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Although inflammatory mechanisms driving hepatocellular carcinoma (HCC) have been proposed, the regulators of anticancer immunity in HCC remain poorly understood. We found that IL27 receptor (IL27R) signaling promotes HCC development in vivo. High IL27EBI3 cytokine or IL27RA expression correlated with poor prognosis for patients with HCC. Loss of IL27R suppressed HCC in vivo in two different models of hepatocarcinogenesis. Mechanistically, IL27R sig­naling within the tumor microenvironment restrains the cytotoxicity of innate cytotoxic lymphocytes. IL27R ablation enhanced their accumulation and activation, whereas depletion or functional impairment of innate cytotoxic cells abrogated the effect of IL27R disruption. Pharmacologic neutralization of IL27 signaling increased infiltration of innate cytotoxic lymphocytes with upregulated cytotoxic molecules and reduced HCC development. Our data reveal an unexpected role of IL27R signaling as an immunologic checkpoint regulating innate cytotoxic lymphocytes and promoting HCC of different etiologies, thus indicating a therapeutic potential for IL27 pathway blockade in HCC. Significance: HCC, the most common form of liver cancer, is characterized by a poor survival rate and limited treatment options. The discovery of a novel IL27-dependent mechanism controlling anticancer cytotoxic immune response will pave the road for new treatment options for this devastating disease. This article is highlighted in the In This Issue feature, p. 1825

Supplementary Data from IL27 Signaling Serves as an Immunologic Checkpoint for Innate Cytotoxic Cells to Promote Hepatocellular Carcinoma

Although inflammatory mechanisms driving hepatocellular carcinoma (HCC) have been proposed, the regulators of anticancer immunity in HCC remain poorly understood. We found that IL27 receptor (IL27R) signaling promotes HCC development in vivo. High IL27EBI3 cytokine or IL27RA expression correlated with poor prognosis for patients with HCC. Loss of IL27R suppressed HCC in vivo in two different models of hepatocarcinogenesis. Mechanistically, IL27R sig­naling within the tumor microenvironment restrains the cytotoxicity of innate cytotoxic lymphocytes. IL27R ablation enhanced their accumulation and activation, whereas depletion or functional impairment of innate cytotoxic cells abrogated the effect of IL27R disruption. Pharmacologic neutralization of IL27 signaling increased infiltration of innate cytotoxic lymphocytes with upregulated cytotoxic molecules and reduced HCC development. Our data reveal an unexpected role of IL27R signaling as an immunologic checkpoint regulating innate cytotoxic lymphocytes and promoting HCC of different etiologies, thus indicating a therapeutic potential for IL27 pathway blockade in HCC.Significance:HCC, the most common form of liver cancer, is characterized by a poor survival rate and limited treatment options. The discovery of a novel IL27-dependent mechanism controlling anticancer cytotoxic immune response will pave the road for new treatment options for this devastating disease.This article is highlighted in the In This Issue feature, p. 1825

Colorectal cancer (CRC) is among the most frequent forms of cancer, and new strategies for its prevention and therapy are urgently needed

The transcription factor ThPOK (encoded by the Zbtb7b gene) controls homeostasis and differentiation of mature helper T cells, while opposing their differentiation to CD4+ intraepithelial lymphocytes (IELs) in the intestinal mucosa. Thus CD4 IEL differentiation requires ThPOK transcriptional repression via reactivation of the ThPOK transcriptional silencer element (SilThPOK). In the present study, we describe a new autoregulatory loop whereby ThPOK binds to the SilThPOK to maintain its own long-term expression in CD4 T cells. Disruption of this loop in vivo prevents persistent ThPOK expression, leads to genome-wide changes in chromatin accessibility and derepresses the colonic regulatory T (Treg) cell gene expression signature. This promotes selective differentiation of naive CD4 T cells into GITRloPD-1loCD25lo (Triplelo) Treg cells and conversion to CD4+ IELs in the gut, thereby providing dominant protection from colitis. Hence, the ThPOK autoregulatory loop represents a key mechanism to physiologically control ThPOK expression and T cell differentiation in the gut, with potential therapeutic relevance. The transcription factor ThPOK is critical for homeostasis and differentiation of mature helper T cells. Here, Kappes and colleagues describe a ThPOK-mediated positive autoregulatory loop that is crucial for tissue-specific Treg cell differentiation, maintenance of intestinal Treg cell integrity and conversion of these cells into CD4+ intraepithelial lymphocytes.

