Where is neutrophils made

Several subsets of neutrophils are found in tissues under homeostatic conditions. We still have much to learn on how these different neutrophil subtypes are generated and recruited to tissues. In addition, various subsets of neutrophils with distinct properties are also detected in pathological conditions particularly in inflammation and in cancer Silvestre-Roig et al.

Neutrophils are the first cell type recruited to sites of inflammation. From there, they can switch phenotypes and generate various subpopulations with different cell functions. Neutrophils can also interact, directly, or via cytokines and chemokines, with other immune cells to modulate both innate and adaptive immune responses. There is not a complete understanding of these subpopulations of neutrophils, but some clear examples showing that bona fide inflammatory subsets occur are mentioned next.

Upon infection with an antibiotic-resistant Staphylococcus aureus , two clear subsets of murine neutrophils can be observed. They differ in cytokine production, macrophage activation potential, expression of TLR, and expression of surface molecules. In systemic inflammation condition, another subset of neutrophils is generated with low doses of endotoxin. These cells have a hypersegmented nucleus and display the phenotype CD62 low CD11b hi CD11c hi , which is similar to the one described for murine aged neutrophils Casanova-Acebes et al.

Also, they are capable of inhibiting T lymphocytes by direct cell contact involving the integrin Mac1, and by local delivery of reactive oxygen species ROS Pillay et al. In certain organs such as liver and adipose tissue, few neutrophils are detected in normal homeostatic conditions.

However, upon an inflammatory state induced by experimental obesity, neutrophil numbers increase rapidly and a metabolic imbalance is slowly generated Talukdar et al. First, neutrophils release elastase from azurophilic granules. This enzyme can destroy insulin receptor substrate 1 IRS1 in adipocytes and hepatocytes, and in consequence induce insulin resistance and lipogenesis Talukdar et al.

Supporting a direct role for this neutrophil subtype in metabolic disorders was the observation that altered levels of elastase or its inhibitor 1-antitrypsin are associated with metabolic syndrome and the onset of diabetes Mansuy-Aubert et al.

As mentioned before, neutrophil effector functions are markedly enhanced after priming. When certain metabolic functions are altered, neutrophils can be primed to present stronger pro-inflammatory responses.

In hyperglycemia human and mouse neutrophils are primed to undergo NETosis Wong et al. In consequence, production of neutrophils is enhanced Xiang et al. Similarly, during hypercholesterolemia, neutrophils showed a primed state characterized by elevated ROS production, increased release of myeloperoxidase MPO and increased expression of CD11b Mazor et al.

Together, these reports show that hyperglycemia and hyperlipidemia generate primed, proinflammatory neutrophils that may contribute to diabetes, adipose tissue inflammation, and cardiovascular inflammation.

Yet, the capacity of neutrophils to undergo NETosis can vary with physiological states, suggesting a neutrophil diversity that could be clinically relevant. In fact, several reports indicate that NETs can influence thrombosis Fuchs et al. As mentioned before some metabolic conditions associated with states of chronic inflammation, can increase neutrophil predisposition to form NETs.

Hence, neutrophils from diabetic patients Wong et al. Nowadays NETs have been described as a player of several pathophysiological processes, including vascular diseases, such as atherosclerosis and venous thrombosis Qi et al.

Hyperlipidemia can damage endothelial cells, promoting lipid deposition and plaque formation. This usually characterizes the onset of atherosclerosis. Hyperlipidemia and also hypercholesterolemia induce neutrophilia, which is positively associated with atherosclerotic plaque burden Drechsler et al.

NETs can also regulate cytokine production from macrophages in atherosclerosis Warnatsch et al. Furthermore, proteinases from NETs contribute to plaque instability Hansson et al. After plaque rupture, thrombin-activated platelets interact with neutrophils at the injured site inducing more formation of NETs Stakos et al.

Thus, neutrophils and NETs are major contributors to atherothrombosis. Different from arteries, thrombosis in veins is usually initiated by endothelial injury Di Nisio et al.

Subsequently, damaged endothelial cells secrete massive amounts of von Willebrand factor and P-selectin, which adhere to platelets and recruit leukocytes Etulain et al. Platelets then interact directly with neutrophils and promote the production of NETs Clark et al.

