Knochenmarksstammzellen werden als mögliche Zellquelle zur Verbesserung kardialer Funktion nach Myokardinfarkt angesehen. Um die Rolle und das Potential verschiedener Knochenmarkszellpopulationen auf das linksventrikuläre Remodeling nach Myokardinfarkt weiter zu untersuchen, wurde auf das Maus-Infarkt-Modell zurückgegriffen. Nach experimentellem Myokardinfarkt durch Ligation der vorderen absteigenden Koronararterie erfolgte entweder die intramyokardiale Injektion von unfraktionierten Knochenmarkszellen oder einer mit Vorläufer- (Lin-) bzw. reifen (Lin+) Zellen angereicherten Knochenmarkszellsubpopulation. Obgleich mit keiner Zellpopulation entscheidend Einfluss auf Überlebensrate und Infarktgröße genommen werden konnte, zeigte sich eine signifikante Verbesserung des linksventrikulären Remodelings nach Injektion von unfraktionierten Knochenmarkszellen, welche hingegen durch Behandlung mit Lin- oder Lin+ Zellen ausblieb. Gemessen wurde dies einerseits auf molekularer Ebene, wo der linksventrikuläre Hypertrophiemarker, bestehend aus betaMHC/alphaMHC-Ratio signifikant gesenkt werden konnte, andererseits auf echokardiographischer Ebene, wo sich eine signifikante Verminderung linksventrikulärer Dilatation nachweisen ließ. Da sich die untersuchten Zellpopulationen hinsichtlich in vitro gemessener Zytokinexpressionslevel teilweise erheblich unterschieden, müssen die beobachteten Resultate im Zusammenhang mit stattgefundener parakrine Zytokinsekretion gesehen werden. / Bone marrow stem cells are considered as a promising cell source to improve cardiac function after myocardial infarction. To further investigate the role and potential of different bone marrow cell preparations on left ventricular remodelling we employed the mouse-infarct-model. After induction of myocardial infarction through ligation of the left descending coronary artery, mice were treated either with intramyocardial injection of unfractionated bone marrow cells, progenitor-enriched (Lin-) or mature (Lin+) cells.Although none of the cell populations showed a pronounced influence on survival rate or infarct size, there was a significant amelioration of left ventricular remodelling after injection of unfractionated bone marrow cells, which could not be accomplished by treatment with Lin- or Lin+ cells. This effect was measured both on molecular basis, where the left ventricular hypertrophie marker, consisting of the betaMHC/alphaMHC-ratio was significantly decreased, and on echocardiographic basis, where a significant reduction of left ventricular dilatation was demonstrated. Considering the substantial differences in in-vitro measured cytokine-levels observed in the investigated cell populations, the results shown could potentially be attributed to paracrine secretion of cytokines.

Thesis advisor: Thomas C. Chiles / Elevating intracellular cAMP levels can result in a wide range of anti-inflammatory effects and growth arrest and apoptosis in cancer cells, marking phosphodiesterases (PDEs) as potential targets for inflammatory diseases and cancer treatment. PDE7A is proposed to be a new therapeutic target for its ubiquitous expression in proinflammatory and immune cells. A Barbituric acid based compound, BC12 was identified as an in vitro PDE7 inhibitor in fission-yeast-based high-throughput screen. Analysis of this compound on the activation of Jurkat T lymphocytes, mouse and human primary T cells reveals inhibition of IL-2 production, though cell viability is not significantly impacted. Real-time RT-PCR and mRNA stability assays indicate that the inhibition of IL-2 production by BC12 is attributable to transcriptional repression without accelerating IL-2 mRNA decay. By contrast, compounds of similar structure with that of BC12 exhibit varying effects on IL-2 production that does not correlate with their in vitro PDE7 inhibitory activity, suggesting that the in vivo target of BC12 responsible for these effects may not be PDE7. Our study further reveals that BC12 inhibits IL-2 transcription through targeting NFAT and NFkB-mediated pathways. Preliminary investigation on other T helper cell cytokine secretion indicates that BC12 has a potential to selectively inhibit Th2 cytokines. Our data suggest that BC12 may present a novel anti-inflammatory drug for its immunosuppressive and potential immunomodulatory effects. Analysis of BC12 on a human B-cell line and a monocytic cell line demonstrate its pro-apoptotic effects in a dose-dependent manner. Titration of BC12 on human diffuse large B-cell lymphoma (DLBCL), LY18 cells, and human primary B cells reveals that BC12 induces cell death more effectively in DLBCL LY18 cells than normal B cells, suggesting the anti-cancer potential of this compound. / Thesis (PhD) — Boston College, 2013. / Submitted to: Boston College. Graduate School of Arts and Sciences. / Discipline: Biology.