Advanced colorectal carcinoma (CRC) is characterized by a high frequency of primary immune evasion and refractoriness to immunotherapy. Given the importance of interferon (IFN)-γ in CRC immunosurveillance, we investigated whether and how acquired IFN-γ resistance in tumor cells would promote tumor growth, and whether IFN-γ sensitivity could be restored.Spontaneous and colitis-associated CRC development was induced in mice with a specific IFN-γ pathway inhibition in intestinal epithelial cells. The influence of IFN-γ pathway gene status and expression on survival was assessed in patients with CRC. The mechanisms underlying IFN-γ resistance were investigated in CRC cell lines.The conditional knockout of the IFN-γ receptor in intestinal epithelial cells enhanced spontaneous and colitis-associated colon tumorigenesis in mice, and the loss of IFN-γ receptor α (IFNγRα) expression by tumor cells predicted poor prognosis in patients with CRC. IFNγRα expression was repressed in human CRC cells through changes in N-glycosylation, which decreased protein stability via proteasome-dependent degradation, inhibiting IFNγR-signaling. Downregulation of the bisecting N-acetylglucosaminyltransferase III (MGAT3) expression was associated with IFN-γ resistance in all IFN-γ-resistant cells, and highly correlated with low IFNγRα expression in CRC tissues. Both ectopic and pharmacological reconstitution of MGAT3 expression with all-trans retinoic acid increased bisecting N-glycosylation, as well as IFNγRα protein stability and signaling.Together, our results demonstrated that tumor-associated changes in N-glycosylation destabilize IFNγRα, causing IFN-γ resistance in CRC. IFN-γ sensitivity could be reestablished through the increase in MGAT3 expression, notably via all-trans retinoic acid treatment, providing new prospects for the treatment of immune-resistant CRC.

Bladder cancer is the fourth most commonly diagnosed malignancy in men worldwide and has one of the highest recurrence rates of all cancers. This cancer type is unique because chronic inflammation caused by Schistosoma haematobium can cause bladder cancer, while inflammation induced by Bacillus Calmette Guerin is the therapeutic cornerstone for this cancer type. Activation of proinflammatory IL- 6/Stat3 axis promotes the development of different cancers by acting on cancer cells as well as by modulating cancer microenvironment. Using a genetic and pharmacological approach in a mouse model, we demonstrated the importance of IL-6 and Stat3 signaling in bladder cancer. Our findings show that pharmacological inhibition of Stat3 with WP1066 effectively delays progression and invasiveness of bladder cancer in N-butyl-N-(4- hydroxybutyl) nitrosamine-induced mouse model. Moreover, either IL-6 blockade or Stat3 inhibition sensitized bladder cancer to anti-PD-L1 immune therapy. Taken together, our study demonstrates an important role of IL-6/Stat3 signaling in bladder cancer and creates a rationale for testing the therapeutic potential of Stat3 inhibitors in human MIBC both alone or in combination with anti-PD-L1 and anti-IL-6 therapy.

A myriad of microbes living together with the host constitute microbiota, which possesses very diverse functions in regulation of host physiology. Recently, it has been unequivocally demonstrated that microbiota regulates cancer initiation, progression and responses to therapy. Here we review known pro-tumorigenic and anti-tumorigenic function of microbiota and mechanisms how microbes can regulate cancer cells and immune and stromal cells within the tumor microenvironment.

γδ T cells physiologically scan the intestinal epithelium, representing a substantial fraction of infiltrating lymphocytes in colorectal cancer (CRC), albeit their role in CRC remains unclear. Using murine CRC models, we found that most γδ T cells in pre- or non-tumor colon express Vγ1+ or Vγ7+ and exhibit a cytotoxic profile. Targeting these γδ T cell subsets, as well as conditionally interfering with γδ T cell function at early stages of tumorigenesis led to heightened tumor development, suggesting anti-CRC functions for Vγ1+ and Vγ7+ subsets. In contrast, RORγt+ γδ T cell subsets, including Vγ4+ and microbiotadependent Vγ6+, accumulated during CRC progression. Conditional deletion of RORγt or Vγ chains revealed redundant roles for IL-17–producing Vγ4+ and Vγ6+ γδ T cells in promoting tumor growth. Our results uncover pro- and anti-tumor roles for γδ T cell subsets.

