early on in therapy when subjects have more limited disease (M1a disease) and good performance status. A small study has indicated that there may be a higher response rate (47%) in patients with NRAS-mutant melanoma, but further validation of this finding is needed. 25 A 2005 study in 36 patients with advanced melanoma who received a combination of ipilimumab (0.1–3 mg/kg every 3 weeks) and IL-2 demonstrated an overall response rate of 22%. 26 Studies evaluating the role of ipilimumab with adoptive cell therapy are ongoing. Another approach to extend or enhance the efficacy of HD IL-2 or ipilimumab therapy is to combine immunotherapy with BRAF inhibitors for treatment of patients with BRAFV600-mutant advanced melanoma. 27 Preclinical studies showed an increase in melanoma antigen expression and the number of tumor-infiltrating lymphocytes in tumor biopsies after BRAF inhibitor therapy, which correlated with a reduction in tumor size and an increase in necrosis. 28, 29 Current efforts are examining tumor biopsies from patients receiving vemurafenib to assess the mechanisms and kinetics of T-cell accumulation within tumors and characterize the specificity and function of immune infiltrating cells to design more successful combination treatments of BRAF inhibitor and immunotherapy regimens.

Interleukin-3

IL-3 was first described as a T-cell product involved in the pathogenesis of Moloney leukemia virus-induced T-cell lymphomas. 30 This molecule is of interest because of its ability to stimulate multilineage hematopoietic progenitors both in vitro and in vivo . 30–37

Biologic properties of IL-3

In vitro , IL-3, in combination with other cytokines, such as stem cell factor (SCF), IL-6, IL-1, GM-CSF, GM-CSF, erythropoietin (EPO), or thrombopoietin (TPO), induces the proliferation of colony-forming unit (CFU)-GM, CFU-Eo, CFU-Baso, BFU-E, and CFU-GEMM in semisolid medium and stimulates the proliferation of purified CD34+ cells in suspension culture. 31 Indeed, IL-3 is combined with other cytokines, in particular SCF, IL-6, IL-1, FL, G-CSF (granulocyte colony-stimulating factor), and/or EPO, in almost all protocols to expand hematopoietic stem and progenitor cells in vitro .

IL-3 in the clinic

IL-3 has been used in a variety of clinical trials peripheral blood stem cell mobilization, postchemotherapy and transplantation, and bone marrow failure states. The majority of studies show only modest effects of IL-3 by itself but significant salutary effects in conjunction with other growth factors. For instance, in mobilization studies, treatment with IL-3 did not mobilize by itself but significantly potentiated G-CSF-induced yield of all progenitor cell types used to restore hematopoiesis after high-dose chemotherapy. After transplantation, the combination of IL-3 and GM-CSF proved more efficient to support bone marrow engraftment than IL-3 or GM-CSF alone. The combination of IL-3 and GM-CSF was more efficient than G-CSF for supporting platelet recovery but was of similar benefit for the reconstitution of myelopoiesis. Following chemotherapy, IL-3 was found to attenuate neutropenia and/or thrombocytopenia in some but not all clinical studies.

Interleukin-4 and interleukin-13

IL-4 and IL-13 are closely related. 38–40 They share biologic and immunoregulatory functions on B cells, monocytes, dendritic cells, and fibroblasts. Both IL-4 and IL-13 genes are located in the same vicinity on chromosome 5. The major regulatory sequences in the IL-4 and IL-13 promoters are identical, thus explaining their restricted expression pattern in activated T cells and mast cells. Furthermore, the IL-4 and IL-13 receptors are multimeric and share at least one common chain—IL-4RA. This, together with similarities in IL-4 and IL-13 signal transduction, explains the striking overlap of biologic properties between these two cytokines. The inability of IL-13 to regulate T-cell differentiation due to a lack of IL-13 receptors on T lymphocytes, however, represents a major difference between these cytokines. Therefore, despite the impact redundancy of these two molecules, regulatory mechanisms are in place to guarantee their distinct functions.

Biologic activities of IL-4 and IL-13

IL-13 elicits many, but not all, of the biologic actions of IL-4. IL-4 is, however, distinguished from IL-13 by its T-cell growth factor activity and its ability to drive differentiation of Th0 precursors toward the Th2 lineage. Th2 cells secrete IL-4 and IL-5 and lead to a preferential stimulation of humoral immunity. In contrast, Th1 cells, which produce IL-2 and IFN-γ, lead to a preferential stimulation of cellular immunity.