Sabe-se que a desnutrição modifica a resposta imune específica e inespecífica do organismo frente a agentes infecciosos comprometendo a produção e a função de células linfo-hemopoéticas, estando associada a modificações da resposta imune, resultando em maior suscetibilidade à infecções, porém os mecanismos exatos que comprometem o sistema imune em estados de desnutrição ainda estão para serem esclarecidos. A literatura relata que aproximadamente 60% das infecções que evoluem para sepse são adquiridas no ambiente hospitalar, envolvendo geralmente bactérias Gram negativas e incidindo especialmente em indivíduos com nutrição inadequada. Considerando tais aspectos e em função da complexidade da interação do estado nutricional e resposta do organismo frente a agentes patogênicos, envolvendo controles celulares e moleculares múltiplos ainda pouco conhecidos, propusemo-nos a estudar alguns aspectos da resposta inflamatória em desnutrição. Camundongos Swiss machos adultos, submetidos a desnutrição protéica-energética, após perda de aproximadamente 25% do peso corpóreo, foram inoculados com lipopolissacarideo de Escherichia coli: Hemograma, mielograma, esplenograma e dosagens sistêmicas de citocinas, foram realizadas. Células coletadas da cavidade peritonial de animais que não foram estimulados com LPS foram utilizadas para as determinações de citocinas e NO in vitro, bem como para quantificação dos receptores CD14 e TLR-4/MD-2 e do fator de transcrição NFκB. Células da cavidade peritonial foram usadas, também, para realização dos testes de espraiamento, fagocitose e atividade cida com Candida albicans. Animais desnutridos apresentaram anemia, leucopenia; severa redução na celularidade da medula óssea, do baço e da cavidade peritonial. A capacidade de espraiamento, fagocitose, atividade cida e síntese de citocinas e NO foram significativamente menores nos animais dos grupo desnutrido. O número de receptores de CD14 e TLR-4/MD-2 e do fator de transrição NFκB, também foram significativamente menores nos animais desnutridos. Estes achados sugerem que. animais desnutridos apresentam resposta deficiente frente ao LPS. A menor expressão de receptores CD14 e TLR-4/MD-2 pode ser responsável, em parte pela imunodepressão observada. Os dados nos levam a inferir que o estado nutricional interfere no estado de ativação de macrófagos e na capacidade de resposta dos animais. / Malnutrition modifies the specific and non-specific immune response of the organism to infectious agents, hampering the production and function of Iympho-hemopoietic cells leading to a higher susceptibility of the orgonism to infections. However, the exact mechanisms by which the immune system is undermined has not yet been fully elucidated. Approximately 60% of infections that evolve to bacteremia are nosocomial, and usually involve Gram negative bacteria in individuais that have inadequate nutrition. Taking this into consideration, and in view of the complexity of the interaction between nutritional state and the organism\' s response to infection, which involves poorly known multiple cellular and molecular controls, we proposed to study a few aspects of the inflammatory response in protein malnutrition. Male, outbred Swiss mice were sumbitted to protein malnutrition and after the loss of about 25% of total body weight they were inoculad whith lipopolissacharide of Escherichia coli: Hemogram, mielogram, splenogram and the determination of systemic production of cytokines were used to evaluate. Cells from the peritoneal cavity of animais who were not inoculated were collected for determination of cytokines and NO production in vitro and to evaluate the expression of NFκB and CD14 and TLR-4/MD-2 receptors. Cells from the peritoneal cavity were used too for the spreading, phagocytosis and killing tests with Candida albicans. Malnourished animais presented anemia, leucopenia a severe reduction on bone marrow, spleen and peritoneal cavity cellularity. The spreading, phagocytosis, killing and the production of cytokines and NO was significantly lower in malnourished animais. The number of CD14 and TLR-4/MD-2 receptors and NFκB was found to be significantly lower in malnourished animais. These findings suggest that malnourished mice present a deficient response to LPS. The smaller expression of CD14 and TLR-4/MD-2 receptors may be partly responsible for the immunodeficiency observed. These data lead us to infer that the nutritional state interferes in the activation of macrophages and in the immune response capacity.