Paneth cells constitutively produce antimicrobial peptides and growth factors that allow for intestinal homeostasis, host protection, and intestinal stem cell replication. Paneth cells rely heavily on the glycolytic metabolic program, which is in part controlled by the kinase complex Mechanistic target of rapamycin (mTORC1). Yet, little is known about mTOR importance in Paneth cell integrity under steady-state and inflammatory conditions. Our results demonstrate that IFN-γ, a crucial mediator of the intestinal inflammation, acts directly on murine Paneth cells to alter their mitochondrial integrity and membrane potential, resulting in an TORC1-dependent cell death mechanism distinct from canonical cell death pathways including apoptosis, necroptosis, and pyroptosis. These results were established with the purified cytokine and a physiologically relevant common Th1-inducing human parasiteToxoplasma gondii. Given the crucial role for IFN-γ, which is a cytokine frequently associated with the development of inflammatory bowel disease and compromised Paneth cell functions, the identified mechanisms underlying mTORC1-dependent Paneth cell death downstream of IFN-γ may provide promising novel approaches for treating intestinal inflammation.

Myeloid-derived suppressor cells (MDSCs) are major negative regulators of immune responses in cancer and chronic infections. It remains unclear if regulation of MDSC activity in different conditions is controlled by similar mechanisms. We compared MDSCs in mice with cancer and lymphocytic choriomeningitis virus (LCMV) infection. Chronic LCMV infection caused the development of monocytic MDSCs (M-MDSCs) but did not induce polymorphonuclear MDSCs (PMN-MDSCs). In contrast, both MDSC populations were present in cancer models. An acquisition of immune-suppressive activity by PMN-MDSCs in cancer was controlled by IRE1α and ATF6 pathways of the endoplasmic reticulum (ER) stress response. Abrogation of PMN-MDSC activity by blockade of the ER stress response resulted in an increase in tumor-specific immune response and reduced tumor progression. In contrast, the ER stress response was dispensable for suppressive activity of M-MDSCs in cancer and LCMV infection. Acquisition of immune-suppressive activity by M-MDSCs in spleens was mediated by IFN-γ signaling. However, it was dispensable for suppressive activity of M-MDSCs in tumor tissues. Suppressive activity of M-MDSCs in tumors was retained due to the effect of IL-6 present at high concentrations in the tumor site. These results demonstrate disease- and population-specific mechanisms of MDSC accumulation and the need for targeting different pathways to achieve inactivation of these cells.

Hepatocellular carcinoma (HCC) is the most common form of liver cancer with poor survival and limited therapeutic options. HCC development is accompanied by underlying chronic inflammation, which represents a major unifying mechanism for tumor promotion. While some tumor-promoting inflammatory mechanisms had been proposed, the identity of immune mechanisms controlling anti-cancer immunity in HCC remain poorly understood. Interleukin (IL)-27 receptor signaling plays an anti-inflammatory role in a variety of infectious and chronic inflammatory diseases. Here, using genetic and pharmacological approaches we found that IL-27 receptor (IL-27R) signaling promotes HCC development in vivo. Genetic loss of IL-27R suppressed HCC in both carcinogen-induced and non-alcoholic steatohepatitis (NASH)-driven models. Mechanistically, the pro-tumorigenic effect was mediated by an immunoregulatory role of IL-27R within the tumor microenvironment, particularly the suppression of cytotoxic Natural killer (NK) cells. Single-cell RNA analysis further established the role of IL-27R signaling in restraining cytotoxic populations of NK cells. IL-27R ablation enhanced the accumulation and activation of cytotoxic NK cells during acute liver injury and in HCC tumors, while depletion or functional impairment of NK cells abrogated the effect of genetic IL-27R disruption. Taken together, our data suggest an unexpected role of IL-27R signaling as a novel immunological checkpoint regulating innate cytotoxic cell activity and promoting development of HCC of different etiologies. Citation Format: Aleksandra M. Mazitova, Turan Aghayev, Jennifer Fang, Amiran Dzutsev, Giorgio Trinchieri, Kerry S. Campbell, Sergei I. Grivennikov, Ekaterina K. Koltsova. IL-27 signaling regulates anti-cancer immune response in hepatocellular carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3130.