IL-4 and IL-13 possess potent antitumor activity in vivo in mice. 41 They can inhibit the proliferation of some human cancer cell lines in vitro and in vivo in nude mice. A similar antiproliferative effect of IL-13 on human breast cancer cells has been described. Moreover, a chimeric protein composed of IL-13 and a truncated form of Pseudomonas exotoxin A exhibits specific cytotoxic activity toward human RCC but not against normal hemopoietic cells. 42

Clinical trials of IL-4

Despite the preclinical promise of IL-4, to date, clinical trials in humans demonstrated that although the molecule is safe and nontoxic, only sporadic antitumor activity is observed in a variety of cancers, including melanoma, lung cancer, and AIDS-related Kaposi’s sarcoma. 43–45

Interleukin-6

IL-6 was first cloned in 1986. 46 It is a typical cytokine, exhibiting functional pleiotropy and redundancy. IL-6 is involved in the immune response, inflammation, and hematopoiesis. IL-6 is a 21- to 30-kDa glycoprotein of 212 amino acids that binds to a specific receptor that requires the same 130-kDa membrane glycoprotein for mediation of signal transduction, as has been described for several cytokines, including IL-2. 47, 48

Biologic activities of IL-6

IL-6 affects the hypothalamic-pituitary axis, bone resorption, and both the humoral and cellular arms of the immune system 49–53 and is a potent and essential factor for the normal development and function of both B and T lymphocytes. 54 IL-6 is also involved in the differentiation of myeloid leukemic cell lines into macrophages, megakaryocyte maturation, neural differentiation, and osteoclast development. As a major inducer of acute-phase protein synthesis in hepatocytes, 55 this cytokine may play a role in the pathogenesis of sepsis.

IL-6 acts as a growth factor for myeloma/plasmacytoma, keratinocytes, mesangial cells, RCC, and Kaposi sarcoma and promotes the growth of hematopoietic stem cells. On the other hand, IL-6 inhibits the growth of myeloid leukemic cell lines and certain carcinoma cell lines. Significant correlations between serum IL-6 activity and serum levels of acute-phase proteins have been demonstrated in a variety of inflammatory conditions. IL-6 has been implicated as a mediator of B symptoms in lymphoma. 56 Elevated serum IL-6 levels have also been associated with an adverse prognosis in both Hodgkin lymphoma and non-Hodgkin lymphoma (NHL). 57–60 In diffuse large-cell lymphoma, IL-6 levels were found to be the single most important independent prognostic factor selected in multivariate analysis for predicting complete remission rate and relapse-free survival. 58 IL-6 levels may also be exploitable as a prognostic factor in RCC and multiple myeloma (MM), and high levels are observed in prostate and ovarian cancers. IL-6 probably also plays an etiologic role in the systemic manifestations of the lymphoproliferative disorder Castleman’s disease. 61 High IL-6 levels are also an adverse prognostic factor in pancreatic cancer. 62

IL-6 in the clinic

In patients undergoing chemotherapy or autologous transplantation, IL-6 has minimal to no platelet-enhancing activity at tolerable doses. Toxicity includes fever and anemia. 63–65 IL-6 has also been tested as an antitumor agent in melanoma and RCC. Response rates have been low (<15%). 55 Because high levels of IL-6 correlate with an adverse outcome in many cancers and function as an autocrine/paracrine growth factor in some tumors, clinical studies of an IL-6 inhibitor may be worthwhile.

IL-6 is one of the most ubiquitously deregulated cytokines in cancer, and increased levels of IL-6 have been observed in virtually every tumor studied. Preclinical and translational findings support a role for IL-6 in diverse malignancies, including breast, lung, colorectal, ovarian, prostate, pancreatic cancers, MM, glioma, melanoma, RCC, leukemia, lymphoma, and Castleman’s disease, and provide a biologic rationale for targeted therapeutic investigations. Various compounds antagonize IL-6 production, including corticosteroids, nonsteroidal anti-inflammatory agents, estrogens, and cytokines. Targeted biologic therapies include IL-6 conjugated toxins and monoclonal antibodies directed against IL-6 and its receptor. As an example, a chimeric murine antihuman IL-6 antibody, CNTO 328, has been used in a phase 1 trial in subjects with B-cell NHL, MM, and Castleman’s disease. 66 The treatment resulted in tumor response and disease control, especially in Castleman’s disease, where striking responses have been seen. 67

Interleukin-7

IL-7 was identified and cloned on the basis of its ability to induce proliferation of B-cell progenitors in the absence of stromal cells. 68–76 It is now known that this cytokine is secreted by stromal cells in the bone marrow and thymus and is irreplaceable in the development of both B and T cells. 69–71 Indeed, the nonredundant nature of IL-7 is underscored by the observation that ablation of IL-7 or parts of the IL-7 receptor in gene knockout mice ineluctably leads to a major defect in lymphocyte development.