FAM3B/PANDER, é uma nova proteína tipo citocina pertencente a família FAM3. Foram reveladas algumas similaridades estruturais entre o FAM3B e outras citocinas associadas ao câncer, como IL-6 e FAM3C suportando a hipótese, de que FAM3B também poderia estar envolvido na progressão tumoral. Considerando que o FAM3B é expresso por tumores de mama, este trabalho dedicou-se a elucidar suas possíveis funções em tumores, a partir de um modelo de superexpressão na linhagem celular tumoral da mama, MDA-MB-231. Ensaios celulares e moleculares sugeriram que o FAM3B é capaz de conferir, proteção à morte celular e de aumentar taxa de migração nas células MDA-MB-231 via, principalmente, Bcl-2 e Bcl-xL. Em concordância, os resultados in vivo demonstraram aumento do volume e peso tumoral, com aumento da expressão de Bcl-2 nos tumores dos animais inoculados com as células MDA-MB-231-FAM3B. Estes resultados indicam que esta proteína, exerce funções nas etapas de invasão e metástase em tumores de mama, o que permite considerar o FAM3B, como um possível candidato a marcador tumoral. / FAM3B/PANDER is a new cytokine-like protein, member of the new FAM3 family. Recent data has been shown similarities between FAM3B/PANDER and others citokines involved in tumor progression, like IL-6 and FAM3C/ILEI, supporting the hypothesis that FAM3B is also involved in tumor progression. In Silico data revealed that FAM3B is expressed by breast tumors and using the overexpression of FAM3B in MDA-MB-231 tumor cell line, the aim was evaluated possible roles of FAM3B in breast tumor progression. The results revealed that FAM3B overexpression inhibits cell death and promotes cell migration in MDA-MB-231 cell line, through, at least in parts, Bcl-2 and Bcl-xL pathways. In agreement, in vivo results shown an increase in volume, weight tumors and increase expression of Bcl-2 in mice with cells overexpressing FAM3B.These results suggest important functions of this protein in cell invasiveness and migration, which allows us to consider, FAM3B as a possible future candidate as a molecular biomarker.

PANDER/FAM3B (PANcreatic-DERived factor) é uma citocina capaz de induzir apoptose em células-β secretoras de insulina e regular a homeostase da glicose nos tecidos periféricos. Considerando que PANDER/FAM3B também é expresso em tumores de próstata, o presente trabalho avaliou o papel desta citocina na inibição da apoptose in vitro, assim como o crescimento e invasividade tumoral de células de carcinoma de próstata da linhagem DU145 in vivo, usando o modelo de superexpressão estável mediada por lentivirus. Os nossos resultados apontam um aumento da viabilidade e uma diminuição da morte celular em células que superexpressam PANDER quando comparadas ao grupo controle. Este efeito protetor é acompanhado por um aumento da expressão de genes anti-apoptoticos e uma diminuição da atividade proteolítica das caspases. Por outro lado, a superexpressão de PANDER/FAM3B por tumores in vivo se correlaciona com aumento da massa tumoral e o aumento de vasos sanguíneos nos tumores. Em síntese, nossos dados demonstram que, em contraste ao papel observado em células β-pancreáticas, o PANDER/FAM3B inibe morte celular e promove a tumorigenicidade e o crescimento tumoral in vivo, sugerindo ao mesmo tempo, algum envolvimento com angiogênese e metástase em células DU145. / PANDER / FAM3B (pancreatic-derived factor) is a cytokine capable of inducing apoptosis in secreting β-cells and insulin to regulate glucose homeostasis in peripheral tissues. Whereas PANDER / FAM3B is also expressed in prostate tumors, this study evaluated the role of this cytokine in the inhibition of apoptosis in vitro as well as tumor growth and invasiveness of prostate carcinoma DU145 cells lineage in vivo, using the model Stable overexpression mediated by lentivirus. Our results suggest that increased viability and decreased cell death in cells that overexpress PANDER when compared to the control group. This protective effect is accompanied by an increased expression of anti-apoptotic genes and a decrease in proteolytic activity of caspases. Moreover, overexpression of PANDER / FAM3B by tumors in vivo correlates with increased tumor mass and the increase of blood vessels in tumors. In summary, our data demonstrate that in contrast to the role observed in pancreatic cells, PANDER / FAM3B inhibits cell death and promotes tumorigenicity and tumor growth in vivo, suggesting at the same time, some involvement in angiogenesis and metastasis in cell DU145.