NETs can also stimulate the activation of coagulation cascades Brill et al. These findings and studies in mice showing an association between the risk of venous thrombosis and high neutrophil counts Ramacciotti et al. Another inflammation process in which NETs play an important role is pancreatitis.

Acute pancreatitis is an inflammatory disorder of pancreas for which no specific treatment is available. Important risk factors of acute pancreatitis are formation of gallstones and alcohol abuse Spanier et al. The most severe cases of the disease are associated with mortality, with acute respiratory distress syndrome being the most frequent cause of death in the early phase of the disease Pandol et al.

Obstruction of the pancreatic duct causes blockage of pancreatic secretion, which is accompanied by disorders in organelle function of pancreatic acinar cells Gukovskaya et al.

These disorders promote co-localization of zymogens-containing vesicles and lysosomes leading to formation of co-localization organelles. In co-localized organelles cathepsin B activates trypsinogen to trypsin Halangk et al. The destroyed tissue will eventually be replaced by fatty tissue, typical of chronic pancreatitis Braganza et al. Neutrophils infiltrate the pancreatic parenchyma during this acute inflammatory response Lankisch et al.

In addition, infiltrating neutrophils aggravate inflammation by releasing NETs in the pancreas and at the sites of systemic injury, namely, the lungs Merza et al. NETs incubated in vitro with pancreatic acinar cells led to trypsin activation in these cells, degradation of the NETs by treatment with deoxyribonuclease DNAse abolished the trypsin activation, reduced local acinar damage, and systemic inflammation Merza et al. AggNETs in turn obstruct secretory flow and thereby perpetuate inflammation Leppkes et al.

Therefore, NETs formed after an initial inflammatory stimulus become the activators of further inflammation in the pancreas. Contrary to the situations mentioned above, Gout is a disease where NETs appear to have a positive effect.

Gout is an acute inflammatory reaction originated from precipitation of uric acid in the form of needle-shaped monosodium urate MSU crystals So and Martinon, Aggregates of MSU crystals known as tophi, induce inflammation in the joints and tissues. Local immune cells such as macrophages and dendritic cells take up the crystals via phagocytosis. The MSU-containing phagosomes then fuse with lysosomes. The low pH in phagolysosomes causes a massive release of sodium and consequently raises intracellular osmolarity, which is balanced by passive water influx through aquaporins.

This process dilutes intracellular sodium and potassium concentrations. Cytokine release then leads to a rapid and dramatic recruitment of neutrophils. This neutrophil influx is accompanied by the infamously intense clinical symptoms of inflammation during an acute gout attack So and Martinon, Continuous recruitment of neutrophils to the site of inflammation results in very high neutrophil densities Shah et al. The release of ATP during NETs formation is of high importance since extracellular nucleotides initiate anti-inflammatory clearance of dead cells by mononuclear phagocytes Elliott et al.

In addition, lactoferrin on NETs abrogates further recruitment of neutrophils and thus contributes to the anti-inflammatory action of NETs in highly infiltrated tissues Bournazou et al. Clearly, in this case NETs have a positive anti-inflammatory effect. The inflammation processes described before show that NETs are much more than a simple anti-microbial tool and they can be produced in very different conditions of neutrophil activation.

However, NETs are a double-edged sword. In addition, neutrophils primed by microbiota-derived products can form NETs more easily than neutrophils newly released from the bone marrow Zhang et al. In contrast, the formation of NETs can be blocked by phagocytosis of small microorganisms via the C-type lectin receptor Dectin-1, which acts as a sensor of microorganism size Branzk et al. Dectin-1 downregulates the translocation of neutrophil elastase NE to the nucleus.

This protease promotes NETosis by degrading histones in the nucleus Branzk et al. This could explain in part why in conditions in which phagocytosis of apoptotic cells is compromised, such as SLE and anti-neutrophil cytoplasmic antibody ANCA -associated vasculitis, a state of persistent inflammation is observed.

These findings imply that heterogeneity in neutrophil function for example for NETs formation is also regulated by physiological signals. It is evident that infection and inflammation can regulate the appearance of neutrophil phenotypes with unique properties.