Biologic activities of IL-7/IL-7 receptor

While most single cytokine knockout mice show relatively normal B- and T-cell compartments, indicating that many cytokine functions are redundant, IL-7-deficient mice present with striking lymphocyte depletion in both the thymus and bone marrow. Collectively, these genetic experiments identify clearly distinct in vivo roles for various lymphoid factors. IL-2 and IL-4 function by influencing mature lymphocyte populations during immune responses, whereas IL-7 plays a singularly dominant role for the production and expansion of lymphocytes. The upregulation of IL-7R occurs at the stage of the clonogenic common lymphoid progenitor that can give rise to all lymphoid lineages at a single-cell level. 74 There are at least three principal means by which IL-7R-mediated signals act in lymphocyte development: enhancement of proliferation, triggering of lineage-specific developmental programs, and maintenance of viability of appropriately selected cells.

High IL-7 levels are found in states of T-cell depletion and may, therefore, play a role in promoting T-cell expansion. 75 High levels of IL-7 are also found in CLL and Burkitt lymphoma, and transgenic mice overexpressing the IL-7 gene show dramatic changes in lymphocyte development, which, in some instances, can result in the formation of lymphoid tumors. 76

Interleukin-8

IL-8 was first identified in 1987 as a potent, proinflammatory chemokine that induces trafficking of neutrophils across the vascular wall (chemotaxis). 77 This molecule belongs to a chemokine superfamily whose members include neutrophil-activating peptide-2, platelet factor-4, growth-related cytokine (GRO), and IFN-inducible protein-10, all of which are responsible for the directional migration of various cells. 78 IL-8 receptor demonstrates strong homology to a gene encoded by human herpesvirus-8 (HHV-8). 79, 80

Biologic activity of IL-8

The chemotactic agents generated by inflammatory stimuli recruit circulating leukocytes, in particular neutrophils, for defensive purposes and direct them to injury sites. Among the neutrophil-affecting chemokines, IL-8 is one of the most potent. 81 On exposure to a chemokine, neutrophils are activated, and within seconds, their shapes change. The process of shape change is crucial. It is modulated by perturbations of cellular integrins and the actin cytoskeleton. The activation and upregulation of integrins also permits the adherence of neutrophils to the endothelial cells of the vessel wall, to allow for subsequent migration into the tissues. Leukocytes follow the IL-8 concentration gradient and accumulate at the location of elevated concentration. These processes play a fundamental role in the host defense as activated leukocytes act to kill and engulf invading bacteria at the site of injury.

IL-8 can induce tumor growth, an effect attributed to its angiogenic activity. On the one hand, the administration of anti-IL-8 to SCID mice bearing xenografts of IL-8-expressing human lung cancer has been shown to have beneficial effects. 82 On the other hand, antitumor effects of IL-8 have also been reported. Of interest in this regard is the fact that increased levels of IL-8 have been discerned in lung carcinomas and in melanomas. IL-8 may be a growth factor for pancreatic cancer and for melanoma. 78 In melanomas, IL-8 levels correlate with the growth and metastatic potential of the tumor cells, and exposure of the cells to IFN (an agent with known antitumor activity in melanoma) decreases IL-8 levels and cancer cell proliferation. 83 Blocking IL-8 or IL-8R has been suggested as a therapeutic strategy. 78

Interleukin-9

Human IL-9 was initially identified and cloned as a mitogenic factor for a human megakaryoblastic leukemia. Subsequently, IL-9 targets were found to encompass a wide range of cells. 84, 85

Biologic activities of IL-9

Cellular elements responsive to IL-9 include erythroid progenitors, human T cells, B cells, fetal thymocytes, thymic lymphomas, and immature neuronal cell lines. 84

IL-9 can support the clonogenic maturation of erythroid progenitors in the presence of EPO. In contrast, granulocyte or macrophage colony formation (CFU-GM, CFU-G, or CFU-M) is usually not influenced by IL-9. IL-9 is more effective on fetal than adult progenitors and in cells that are activated. In addition to its proliferative activity, IL-9 also seems to be a potent regulator of mast cell effector molecules.

There is an interesting paradox between the unresponsiveness of normal T cells to IL-9 and the potent activity of this molecule on lymphoma cells. This contrast is illustrated by the observation that murine T cells acquire the ability to respond to IL-9 after a long period of in vitro culture, while they simultaneously acquire characteristics of tumor cell lines. Observations made with transgenic mice also demonstrate the oncogenic potential of dysregulated IL-9 production as 5–10% of mice that overexpress this cytokine develop lymphoblastic lymphomas. 85 In line with these data, constitutive IL-9 production by human Hodgkin lymphomas and large-cell anaplastic lymphomas has now been clearly documented. 84 Even so, the pathophysiologic role of IL-9 remains elusive.