A desnutrição modifica a resistência à infecção, alterando grande variedade de processos fisiológicos, incluindo a hematopoiese e a resposta imune. Os mecanismos exatos que comprometem o sistema imunológico em condições de desnutrição ainda não estão totalmente compreendidos, sendo que a desnutrição modifica a funcionalidade das células efetoras na resposta inflamatória/infecciosa, causando redução da síntese de citocinas pró-inflamatórias, além de alterações na hematopoiese. Os linfócitos têm papel-chave tanto na resposta imune inata quanto adaptativa. Nesse contexto, propusemo-nos a avaliar alguns aspectos da resposta linfoide esplênica em um modelo de desnutrição proteica. Camundongos BALB/c, machos, submetidos à desnutrição proteica, após perda de aproximadamene 20% do peso corpóreo, foram eutanasiados. Hemograma, mielograma, esplenograma e análise das concentrações séricas de proteína totais, albumina, pré-albumina e dosagem de imunoglobulinas G e M foram realizados. As células do timo e do baço foram coletadas e analisadas por citometria de fluxo - e foi realizado o ciclo celular. Para avaliação da linfoproliferação, as células esplênicas foram cultivadas, in vitro, e estimuladas com LPS ou ConA durante 72 horas. Foram quantificadas a capacidade de produção de INF-γ, IL-2 e IL-10 e a expressão de STAT- 1 e STAT-3. Os animais desnutridos apresentaram anemia, leucopenia e severa redução na celulariedade da medula óssea, com comprometimento no setor mieloide e diminuição da celulariedade do baço, com redução da população de linfócitos. Os animais desnutridos apresentaram alterações no desenvolvimento celular linfoide, alteração na capacidade de proliferação, menor produção de citocinas pró-inflamatórias, como IL-2, e, ao mesmo tempo, aumento da produção de IL-10. Os animais também apresentaram maior porcentagem de células CD3+ e CD4+ no baço em relação aos animais controle. Quando as células esplênicas foram estimuladas, apresentaram maior expressão de STAT-3 e menor expressão de STAT-1. Baseando-se nestes resultados, discutimos se deficiências nutricionais alteram a imunocompetência e aumentam o risco de infecções. / Malnutrition modifies the resistance against infection by changing a variety of physiological processes, including the hematopoiesis and the immune response. The exact mechanisms that compromise the immune system in conditions of malnutrition is not yet thoroughly understood, since malnutrition modifies the functionality of the effector cells in inflammatory / infectious response, causing the reduction of the synthesis of proinflammatory cytokines, besides causing alterations in hematopoiesis. The lymphocytes have a key role in both innate and adaptive immune responses, in this context, we proposed to evaluate some aspects of lymphoid spleen cells response in a model of protein malnutrition. Male mice BALB / c submitted to protein malnutrition after approximately 20% loss of body weight were euthanized. The complete blood count, bone marrow examination, spleen cells count and analysis of serum total protein, albumin, pre-albumin and immunoglobulin G and M were performed. The thymus and spleen cells were collected and analyzed by flow cytometry as well as the cell cycle performed. For evaluation of the lymphocyte proliferation the spleen cells were cultivated in vitro and stimulated with LPS or ConA for 72 hours and the ability to produce IFN-γ, IL-2 and IL-10 as well as the expression of STAT-1 and STAT -3, were quantified. The malnourished animals presented anemia, leukopenia and severe reduction in celulariedade bone marrow with myeloid commitment in the sector and decreased celulariedade spleen, with reduced lymphocyte population. The malnourished animals showed changes in lymphoid cell development, abnormal proliferation capacity, reduced production of proinflammatory cytokines such as IL-2, and at the same time, increased production of IL-10. The animals also had higher percentages of CD3 + and CD4 + spleen compared to control animals. When spleen cells were stimulated, showed higher expression of STAT-3 and lower expression of STAT-1. Based on these results, we discussed whether nutritional deficiencies alter immunocompetence and increase the risk of infections.