Hepatocellular carcinoma (HCC) is the most common form of liver cancer with poor survival and limited therapeutic options. HCC has different etiologies, typically associated with viral or carcinogenic insults or fatty liver disease and underlying chronic inflammation presents as a major unifying mechanism for tumor promotion. On the other hand, mechanisms of how inflammatory response can regulate anti-cancer immunity in HCC remain incompletely understood.Interleukin (IL)-27 receptor signaling plays an anti-inflammatory role in a variety of infectious and chronic inflammatory diseases. Here, using genetic and pharmacological approaches we found that IL-27 receptor (IL-27R) signaling promotes HCC development in vivo. Genetic loss of IL-27R suppressed HCC development in both toxin/carcinogen-induced diethylnitrosamine (DEN) and non-alcoholic steatohepatitis (NASH)-driven models. Elevated expression of IL-27RA rendered poor prognosis to HCC patients. Mechanistically, the pro-tumorigenic effect was mediated by immunoregulatory role of IL-27R signaling within the tumor microenvironment, particularly the suppression of Natural killer (NK) cells. IL-27R ablation enhanced the accumulation and activation of cytotoxic NK cells during acute liver injury and in HCC tumors, while depletion of NK cells abrogated the effect of genetic IL-27R disruption.Taken together, our data suggest an unexpected role of IL-27R signaling as a novel immunological checkpoint regulating NK cell activity and promoting development of HCC of different etiologies.

The transcription factor ThPOK is critical for homeostasis and differentiation of mature T helper cells. Recent studies have shown that the ThPOK transcriptional silencer element (Sil(ThPOK)) can be reactivated in mature CD4 T cell populations to repress ThPOK and thereby promote differentiation to CD4+ IELs in the intestinal mucosa. However, the molecular basis for this dynamic regulation of Sil(ThPOK) activity remains unknown. Here, using a combination of reverse genetic and reporter approaches, along with global RNA-, ATAC- and ChIP-seq analyses, we demonstrate an important mechanism of ThPOK regulation by which ThPOK binds to the Sil(ThPOK) to drive an auto-feedback loop that maintains its own long-term expression. Disruption of this loop in vivo prevents persistent ThPOK expression in mature peripheral CD4 T cells, leading to genome-wide changes in chromatin accessibility, widespread transcriptional deregulation, and creation of an unusual metastable state, characterized by gain of colonic Treg, cytotoxic T-cell and early DN thymic progenitor specific gene signatures in naïve CD4 T cells. Functionally, abrogation of the ThPOK autoregulatory loop promotes selective differentiation of naive CD4 T cells into GITR(lo) PD-1(lo) CD25(lo) (Triple(lo))Tregs in vitro and in vivo, as well as their conversion to CD4+ IELs in the gut, which provide dominant protection from colitis. Thus, the ThPOK autoregulatory loop represents a key mechanism to physiologically control ThPOK expression and T cell differentiation in the gut, with potential therapeutic relevance.

The cytokine IFN-γ has well-established antibacterial properties against the bacterium Salmonella enterica in phagocytes, but less is known about the effects of IFN-γ on Salmonella-infected nonphagocytic cells, such as intestinal epithelial cells (IECs) and fibroblasts. In this article, we show that exposing human and murine IECs and fibroblasts to IFN-γ following infection with Salmonella triggers a novel form of cell death that is neither pyroptosis nor any of the major known forms of programmed cell death. Cell death required IFN-γ-signaling via STAT1-IRF1–mediated induction of guanylate binding proteins and the presence of live Salmonella in the cytosol. In vivo, ablating IFN-γ signaling selectively in murine IECs led to higher bacterial burden in colon contents and increased inflammation in the intestine of infected mice. Together, these results demonstrate that IFN-γ signaling triggers release of Salmonella from the Salmonella-containing vacuole into the cytosol of infected nonphagocytic cells, resulting in a form of nonpyroptotic cell death that prevents bacterial spread in the gut.