Interleukin-10

IL-10 is a pleiotropic cytokine discovered in 1989 as an activity produced by murine type 2 helper T cells (Th2). 86, 87 It was initially designated as cytokine synthesis inhibitory factor because of its ability to inhibit the production of certain cytokines. 88 Of interest, IL-10 exhibits strong DNA and amino acid sequence homology to an open reading frame—BCRF1—in the Epstein–Barr virus (EBV) genome. 88 Indeed, the BCRF1 protein product displays many of the biologic properties of cellular IL-10 and has, therefore, been termed viral IL-10.

Biologic activities of IL-10

IL-10 inhibits the synthesis of Th1-derived cytokines, including IL-2, IFN-γ, GM-CSF, and lymphotoxin and of monocyte-derived IL-1α and β, IL-6, IL-8, TNF-α, GM-CSF, and G-CSF. Exogenous IL-10 can also suppress expression of IL-10. 87 At the same time, IL-10 induces the synthesis of the IL-1 receptor antagonist by macrophages. IL-10 also suppresses the CD28 costimulatory pathway and hence acts as a decisive mechanism in determining if a T cell will contribute to an immune response or become anergic.

From the molecular standpoint, IL-10 suppresses cytokine expression at a transcriptional and posttranscriptional level. 89 Both these mechanisms appear to require new protein synthesis. At a cellular level, Th1 cytokines synthesis inhibition is mediated indirectly through the effect of IL-10 on APC, as suppression occurs when macrophages, but not B cells, are used as APC. 90

In the presence of monocytes/macrophages, IL-10 inhibits proliferation of resting T cells, including Th0, Th1, and Th2 CD4+ T-cell clones. This inhibition can only be partially reversed by high concentrations of IL-2, suggesting that the reduced proliferation is only partially a reflection of reduced IL-2 production. IL-10 can also enhance the cytotoxic activity of CD8+ T cells. All these effects support an important role of IL-10 in regulating inflammatory responses. In contrast to the inhibitory effects on other lineages, IL-10 has a stimulatory effect on B cells and mast cells. 91 For instance, IL-10 strongly stimulates proliferation and differentiation of activated B cells.

The role of IL-10 in cancer should be considered within the frame of a highly complex biological puzzle. It is known that IL-10 can have pleiotropic effects on adaptive and innate immunity cell mediators. Although several studies show that IL-10 can actively mediate immune suppression, some experimental models describe relatively opposite conclusions. Recent data on the relationship between IL-10 and anticancer immunity support an effective immune attack against malignant cells, which challenges the common belief that IL-10 acts as an immunosuppressive factor promoting tumor immune escape.

Interleukin-11

Originally characterized as a thrombopoietic factor, IL-11 is now known to be expressed and have activity in a multitude of other systems, including the gut, testes, and the central nervous system. 92, 93 Clinically, this cytokine has been approved by the FDA for amelioration of chemotherapy-induced thrombocytopenia.


2. The Glioblastoma TME

In common with other solid tumours, the glioblastoma microenvironment harbours an array of non-malignant (stromal) cell types in addition to the cancer cells themselves [18,19]. The main stromal cell types in glioblastoma are cells of the immune system𠅍iscussed in detail below𠅊nd cells associated with the structure and function of blood vessels (endothelial cells and pericytes) [20]. In contrast to most other tumour types, fibroblasts are not known to be a significant component of the glioblastoma TME. Vessels promote the growth and survival of glioblastoma cells, by facilitating blood perfusion and hence the provision of essential oxygen and nutrients. In addition, the perivascular zone can serve as a specialised niche to support the survival and function of glioma stem cells (GSCs), which are self-renewing, multipotent cells thought to produce the bulk of the malignant cells in glioblastoma [21]. In contrast, the role of immune cell populations is more complex, and the balance of pro-tumour versus anti-tumour populations likely plays a critical role in determining the trajectory of tumour growth and spread.

For many years, the brain was viewed as an immune privileged site, protected from the regular surveillance systems that operate in the periphery [22]. This concept was supported by a perceived lack of lymphatic vessels in the brain, thereby separating the brain from central lymphocyte circulation pathways, and the presence of the blood𠄻rain barrier (BBB), which restricts the entry of leukocytes from the blood. However, functional lymphatic vessels have recently been discovered to line the dural sinuses of mice, and potentially analogous structures exist in human dura [23], suggesting that the brain is not in fact immunologically separate from the periphery. In addition, the BBB is frequently compromised in glioblastoma [22], and priming of tumour-specific T cells has been detected in glioblastoma patients [16]. Thus, it is clear that glioblastoma tumours interact with the immune system, but immune-mediated tumour control is likely hampered by an overwhelmingly immunosuppressive TME.