Unfortunately, nowadays we can only incompletely describe the function of these neutrophil subtypes. It would become important to characterize these cells at the level of surface markers, functional responses, and transcriptional profiles in order to understand their role in multiple diseases.

The changes in neutrophil phenotype during cancer are perhaps the most impressive and best studied so far. Neutrophils play important and contradictory roles in cancer development, as reflected by several recent reviews Sionov et al. In tumor-bearing mice, the number of circulating neutrophils increases along with tumor progression.

Similarly, in patients with advanced cancer counts of neutrophils in blood are also increased. It is not clear how tumors can induce neutrophilia, but a common mechanism seems to be the production by tumors of cytokines that influence granulopoiesis, including G-CSF McGary et al. The presence of elevated numbers of neutrophils in the circulation is associated with poor outcome in several types of cancers Schmidt et al. In addition, the presence of neutrophils in tumors also seems to be an indicator of poor outcome Sionov et al.

For this reason, the counts of neutrophils in blood in relation to other leukocytes have been suggested as a prognostic value in cancer. Therefore, the neutrophil to lymphocyte ratio NLR was introduced as a simple and inexpensive biomarker for many types of cancer Peng et al. In general, the blood NLR is high in patients with more advanced or aggressive cancers Guthrie et al.

Despite the simplicity for using the NLR, it has not been accepted in many clinical settings. One reason for this is that neutrophilia can be the result of elevated granulopoiesis and as a consequence, it is not always a bad sign for cancer progression. Another reason is that neutrophilia does not correlate with poor clinical outcome in all types of cancer.

In gastric cancer, for example a high NLR is indicative of positive prognosis Caruso et al. This is indicative of the great plasticity neutrophils have. They can directly kill tumor cells and control cancer Yan et al.

Therefore, the exact role of neutrophils within the tumor is a controversial matter Sionov et al. In several types of cancer, not only an increase in the number of neutrophils in blood is observed, but also an increase in immature myeloid cells Brandau et al. These immature cells are at various stages of differentiation, accumulate in the spleen of tumor-bearing animals, and present an immunosuppressive phenotype that supports tumor progression Nagaraj et al.

Hence, some researchers considerer them to be a bona fide phenotype of neutrophils Pillay et al. However, the relationship among these cells is not clear since immature neutrophils do not have immunosuppressive properties Solito et al. Figure 4. Neutrophil subsets that can be separated through density gradient centrifugation. The bulk of mature normal neutrophils PMN are denser and separate at the bottom of the density gradient. These cells, named high-density neutrophils present the classical neutrophil morphology with a lobulated nucleus and many granules.

In the upper part of the gradient, the less dense peripheral blood mononuclear cells PBMC are separated. Among these cells, low-density neutrophils can be found.

They comprise immature and mature neutrophils with immunosuppressive properties. In humans the problem is more complex, since the Ly6G antigen does not exist.

MDSC share all these markers, making it impossible to differentiate these cells from mature neutrophils. At present, it is not possible to distinguish whether MDSC are indeed subpopulations of neutrophils or a separate cell type Solito et al. An interesting subpopulation of neutrophils is the so-called low-density neutrophils LDNs.

In contrast, LDNs are found in the low-density fraction Sagiv et al. Interestingly, the proportion of LDNs in the low-density fraction increases with tumor growth and progression Mishalian et al.

These LDNs are a subpopulation of neutrophils with characteristics and functions not well described. Although, LDNs were first reported in the blood of patients with SLE, rheumatoid arthritis, or rheumatic fever Hacbarth and Kajdacsy-Balla, , they attracted attention only recently because they seem to be associated with cancer Brandau et al.

These LDNs have been found in many other pathological conditions including sepsis Morisaki et al. In addition, LDNs have been reported in natural pregnancy Ssemaganda et al.

During pregnancy, downregulation of T cell functions is required to ensure materno-fetal tolerance. Arginase activity is significantly increased in the peripheral blood of pregnant women and also in term placentae Kropf et al. This phenotype is suggestive of a activated neutrophil Fortunati et al. Thus, situations of chronic inflammation and immunosuppression appear to induce neutrophil diversity.