Elevated levels of the inflammatory cytokine interleukin-6, IL-6, have been linked with poor prognosis in ovarian cancer patients by influencing tumour growth, invasion, angiogenesis and chemo-resistance. A clinical trial conducted in parallel with pre-clinical studies showed an anti-IL-6 antibody to have some activity in ovarian cancer patients and in xenograft models, via reduction in pro-inflammatory and angiogenic factors such as TNF-α, IL-8 and VEGF. Anti-IL-6 treatment also showed significant reductions in vascular area with decreased expression of an angiogenic factor Jagged1. The aim of my study was to investigate the effects of IL-6 on normal and tumour angiogenesis. I found that recombinant IL-6 stimulates angiogenesis in mouse and rat aortic ring assays and that it can also stimulate growth and migration of endothelial cells in vitro. IL-6 has similar potency as VEGF in inducing vessel sprouting. IL-6 itself does not induce VEGF in the endothelial cells I tested. Investigation of the effects of IL-6 on vessel maturation revealed a significant reduction in pericyte coverage of vessels treated with IL-6 compared with VEGF. Collectively, these data led to my hypothesis that ‘IL-6 drives aberrant angiogenesis, independent of VEGF signalling’. Investigating the mechanism by which IL-6 drives angiogenesis and leads to defective pericyte formation, I showed a link between IL-6 and the Notch ligands, Jagged1 and DLL4. My data suggested that IL-6 could stimulate Jagged1 in endothelial cells, whereas VEGF induces DLL4, the Notch ligand known to be involved in inducing stalk phenotype. Exploring previous findings to get a better understanding of the interaction of Notch ligands and pericyte recruitment also suggested a role of Angiopoeitin-2 in relation to IL-6 signalling. These observations were extended in IGROV-1 ovarian cancer xenografts treated with an anti-IL-6 antibody and by analysis of gene expression datasets from ovarian cancer biopsies. My results suggest therapeutic potential of combining inhibitors of IL-6 and VEGF in ovarian cancer.

The purpose of this study was to quantify the effects of phacoemulsification and posterior segment chamber lens placement on vitreous inflammatory and neovascular growth factors. More specifically, the effect of immediately preceding cataract surgery was compared to a history of cataract surgery. This study involved a retrospective review and analysis of vitreous samples from a total of twenty seven patients separated into three groups. Group 1, seven patients who underwent a pars plana vitrectomy with macular surgery, group 2, fourteen patients who underwent a combined cataracts and pars plana vitrectomy procedure and group 3, six patients with a history of cataract surgery who underwent a pars plana vitrectomy. The twenty seven patients were picked from a pool of 100 patients who all received pars plana vitrectomy at Beth Israel Deaconess Medical Center with surgeon Dr. Jorge Arroyo. Exclusion factors included active ocular pathologies such as vitreous hemorrhage and retinal detachment. Undiluted vitreous samples from each group were taken before beginning the pars plana vitrectomy. The vitreous samples were analyzed for concentrations of fourteen specific vitreous cytokines and neovascular growth factors including but not limited to TNF Alpha; and SCD40L. These fourteen cytokines and growth factors were chosen through a literature review on the post-surgery ocular inflammatory response. Statistical analysis was done on the average means of the cytokine levels for each group using SPSS 20 for windows. A comparison of means analysis found no significant difference in the means of the fourteen cytokines for group 1 and group 2. A second comparison of means with a pooled control group of both group 1 and group 2 patients versus group 3 was also run. In this analysis, only SCD40L or soluble CD40 ligand was shown to have a significant difference between groups. SCD40L levels were significantly higher (significance level of .038) in group 3, the history of cataract group with a mean of (9.50±4.76) than in the control group with a mean of (5.50±3.35). The findings of this study indicate that the protein SCD40L may play an important role in mediating the inflammatory response seen post cataract surgery and may be useful as a target for novel therapies against the inflammatory response.