Cells of the myeloid lineage are a major component of the glioblastoma TME [20,24,25,26]. In fact, these cells are reported to constitute a remarkable 30�% of the glioblastoma tumour mass. Myeloid cell types within the glioblastoma TME include brain-resident microglia and infiltrating macrophages, which are collectively referred to as glioma-associated microglia and macrophages (GAMs), as well as myeloid-derived suppressor cells (MDSCs). Microglia are derived from primitive yolk sac progenitors that enter the brain during embryogenesis and reside as a local resident population throughout life [25,26]. They play many critical roles under conditions of homeostasis, including synaptic pruning and the regulation of sleep and memory, as well as serving as local sensors of neuronal damage and infection. In contrast, infiltrating macrophages are thought to enter the tumour as blood-borne monocytes, which are recruited in response to inflammatory stimuli, and then differentiate to macrophages once they enter the TME [19,25]. Finally, MDSCs are immature myeloid lineage cells with inherent immunosuppressive properties [19,27]. They arise through a pathological (tumour-driven) block in normal myeloid differentiation pathways, leading to the accumulation of an abnormal population of partially differentiated myeloid cells. MDSCs exploit a number of immunosuppressive mechanisms to inhibit adaptive immune responses, including the depletion of nutrients required for effective T cell responses, the generation of oxidative stress conditions that inhibit T cell function, and the activation and expansion of regulatory T cells (Tregs) [19,27].

In contrast to the inherent immunosuppressive properties of MDSCs, macrophages and microglia are more plastic cell types that can be readily polarised according to their local environment, resulting in highly divergent functions [19,25,27]. According to the M1/M2 paradigm, 𠆌lassically activated’ macrophages (M1) assume an inflammatory phenotype characterised by efficient phagocytosis and antigen presentation, and abundant production of pro-inflammatory cytokines. In contrast, 𠆊lternatively activated’ macrophages (M2) largely produce anti-inflammatory cytokines and support tissue remodelling and matrix deposition. However, this proposed dichotomy is largely based on in vitro studies, and it is likely that macrophages in tissues rarely exist in such clearly defined states. Indeed, although GAMs clearly can express markers of the immunosuppressive M2 phenotype, including TGF-β, IL-10, CD163 and CD204, unbiased transcriptomic analyses characterised GAMs in patient tissues as more in keeping with a non-polarised M0 phenotype [28], or a mixed M1/M2 phenotype [29].

GAMs are most commonly considered to have a pro-tumorigenic function. For example, they possess multiple immunosuppressive properties, secrete factors that actively promote tumour cell proliferation and invasion, and in certain animal models it has been demonstrated that depletion of GAMs can significantly reduce tumour growth [19,25,26,30,31]. Furthermore, in patient glioblastoma tissues, the proportion of M2 macrophages is reported to positively correlate with the rate of tumour cell proliferation [32]. However, several other studies have shown conflicting results. For example, in some animal models GAM depletion actually enhances tumour growth [25,33], while a high frequency of either total or M2-phenotype GAMs in patient glioblastoma tissues correlate with improved survival [29]. The role of GAMs in the growth and progression of glioblastomas is therefore likely to be complex and highly context-dependent and requires further study.

Although GAMs represent the predominant immune cell population in glioblastoma, significant populations of lymphocytes are also present. These are primarily T cells, although natural killer (NK) cells and B cells have also been identified in human glioblastomas, the latter being relatively rare [19]. The T cell population in glioblastoma generally displays a profoundly exhausted phenotype, characterised by expression of LAG3, TIGIT, CD39 and especially programmed cell death 1 (PD1) [34]. T cell anti-tumour activity can also be inhibited by indoleamine 2,3-dioxygenase (IDO), an enzyme present in the TME responsible for catalysing the oxidation of tryptophan to downstream metabolites belonging to the kynurenine pathway. This can, through a variety of mechanisms, lead to T cell dysfunction, an effect that is particularly pronounced in the setting of advanced age [35,36]. Furthermore, Tregs are enriched in glioblastoma lesions compared to peripheral blood, and are expected to further inhibit the function of effector T cells, as well as NK cells [19,37]. This severely immunosuppressed microenvironment likely contributes to the apparent inability of infiltrating T cells to control tumour growth. This effect is compounded by the inherent low immunogenicity of glioblastoma tumours, which generally lack the high mutation rate thought to be required for robust anti-tumour T cell responses [38]. However, it is worth noting that a high effector CD8+ T cell frequency in patient glioblastoma tissues is associated with prolonged survival [39]. In addition, ICI therapies, which promote anti-tumour T cell responses, can induce regression of glioblastomas harbouring germline mismatch repair deficiency, which are characterised by a greatly elevated mutation rate [12]. Thus T cells may have the potential to control glioblastoma growth in circumstances where their frequency and function are optimal, highlighting the therapeutic potential of T cell-based therapies in this disease.