Very little is known about the function of these subpopulations of neutrophils. These NETs could then assist the clustering and growth of free tumor cells disseminated in the abdomen. The origin of LDNs remains unclear.

Since LDNs are a mixture of cells with segmented or banded nuclei and myelocyte-like cells, one thought is that LDNs are immature neutrophils that are released from the bone marrow during chronic inflammation or immunosuppression Denny et al. Another possibility is that these LDNs are activated neutrophils that have undergone degranulation and therefore they have a reduced density Rocha et al.

In mouse models of cancer, these LDNs seem to derive either from immature cells released by the bone marrow Youn et al.

It is important to notice that these LDNs are still not properly characterized. The immunosuppressive function of these cells has not been directly determined and the transition of mature normal density to LDNs appears to involve an increase in volume rather than degranulation Sagiv et al. Thus, most likely these LDNs are not activated normal neutrophils.

In addition, since SLE patients have chronic inflammation, it is unlikely that their LDNs present immunosuppressive activity. All these possibilities need to be further studied in the future. However, one serious limitation for the characterization of these cells is that there are not specific molecular markers that could distinguish among these possible neutrophils subpopulations.

The phenotypic changes of circulating neutrophils during tumor progression are also related to infiltration of neutrophils into tumors. Figure 5. Neutrophils in the circulation display different phenotypes. Mature normal neutrophils PMN leave the bone marrow and display the classical pro-inflammatory and anti-tumor properties of these cells. It is thought that these PMN can migrate into tumors and display an anti-tumor N1 phenotype.

In tumor-bearing mice, immature neutrophils, such as band cells, also leave the bone marrow into the circulation. These cells can infiltrate tumors and display pro-tumor N2 phenotype. The exact origin of recruited neutrophils is not known. Also, it is not clear if N1 cells can change into N2 cells and vice versa under the influence of the tumor microenvironment. Depending on the phenotype displayed by TANs in tumor-bearing mice, they have been classified as N1 or N2 Fridlender et al.

This classification is analogous to antitumor tumor-infiltrating macrophages M1 or protumor macrophages M2 Galdiero et al. Murine N1 TANs are proinflammatory and antitumorigenic. This means that TANs can display an antitumor N1 phenotype or a pro-tumor N2 phenotype depending on the tumor microenvironment Sionov et al.

In addition, in tumor-bearing animals the LDNs increased progressively in circulation, were not cytotoxic, and had reduced expression of cytokines Sagiv et al. This is consistent with the idea that some of the LDNs are indeed immature neutrophils Sagiv et al. It is important to emphasize here that the paradigm of N1 and N2 TANs has been described only in murine models of cancer, and that the nature and function of TANs in the tumor microenvironment remains largely unknown, particularly with human tumors.

There are only two reports on isolation of human TANs, but they describe controversial data on their immunosuppression capacity. In one report, TANs were isolated from digested human lung tumors Eruslanov et al. In another report, TANs were isolated from a colorectal tumor and found to present a typical neutrophil morphology.

Latter, the cells acquire an N2 phenotype and become immunosuppressive Wu et al. Recently, another important role of neutrophils in anticancer therapy has been described. Immunogenic cell death can stimulate neutrophils to utilize cytotoxicity against residual live cancer cells after therapy. The concept of immunosurveillance explains how only the most immunoevasive or highly mutagenic neoplastic cells are able to generate clinically relevant tumors Dunn et al.

Yet, the immune system is capable of recognizing altered-self molecules in damaged cells through endogenously derived danger signals or alarmins Bianchi, ; Garg et al. Anti-cancer therapy-induced cancer cell death can be subdivided into three distinct types i. The details of these types of cell death are beyond the scope of the present publication, but the reader is directed to some excellent recent reviews Green et al.

Through immunogenic cell death, after some types of anti-cancer therapy Galluzzi et al. These signals trigger neutrophil phagocytosis of cancer cells and pro-inflammatory stimulation, leading to a change in neutrophil phenotype Garg et al. As a result, neutrophils stimulated by immunogenic cell death showed cytotoxicity against residual live cancer cells Garg et al. Thus, neutrophils interacting with immunogenic apoptotic cells gain a pro-inflammatory profile, culminating into neutrophil dependent cytotoxicity against residual cancer cells.