Pancreatic ductal adenocarcinoma (PDAC) represents a significant healthcare problem which is on the rise, primarily because of the late stage diagnosis, early metastasis, and rapidly emerging resistance to therapy. As cancer cells in a typical pancreatic tumor are outnumbered by stromal and immune cells, the role of tumor microenvironment in cancer growth and progression is essential. One of the key regulators of tumor microenvironment in numerous cancers is IFN-γ signaling pathway, which is implicated into regulation of multiple cell types. Using a genetic murine model with conditional IFN-γR2 deficiency in myeloid cells, we performed an orthotopic injection of cell line derived from Pdx1Cre-Kras-P53 model (“KPC” cells). The absence of IFN-γ signaling in myeloid cells led to an increase of tumor burden and size of tumors in these mice. Further analysis demonstrated that IFN-γR2 deletion in myeloid cells changes the differentiation of cancer cells in primary tumors, resulting in a loss of characteristic histological pattern and shifting it towards more aggressive phenotype. Subsequent intrasplenic injection/liver spread model revealed that loss of IFN-γR2 in myeloid cells led to increased metastatic outgrowth of PDAC KPC cells. This effect was partially dependent on the gut microbiota. Deficiency of IFN-γR2 in myeloid cells increases recruitment and infiltration of monocytes in normal and tumor liver tissue. Meanwhile, deletion of IFN-γR2 in several subsets of cancer-associated fibroblasts demonstrated that IFN-γ signaling in different subsets of fibroblast/myofibroblasts may play opposing role in PDAC tumor growth. Our data identifies cell type specific IFN-γ signaling pathway as an important regulator of pancreatic cancer progression and metastasis, that can be potentially exploited as a novel immunooncology target or a biomarker of the disease progression. Citation Format: Elena Ivleva, Ashley Crisostomo, Natalia Andreeva, Sergei Grivennikov. IFN-γ signaling in myeloid and fibroblastic cells regulates pancreatic cancer growth and metastasis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1757.

// Niklas K. Finnberg 1 , Prashanth Gokare 1 , Avital Lev 1 , Sergei I. Grivennikov 2 , Alexander W. MacFarlane IV 3 , Kerry S. Campbell 3 , Ryan M. Winters 4 , Karen Kaputa 4 , Jeffrey M. Farma 5 , Abbas El-Sayed Abbas 6 , Luigi Grasso 7 , Nicholas C. Nicolaides 7 and Wafik S. El-Deiry 1 1 Department of Hematology/Oncology and Molecular Therapeutics Program, Laboratory of Translational Oncology and Experimental Cancer Therapeutics, Fox Chase Cancer Center, Philadelphia, PA, USA 2 Cancer Prevention and Control Program, Fox Chase Cancer Center, Philadelphia, PA, USA 3 Division of Basic Science, Fox Chase Cancer Center, Philadelphia, PA, USA 4 Biosample Repository Facility, Fox Chase Cancer Center, Philadelphia, PA, USA 5 Division of General Surgery, Department of Surgical Oncology, Fox Chase Cancer Center, Philadelphia, PA, USA 6 Division of Thoracic Surgery, Department of Surgical Oncology, Fox Chase Cancer Center, Philadelphia, PA, USA 7 Morphotek Inc., Philadelphia, PA, USA Correspondence to: Wafik S. El-Deiry, email: // Keywords : 3D, organoid, colorectal cancer, immune cells, biomarkers Received : June 24, 2017 Accepted : July 20, 2017 Published : August 05, 2017 Abstract We have developed 3D-tumoroids and tumor slice in vitro culture systems from surgical tumor specimens derived from patients with colorectal cancer (CRC) or lung cancer to evaluate immune cell populations infiltrating cultured tissues. The system incorporates patient’s peripherally and tumor-derived immune cells into tumoroid in vitro cultures to evaluate the ability of the culture to mimic an immunosuppressive tumor microenvironment (ITM). This system enables analysis of tumor response to standard therapy within weeks of surgical resection. Here we show that tumoroid cultures from a CRC patient are highly sensitive to the thymidylate synthase inhibitor 5-fluorouracil (adrucil) but less sensitive to the combination of nucleoside analog trifluridine and thymidine phosphorylase inhibitor tipiracil (Lonsurf). Moreover, re-introduction of isolated immune cells derived from surrounding and infiltrating tumor tissue as well as CD45+ tumor infiltrating hematopoietic cells displayed prolonged (>10 days) survival in co-culture. Established tumor slice cultures were found to contain both an outer epithelial and inner stromal cell compartment mimicking tumor structure in vivo . Collectively, these data suggest that, 3D-tumoroid and slice culture assays may provide a feasible in vitro approach to assess efficacy of novel therapeutics in the context of heterogeneous tumor-associated cell types including immune and non-transformed stromal cells. In addition, delineating the impact of therapeutics on immune cells, and cell types involved in therapeutic resistance mechanisms may be possible in general or for patient-specific responses.