Interferon Types

Interferons are currently classified into three groups: type I, type II and type III IFNs. The type I IFNs include all IFNαs, IFNβ, IFNε, IFNκ, IFNω and IFNν. Humans have 12 different IFNαs and a single IFNβ. Type I IFN genes are clustered on the human chromosome 9. Each subtype is encoded by its own gene and regulated by its own promoter, and none of them contain introns. The different IFNαs and IFNβ differ substantially in their specific antiviral activities and in the ratios of antiviral to antiproliferative activities. However, the molecular basis of these differences is not yet known. All type I IFNs bind to the same interferon alpha/beta receptor (IFNAR) which consists of two major subunits: IFNAR1 and IFNAR2c (the βL subunit).

There is only one class II IFN, IFNγ. Interferon gamma is produced by T lymphocytes when stimulated with antigens or mitogens. IFNγ binds to a distinct receptor, the interferon gamma receptor (IFNGR) consisting of the two subunits IFNGR1 (previously α chain) and IFNGR2 (previously β chain or accessory factor).

The more recently described type III IFNs IFNλ2, IFNλ3 and IFNλ1 are also known as IL28A, IL28B and IL29 respectively. The same as type I IFNs, they are also induced by viral infections. They signal through the IFN-λ receptor consisting of the IL-10R2 chain shared with the IL-10 receptor, and a unique IFNλ chain.


Role of Chemokines in Neutrophil Heterogeneity

Despite the previous belief that differentiated neutrophils were a homogeneous population, the existence of different circulating subsets was demonstrated in varied health and disease contexts, both in mice and humans (51) (Figure 1). A consensus on the phenotype of these subpopulations is still missing and under steady-state conditions heterogeneity may arise mainly from the aging process of circulating neutrophils (52). Indeed, neutrophils oscillate in a circadian manner in numbers, morphology, and phenotype (53, 54). This process is regulated by gut microbiota (55) and is controlled by neutrophils themselves through the circadian expression of the transcription factor Bmal1 that controls the production of CXCL2. In turn, CXCL2 acting on CXCR2 induces neutrophil aging (56).

During inflammatory conditions, increased levels of a neutrophil circulating population that shared characteristics with BM immature neutrophils was described both in mice and humans. These cells express low levels of CD16 and are CD10 − (57�). The functional properties of this subset are still controversial, they were described having either immunosuppressive activity (60) or promoting T-cell survival and proliferation (57).

Other circulating neutrophils subpopulations were described: olfactomedin 4 (OLFM4)-positive neutrophils in healthy donors (61), T-cell receptor (TCR)�sed variable immunoreceptor neutrophils (62), and CD177 + neutrophils during inflammatory diseases both in mice (63) and humans (64).

In addition, a reverse transendothelial migrating neutrophil subset (rTEM) was described in a murine model of sterile injury (65). These neutrophils are CD54 hi and, in order to reverse transmigrate into vasculature, downregulate CXCR1. Concomitantly, they upregulate CXCR4 to go into the lungs, before being cleared in BM (66). This subset represents a phenotypically and functionally distinct population different from circulating neutrophils (CD54 lo CXCR1 hi ) and express vascular endothelial growth factor receptor (VEGFR) 1, indicating a possible role in angiogenesis (67, 68). Similar cells, with increased levels of CD54 and CD18 and downregulation of CD62L and CXCR1 and 2, were found in patients with chronic inflammatory diseases, suggesting a role of rTME neutrophils in the persistence of inflammation (67). Moreover, around 1% of circulating neutrophils after ischemia-reperfusion were found to be CD54 hi and producing ROS into lungs (65). On the contrary, neutrophils that migrate away from the inflammation site in interstitial tissues are called reverse interstitial migration (rIM) neutrophils and are supposed to contribute to the resolution of inflammation. The role of chemokine receptors in this process is still not clear (69).

Finally, in circulation it is possible to identify aged or senescent neutrophils (54, 70, 71). Ex vivo aging experiments have shown that neutrophils kept in culture downregulate the expression of CXCR2 (44) and re-express CXCR4 in a time-dependent way (22), suggesting a preferentially homing of senescent cells to the BM in response to CXCL12 (21). In mice aged neutrophils display circadian oscillations and, in addition to high levels of CXCR4, are characterized by an increased surface expression of CCR5 and decreased expression of CD62L (53, 72). CXCR4 upregulation seems involved not only in guiding neutrophils back to the BM but also in their migration within the marrow tissue in order to be engulfed with greater efficacy by macrophages (17, 19, 53, 54, 72). CCR5 was reported to work as a chemokine scavenger promoting the resolution of the inflammatory response (73). Aged neutrophils were found in lungs, where pulmonary vasculature expresses CXCL12, and this could either supply the pool of circulating neutrophils or respond to injury (45, 68).