The realization that neutrophils do in fact perform many more functions than just antimicrobial responses, and the fact that neutrophils with different phenotypes have been reported in various tissues and pathological conditions, suggest that indeed different neutrophils exist.

However, in most reports the evidence is circumstantial and we are in need for solid experimental proof that the cells described are in fact novel neutrophil subsets. What we have learned for sure so far is that in various pathological conditions, particularly cancer, distinct populations of mature and immature neutrophils are found in circulation. However, we have to highlight that the actual cell type responsible for the immunosuppressive properties of MDSC remains a mystery.

Many questions remain open, but at least two topics seem to be relevant at the moment. One important topic that needs to be addressed is whether mature neutrophils in circulation can be reprogrammed by external stimuli, or whether defined phenotypes are programmed in the bone marrow and neutrophils exit with a particular phenotypic signature.

Evidence suggests that neutrophils are very plastic cells and in consequence the various subtypes described seem to be acquired in the tissues. However, these possibilities need to be formally tested. Another relevant topic is that the many functions described have not been assigned to particular phenotypes of neutrophils.

This remains a complex issue, as there are currently no appropriate molecular markers to readily identify these different neutrophil subpopulations. This confusing scenario is the fuel for new and even more exciting research. We expect to learn new tricks from our favorite cell type in the near future.

The author confirms being the sole contributor of this work and approved it for publication. The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Eileen Uribe-Querol for preparing the figures. Aaes, T. Vaccination with necroptotic cancer cells induces efficient anti-tumor immunity. Cell Rep. Adrover, J. Aging: a temporal dimension for neutrophils. Trends Immunol. Beauvillain, C. CCR7 is involved in the migration of neutrophils to lymph nodes. Blood , — Berger-Achituv, S. A proposed role for neutrophil extracellular traps in cancer immunoediting. Beyrau, M.

Neutrophil heterogeneity in health and disease: a revitalized avenue in inflammation and immunity. Open Biol. Bezu, L. Combinatorial strategies for the induction of immunogenic cell death. Bianchi, M. Bonaventura, A. The pathophysiological role of neutrophil extracellular traps in inflammatory diseases. Borregaard, N. Neutrophils, from marrow to microbes. Immunity 33, — Bournazou, I.

Apoptotic human cells inhibit migration of granulocytes via release of lactoferrin. Isolation of mononuclear cells and granulocytes from human blood. Isolation of monuclear cells by one centrifugation, and of granulocytes by combining centrifugation and sedimentation at 1 g.

Scand, J. PubMed Abstract Google Scholar. Braganza, J. Chronic pancreatitis. Lancet , — Brandau, S. The kinship of neutrophils and granulocytic myeloid-derived suppressor cells in cancer: cousins, siblings or twins? Cancer Biol. Myeloid-derived suppressor cells in the peripheral blood of cancer patients contain a subset of immature neutrophils with impaired migratory properties. Branzk, N. Neutrophils sense microbe size and selectively release neutrophil extracellular traps in response to large pathogens.

Bratton, D. Neutrophil clearance: when the party is over, clean-up begins. Brill, A. Neutrophil extracellular traps promote deep vein thrombosis in mice. Buckley, C. Identification of a phenotypically and functionally distinct population of long-lived neutrophils in a model of reverse endothelial migration. Busso, N. Microcrystals as DAMPs and their role in joint inflammation.

Rheumatology 51, — Carmona-Rivera, C. Low-density granulocytes: a distinct class of neutrophils in systemic autoimmunity. Caruso, R. Prognostic value of intratumoral neutrophils in advanced gastric carcinoma in a high-risk area in northern Italy.

Casanova-Acebes, M. Rhythmic modulation of the hematopoietic niche through neutrophil clearance. Cell , — Cerutti, A. The B cell helper side of neutrophils. Chavakis, E. Novel aspects in the regulation of the leukocyte adhesion cascade. Chen, F. Neutrophils prime a long-lived effector macrophage phenotype that mediates accelerated helminth expulsion. Chistiakov, D. Neutrophil's weapons in atherosclerosis.