O aumento da adiposidade corpórea pode gerar diversos mediadores inflamatórios com capacidade de influenciar a proliferação e a diferenciação hematopoética e, consequentemente, a complexa regulação da hematopoese. Por isso, propusemo-nos, neste trabalho, avaliar a influência do aumento da adiposidade corpórea sobre a proliferação e a diferenciação de células hematopoéticas, bem como sua capacidade em sintetizar citocinas. Ratos Wistar, machos foram alimentados com uma dieta rica em lipídios durante 14 semanas. Após esse período foram avaliados hemograma, mielograma, perfil lipídico, concentrações séricas de leptina, insulina e adiponectina. Citômetria de fluxo foi utilizada para avaliação da porcentagem de células CD34+/CD133+, bem como o ciclo celular de células medulares. Células medulares foram utilizadas para avaliar a atividade proliferativa in vitro e a capacidade de diferenciação, in vitro, na presença de IL-3, EPO, GM-CSF e G-CSF. Animais, alimentados com dieta hiperlipídica, apresentaram maiores concentrações de leptina circulante, com aumento de gordura corporal, aumento da concetração de proteína C reativa, colesterol total, LDL, VLDL e triacilglicerol. O hemograma apresentou neutrofilia absoluta e a medula óssea apresentou-se hipercelular com aumento do número de granulócitos maduros e da população celular CD133-/CD34+. Os resultados dos testes in vitro demonstraram aumento da capacidade de síntese de IL-3 e aumento de G-CSF, com aumento do potencial proliferativo, também evidenciado pelo maior número de células medulares na fase S/G2/M, bem como o aumento da diferenciação granulocítica. Esses resultados sugerem que a leucocitose e neutrofilia observadas em situações de aumento da adiposidade corpórea são decorrentes de uma complexa modulação do sistema hematopoético. / The body fat increase can generate various inflammatory mediators, that are capable to influence the proliferation and differentiation of hematopoietic cells and consequently modulate the complex regulation of the hematopoiesis. In this study we have proposed to evaluate the effect of increase body fat on the proliferation and differentiation of hematopoietic cells, as well as its ability to synthesize cytokines. Male Wistar rats were subjected to a high fat diet during a period of 14 weeks. After that period were evaluated hemogram, mielogram, lipid profile and the serum concentrations of leptin, insulin and adiponectin. Flow cytometry was used to evaluate the percentage of CD34+/CD133+, as well as the cell cycle of bone marrow cells. Bone marrow cells were used to perform the proliferation and differentiation capacity in vitro in the presence of IL-3, EPO, GM-CSF and G-CSF. Animals fed high-fat diet had higher concentrations of circulating leptin with increase body fat, and increase of C-reactive protein, total cholesterol, LDL, VLDL and triacylglycerol concentrations. The hemogram showed absolute neutrophilia and a hypercellular bone marrow with increase of granulocytic mature population and CD133-/CD34+ cells. The results in vitro, showed an increase of IL-3 and G-CSF production, and higher proliferative potential with an increase in S/G2/M bone marrow cell cycle phases, as well as an increase of the granulocytic differentiation. The results suggest that leukocytosis and neutrophilia observed in this model of body fat increase are in fact a result of a complex modulation of the hematopoietic system.

Immune cells must accurately interpret environmental signals to make robust cell fate decisions and control inflammatory signalling. Many signals (e.g. Tumor Necrosis Factor alpha (TNFα) or interferon gamma (IFNγ)) converge on just a few key signalling systems such as Nuclear Factor kappa B (NF-κB) or Signal Transducers and Activators of Transcription (STAT), which exhibit complex activation dynamics that control patterns of downstream gene expression. Often, seemingly identical cells show heterogeneous or random behaviour to a common stimulus. Therefore, a key question is how can immune cells coordinate inflammatory signalling in the presence of this noise. The NF-kB system dynamics were studied in response to rapidly changing inflammatory signals. It was shown that pulsed TNFa cytokine stimulations induced digital single-cell NF-kB responses, with only a fraction of cells able to respond to repeated pulses. These responses appeared to be reproducible in individual cells, but heterogeneous in the population. Mathematical models of the NF-kB signalling network suggested that single-cell responses were governed through a refractory state potentially encoded via 'extrinsic' noise in the levels of signalling molecules related to the TNFa signal transduction pathway. Such signal processing enabled robust and reproducible single cell responses and maintained acute tissue-level signalling, with fewer cells responding to shorter pulsing intervals. The NF-kB system is involved in effector cytokine propagation in response to pathogen infection. It was shown that in macrophages, the dose of TLR4 stimulation (mimicking the pathogen infection) was encoded in graded (yet heterogeneous) NF-kB dynamics in single cells. This resulted in analogue inflammatory gene expression patterns in the population. However, individual cells substantially differed in their ability to encode TLR4 signal and to regulate TNFa expression, which was explained by extrinsic noise in the NF-kB system. Quantitative mathematical modelling showed that tissue-level environment modulates heterogeneous single-cell TNFa outputs; by effectively removing it from circulation. This may determine the interaction distance between tissue-resident immune cells to enable propagation of cellular inflammation. Heterogeneity of single cell macrophage signalling was also observed in NF-kB and STAT1 system responses to a range of IFN stimulation doses. Although each system showed substantial variability between cells, their responses were surprisingly well correlated in individual cells. It was however apparent (based on gene expression studies) that individual cells may not be able to precisely discriminate different IFNg doses. Overall, this work suggests that heterogeneity in the NF-kB (and other) regulatory networks might be a part of an inherent design motif in the inflammatory response, which enables robust control of the tissue-level inflammatory response by preventing homogeneous and thus potentially harmful activation.