New data from single cell sequencing of murine circulating neutrophils confirm the presence of three transcriptionally different neutrophil subpopulations. The first expresses high levels of inflammatory genes and the highest levels of CXCR2 arising mainly from BM mature neutrophils. The second expresses interferon-stimulated genes and derives from BM immature neutrophils. Both populations mature in an aged subset CXCR4 positive with high phagocytic activity and still highly transcriptionally functional (41). The correlation of these subpopulations of neutrophils with the others described in the foregoing is still missing. In addition, the role of chemokines in the mobilization and function of these neutrophil subpopulations is not known. Of relevance, at least in mice, mobilization of immature neutrophils could be CXCR2 independent because they are referred to as CXCR2 negative (44).

Finally, neutrophil heterogeneity has been described in tumors where tumor-associated neutrophils (TANs) can exist in two different functional states: N1 proinflammatory and antitumoral subset and an antiinflammatory tumor promoting N2 population, distinguished for the expression of adhesion molecules, cytokines and inflammatory mediators, chemokines, and chemokine receptors (4, 74). N1 phenotype has been associated with IFN-β polarization both in mice and humans. These cells have an activated phenotype (CD62L lo CD54 + ) express the chemokine receptors CCR5, CCR7, CXCR3, and CXCR4 and produce the proinflammatory chemokines and cytokines: CCL2, CXCL8, CCL3, and interleukin-6 (IL-6). Moreover, this subset has been associated with stimulation of T-cell responses and ROS production (4, 75, 76). In contrast, N2 neutrophils are induced by transforming growth factor- β (TGF-β) stimulation. Protumoral N2 neutrophils display high levels of CXCR4, VEGF, and matrix metalloproteinase 9 (MMP-9) (77), and produce high levels of CCL2, CCL5, neutrophil elastase (NE), cathepsin G (CG), and arginase 1 (78�).

Therefore, results obtained in preclinical mouse models and in humans suggest that the interplay between CXCR2 and CXCR4 dictates not only BM neutrophil mobilization and retention but also neutrophil diversity in homeostasis. CXCR2 signaling promotes neutrophil aging and CXCR4 guides their homing back to the BM. Furthermore, diversity of tissue infiltrating neutrophils is also associated with a distinct pattern of chemokine receptors in particular N1 neutrophils express inflammatory CC chemokine receptors important for their effector functions (see later).


Diagnostic value of cytokines and chemokines in lyme neuroborreliosis

The aims of the present study were to assess the concentrations of different cytokines and chemokines in blood serum and cerebrospinal fluid (CSF) samples of patients with Lyme neuroborreliosis and to identify the possible marker(s) that would enable a distinction between clinically evident and suspected Lyme neuroborreliosis, as well as between Lyme neuroborreliosis and tick-borne encephalitis (TBE). Our additional interest was to evaluate the relationship between cytokine and chemokine concentrations and Borrelia burgdorferi sensu lato isolation from CSF, as well as intrathecal synthesis of specific borrelial antibodies. We found that higher concentrations of CXCL13 and lower concentrations of interleukin 10 (IL-10) in serum were associated with higher odds for clinically evident Lyme neuroborreliosis compared to suspected Lyme neuroborreliosis, as well as to TBE. The concentrations of IL-2, IL-5, IL-6, IL-10, and CXCL13 in the CSF were higher in patients with evident Lyme neuroborreliosis than in those who were only suspected to have the disease. A comparison of CSF cytokine and chemokine levels in patients with and without intrathecal synthesis of specific borrelial antibodies revealed that CXCL13 CSF concentration is significantly associated with intrathecal synthesis of borrelial antibodies. A comparison of the cytokine and chemokine CSF concentrations in patients with clinically evident Lyme neuroborreliosis according to CSF culture results revealed that higher concentrations of gamma interferon (IFN-γ) were associated with lower odds of Borrelia isolation. Although several differences in the blood serum and CSF concentrations of various cytokines and chemokines between the groups were found, the distinctive power of the majority of these findings is low. Further research on well-defined groups of patients is needed to appraise the potential diagnostic usefulness of these concentrations.