Clark, S. Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood. Clarke, T. Recognition of peptidoglycan from the microbiota by Nod1 enhances systemic innate immunity.

Cloke, T. Neutrophilic granulocytes or polymorphonuclear neutrophils PMNs are the most abundant white blood cell in humans and mice. They are characterised by the multi-lobed shape of their nucleus Figure 1, left which distinguished them from other white blood cells of lymphoid or myeloid origin, such as lymphocytes and monocytes.

Neutrophils are the first white blood cells recruited to sites of acute inflammation, in response to chemotactic cues such as CXCL8 interleukin-8, IL-8 produced by stressed tissue cells and tissue-resident immune cells such as macrophages.

Neutrophils therefore comprise a large proportion of the early cellular infiltrate in inflamed tissues and are the major constituent of pus. Phagocytosis is an active, receptor mediated process during which a pathogen is internalised into a specialised vacuole, the phagosome Figure 1, right. The interaction with the pathogen can be direct, through recognition of pathogen-associated molecular patterns PAMPs by neutrophil pattern recognition receptors PRRs , or indirect, through recognition of opsonised microbes by Fc receptors or complement receptors.

The phagosome undergoes a rapid maturation process that involves the fusion with neutrophil granules and the targeted delivery of antimicrobial molecules and generation of reactive oxygen species ROS. Degranulation of specific granules on the neutrophil surface and the extrusion of nucleic acids to form neutrophil extracellular traps NETs create an antimicrobial milieu at the inflammatory site and contributes to killing of extracellular pathogens. Neutrophils at the interface between innate and adaptive immunity Neutrophils have historically been viewed as short-lived effector cells of the innate immune system as they undergo spontaneous apoptosis in vitro unless rescued by survival signals such as inflammatory cytokines or microbial compounds Figure 1, left.

Tonks, N. Redox redux: revisiting PTPs and the control of cell signaling. Cell , — Steinberg, B. STKE pe11 Buchanan, J. DNase expression allows the pathogen group A Streptococcus to escape killing in neutrophil extracellular traps. Beiter, K.

An endonuclease allows Streptococcus pneumoniae to escape from neutrophil extracellular traps. Wartha, F. Capsule and D -alanylated lipoteichoic acids protect Streptococcus pneumoniae against neutrophil extracellular traps. Urban, C. Neutrophil extracellular traps capture and kill Candida albicans yeast and hyphal forms. Hirsch, J. Bactericidal action of histone. Cho, J. Cathepsin D produces antimicrobial peptide parasin I from histone H2A in the skin mucosa of fish.

Kim, H. Pepsin-mediated processing of the cytoplasmic histone H2A to strong antimicrobial peptide buforin I. Park, C. Structure—activity analysis of buforin II, a histone H2A-derived antimicrobial peptide: the proline hinge is responsible for the cell-penetrating ability of buforin II. Natl Acad. USA 97 , — Patat, S.

Antimicrobial activity of histones from hemocytes of the Pacific white shrimp. Sumby, P. Extracellular deoxyribonuclease made by group A Streptococcus assists pathogenesis by enhancing evasion of the innate immune response. USA , — Gupta, A. Induction of neutrophil extracellular DNA lattices by placental microparticles and IL-8 and their presence in preeclampsia. Alghamdi, A.

Seminal DNase frees spermatozoa entangled in neutrophil extracellular traps. Lippolis, J. Neutrophil extracellular trap formation by bovine neutrophils is not inhibited by milk. Palic, D. Zebrafish Danio rerio whole kidney assays to measure neutrophil extracellular trap release and degranulation of primary granules.

Fairhurst, A. Systemic lupus erythematosus: multiple immunological phenotypes in a complex genetic disease. Duranton, J. FEBS Lett. McCauley, T. Purification and characterization of fertility-associated antigen FAA in bovine seminal fluid. Zhong, X. Elevation of both maternal and fetal extracellular circulating deoxyribonucleic acid concentrations in the plasma of pregnant women with preeclampsia.