The link between inflammation and cancer was first proposed by R. Virchow. It was later realized that it is chronic inflammation that may promote cancer, whereas acute inflammation can actually block tumor development or even result in cure. Many molecular mediators of these diverse processes have been characterized only during the past 3 decades thanks to the advances in molecular and cellular techniques, as well as due to technologies of reverse genetics. In this chapter we discuss the role of Toll-like receptor (TLR) 4 signaling in cancer and contributions of proinflammatory cytokine signaling (whose expression may be driven by TLR-mediated signals) to tumor-promoting microenvironment. We also discuss recent clinical advances to target these pro-tumorigenic pathways at distinct stages of tumorigenesis.

Relationship between inflammation and cancer is now well-established and represents a paradigm that our immune response does not necessarily serves solely to protect us from infections and cancer. Many specific mechanisms that link chronic inflammation to cancer promotion and metastasis have been uncovered in the recent years. Here we are focusing on the effects that tumors may exert on inflammatory cascades, tuning the immune system ability to cause tumor promotion or regression. In particular, we discuss the contributions of chemokines, cytokines and exosomes to the processes such as induction of inflammation and tumorigenesis. Overall, tumor-elicited inflammation is a key driver of tumor progression and an essential component of tumor microenvironment.

IntroductionAberrant DNA methylation is frequently observed in colorectal cancer (CRC), but the underlying mechanisms are poorly understood. Ten-Eleven Translocation (TET) dioxygenases and DNA repair enzyme Thymine DNA Glycosylase (TDG) are involved in active DNA demethylation by generating and removing, respectively, novel oxidized cytosine species. Mutations ofTET1andTDG, and alterations of the levels of oxidized cytosine species have been identified in human CRC cases, but the biological significance of the TET-TDG demethylation axis in intestinal tumorigenesis is unclear.Material and MethodsWe generatedApcMinmice with additional inactivation ofTet1and/orTdg, and characterized the methylome and transcriptome of intestinal adenomas by DREAM and RNA sequencing, respectively.ResultsTet1-and/orTdg-deficientApcMinmice show enhanced intestinal tumorigenesis in comparison to wild typeTet1andTdg ApcMinmice. Specifically,Tet1and/orTdg-deficientApcMinadenomas manifested increased size or features of erosion and stroma activation. Methylome analysis revealed progressive loss of global DNA hypomethylation in colonic adenomas fromTet1-andTdg-deficientApcMinmice, and hypermethylation of CpG islands inTet1-deficientApcMinmice. In addition, RNA sequencing showed upregulation of genes in inflammatory and immune response pathways inTet1-andTdg-mutant colonic adenomas compared to controlApcMinadenomas.ConclusionsTaken together, these findings demonstrate the important role of active DNA demethylation mediated by TET-TDG in reducing intestinal tumor formation, by modulating the epigenome and inflammatory/immune responses. This study highlights a novel mechanism of epigenetic deregulation during intestinal tumorigenesis with diagnostic, therapeutic and prognostic implications.