Figures

CXCL13 concentrations in sera from…

CXCL13 concentrations in sera from patients with clinically evident and microbiologically confirmed Lyme…

CXCL13 concentrations in CSF from…

CXCL13 concentrations in CSF from patients with clinically evident and microbiologically confirmed Lyme…


CONCLUSIONS

IL-36α, IL-36β, and IL-36γ signal through the same receptor and appear to have the identical effects on cells and in vivo, which calls into question the need for 3 ligands. There is evidence that the expression patterns for IL-36 ligands are different, with IL-36γ as the most highly expressed and inducible in skin and lung. In addition, there is very little sequence homology among the 3 ligands around the site of cleavage, suggesting differential mechanisms for producing the active form. Therefore, whereas all IL-36 ligands appear to have the same activity, their expression and activation are likely differentially regulated. More work needs to be done to address the regulation of IL-36 cytokines at the expression level, as well as to define and understand the forces governing their truncation. Whereas it is clear that processing these ligands must occur for their activity, the identity of the enzymes involved in the regulation of this process is unknown. Demonstration that truncation of IL-36 ligands occurs in vivo and under certain circumstances and the search for the proteases involved are active areas of research.

Recent data have emerged demonstrating that IL-36 cytokines play a crucial role in the skin, most dramatically evidenced by the genetic variants in IL36RN, leading to the life-threatening disease GPP, the marked up-regulation of IL-36 cytokines in psoriatic lesional tissue, and the ability of IL-36R neutralization to decrease the inflammation and skin thickening of human psoriatic tissue when engrafted onto mice. Despite convincing evidence for a role for these cytokines in skin pathology and the strong ability to induce inflammatory responses in the lung, it is not yet clear whether IL-36 plays a role in pathologic conditions in the lung, such as asthma or COPD, and whereas IL-36 cytokines are expressed in other tissues, including the kidney, brain, and gut, it is unclear what role they play in the physiology or pathophysiology of these tissues. In addition, whereas IL-36 clearly acts on DCs to enhance their maturation and production of cytokines and acts on nai¨ve T cells in mice, the full role of these cytokines on human immune cells is not yet clear. Can IL-36 act on human αβ T cells or γδ T cells if provided in the right context? Likewise, the role of IL-36 on macrophages, in particular, of different subtypes and activation states has not been thoroughly explored. Similar to its fellow family members—IL-1, IL-18, and IL-33—IL-36 likely plays a prominent role in human health, and further studies assessing the expression and activity of IL-36 in the context of disease will likely uncover additional roles for these cytokines.


Interleukins

Interleukins are a group of cytokines that act as chemical signals between white blood cells. Interleukin-2 (IL-2) helps immune system cells grow and divide more quickly. A man-made version of IL-2 is approved to treat advanced kidney cancer and metastatic melanoma. IL-2 can be used as a single drug treatment for these cancers, or it can be combined with chemotherapy or with other cytokines such as interferon-alfa.

Side effects of IL-2 can include flu-like symptoms such as chills, fever, fatigue, and confusion. Some have nausea, vomiting, or diarrhea. Many people develop low blood pressure, which can be treated with other medicines. Rare but potentially serious side effects include an abnormal heartbeat, chest pain, and other heart problems. Because of these possible side effects, if IL-2 is given in high doses, it must be done in a hospital.

Other interleukins, such as IL-7, IL-12, and IL-21, continue to be studied for use against cancer too, both as adjuvants and as stand-alone agents.


FINAL REMARKS

The chemokine/cytokine network is profoundly involved in the control of HIV infection as it is both a main target of the HIV-induced dysregulation and, at the same time, a complex modulator of the susceptibility of immune cells to infection and replication. The recent findings that chemokines can affect binding, entry, and post-entry events, and that cytokines can influence HIV infection by modulating the expression of chemokines and their receptors as well as the extent of viral replication, further support the general model that multiple steps of the life cycle of HIV are regulated by this network (Table 2and 3). Macrophages serve as a major reservoir and vehicle for dissemination of HIV in different tissues. Thus, HIV harbored in these cells may escape immune surveillance and antiviral therapy. Although highly active antiretroviral therapy (HAART) significantly suppresses viral replication, ongoing viral replication and spreading has been observed during HAART, especially in macrophages with respect to resting T cells [59 ]. Therefore, macrophages can play a key role in regulating the intensity and progression of HIV disease even during therapy, and their secretory products have been implicated in the pathogenesis of AIDS [234, 10, 11 ]. In conclusion, exploitation of knowledge on the interactions between HIV and macrophages in their milieu of cytokines and chemokines may lead to novel and more effective strategies of preventive or therapeutic interventions. Unraveling this complex network of interactions is of relevance for the eradication of tissue viral reservoirs of long-lived, latently infected cells, as well as for the development of molecules capable of interfering with HIV entry by targeting chemokine receptors.