Inflammation predisposes to the development of cancer and promotes all stages of tumorigenesis. Cancer cells as well as surrounding stromal and inflammatory cells engage in well-orchestrated reciprocal interactions to form an inflammatory tumor microenvironment (TME). Cells within the TME are highly plastic, continuously changing their phenotypic and functional characteristics. Here we review the origins of inflammation in tumors, and the mechanisms whereby inflammation drives tumor initiation, growth, progression and metastasis. We discuss how tumor promoting inflammation closely resembles inflammatory processes typically found during development, immunity, maintenance of tissue homeostasis or tissue repair, and illuminate the distinctions between tissue-protective and pro-tumorigenic inflammation, including spatio-temporal considerations. Defining the cornerstone rules of engagement governing molecular and cellular mechanisms of tumor-promoting inflammation will be essential for the further development of anti-cancer therapies.

The idea of a potential link between cancer and inflammation was first proposed by R. Virchow in the nineteenth century. However, clear evidence regarding a key role of inflammation in oncogenesis appeared only during the last decade. Now the tumor microenvironment is commonly considered as an obligatory and significant component of almost all types of cancer, and the cells infiltrating such microenvironment are a source of inflammatory cytokines. Such cytokines play a key role in regulating inflammation during both normal immune response and developing cancer. In this review, we explore the role of two inflammatory cytokines interleukin 1 and interleukin 6 in cancer development. These cytokines have pleiotropic effects on various cell types in the tumor microenvironment, particularly being able to regulate pro-oncogenic transcription factors NF-κB and STAT3. For this reason, such cytokines influence key parameters of oncogenesis, increasing cell resistance to apoptosis, proliferation of cancer cells, angiogenesis, invasion and malignancy as well as the ability of tumor cells to respond to anticancer therapy. Here we summarize novel experimental data regarding mechanisms underlying the interaction between chronic inflammation and malignant neoplasms.

Inflammation promotes regeneration of injured tissues through poorly understood mechanisms, some of which involve interleukin (IL)-6 family members, the expression of which is elevated in many diseases including inflammatory bowel diseases and colorectal cancer. Here we show in mice and human cells that gp130, a co-receptor for IL-6 cytokines, triggers activation of YAP and Notch, transcriptional regulators that control tissue growth and regeneration, independently of the gp130 effector STAT3. Through YAP and Notch, intestinal gp130 signalling stimulates epithelial cell proliferation, causes aberrant differentiation and confers resistance to mucosal erosion. gp130 associates with the related tyrosine kinases Src and Yes, which are activated on receptor engagement to phosphorylate YAP and induce its stabilization and nuclear translocation. This signalling module is strongly activated upon mucosal injury to promote healing and maintain barrier function.

Summary Although commensal flora is involved in the regulation of immunity, the interplay between cytokine signaling and microbiota in atherosclerosis remains unknown. We found that interleukin (IL)-23 and its downstream target IL-22 restricted atherosclerosis by repressing pro-atherogenic microbiota. Inactivation of IL-23-IL-22 signaling led to deterioration of the intestinal barrier, dysbiosis, and expansion of pathogenic bacteria with distinct biosynthetic and metabolic properties, causing systemic increase in pro-atherogenic metabolites such as lipopolysaccharide (LPS) and trimethylamine N-oxide (TMAO). Augmented disease in the absence of the IL-23-IL-22 pathway was mediated in part by pro-atherogenic osteopontin, controlled by microbial metabolites. Microbiota transfer from IL-23-deficient mice accelerated atherosclerosis, whereas microbial depletion or IL-22 supplementation reduced inflammation and ameliorated disease. Our work uncovers the IL-23-IL-22 signaling as a regulator of atherosclerosis that restrains expansion of pro-atherogenic microbiota and argues for informed use of cytokine blockers to avoid cardiovascular side effects driven by microbiota and inflammation.

Inflammation has long been suspected to play a major role in the pathogenesis of cancer. Only recently, however, have some mechanisms of its tumor promoting effects become known. Microbes, both commensal and pathogenic, are critical regulators of the host immune system and, ultimately, of inflammation. Consequently, microbes have the potential power to influence tumor progression as well, through a wide variety of routes, including chronic activation of inflammation, alteration of tumor microenvironment, induction of genotoxic responses, and metabolism. In this review, we will provide a general overview of commensal microbiota, inflammation, and cancer, as well as how microbes fit into this emerging field.

Summary MicroRNA deregulation is frequent in human colorectal cancers (CRCs), but little is known as to whether it represents a bystander event or actually drives tumor progression in vivo. We show that miR-135b overexpression is triggered in mice and humans by APC loss, PTEN/PI3K pathway deregulation, and SRC overexpression and promotes tumor transformation and progression. We show that miR-135b upregulation is common in sporadic and inflammatory bowel disease-associated human CRCs and correlates with tumor stage and poor clinical outcome. Inhibition of miR-135b in CRC mouse models reduces tumor growth by controlling genes involved in proliferation, invasion, and apoptosis. We identify miR-135b as a key downsteam effector of oncogenic pathways and a potential target for CRC treatment., Highlights • miR-135b is overexpressed in mouse and human colorectal cancer • miR-135b overexpression is associated with poor clinical outcome • miR-135b activation is triggered by oncogenic pathways in colorectal cancer • miR-135b represents a therapeutic target for colorectal cancer, Valeri et al. identify miR-135b as a key oncogenic pathway effector involved in transformation and colorectal cancer (CRC) progression. Upregulation of miR-135b in human CRCs correlates with poor clinical outcome. miR-135b targets several tumor suppressor genes and is a potential target for CRC therapy.

Compromised integrity of the epithelial barrier function of colon tissue associated with inflammatory bowel diseases (IBDs) or arising from tumor-elicited inflammation (TEI) are associated with and support formation of colorectal cancer (CRC). Apical tight junction (TJ) proteins, including multiple specific claudins, are critical in regulating TJ barrier function and paracellular permeability. Changes in TJ composition in the kidney are strongly linked to the etiology of autosomal dominant polycystic kidney disease (ADPKD), with ADPKD-inducing mutations in PKD1 causing non-leaky barriers that can withstand high hydrostatic pressure within renal cysts a hallmark of this common (1 in 500) inherited disease. Intriguingly, a large population study has found a decreased incidence of CRC in ADPKD patients. Based on these and other suggestive data, we hypothesized that loss of PKD1 would hinder development of ulcerative colitis and initiation of colitis associated cancer, based on reorganizing TJs and TEI in the colon. To evaluate the impact of Pkd1 loss on colon barrier function, we treated 10-12 week old wt or Pkd1fl/fl mice with tamoxifen-induced, Cre expression from the CreERT2 cassette for 5 days with 2.5% DSS in drinking water to induce acute colitis, compared to a no-DSS control group. For the wtcohort, treatment with DSS increased orally gavaged FITC-dextran detectable in serum versus controls. In contrast, no Pkd1-/- mice showed a similar DSS-dependent response to FITC-dextran, suggesting reinforced colon barrier function; further, histopathological assessment confirmed less DSS-induced damage in Pkd1fl/fl mice. Further, based on immunofluorescence (IF) analysis of tissue sections, claudins 4 and 7 associated with increased barrier function) strongly elevated in the colonic epithelium of Pkd1fl/fl versus wt mice, with the expressed proteins having consistently greater localization to cell junctions in Pkd1fl/fl versus wt mice. We also investigated the formation of CRC tumors, using a CDX2-ERT2/Cre model for tamoxifen-induced loss of Apc and/or Pkd1 in the colon to compare tumorigenesis in the Apcfl/fl, Apcfl/fl Pkd1fl/fl, and Apcfl/flPkd1fl/+ genotypes. We observed a highly significant progressive increase in the number and size of Apc mutation-induced tumors based on retention of the Pkd1 gene. Preliminary qRT-PCR analysis of these tumors shows that a Pkd1fl/fl genotype substantially increases CLDN4 in normal and tumor tissue, and decreases TNFα expression in tumors, suggesting reduced inflammation. This work and extended mechanistic characterization identifies PKD1 as a vital regulator of signaling systems already associated with colitis and CRC. Understanding the role of PKD1 and its effectors may provide useful information to genetic counselors assessing risk of IBD and CRC, and suggest therapeutic strategies. Citation Format: Anna S. Nikonova, Anna Kiseleva, Ilya Serebriiskii, Sergei Grivennikov, Erica A. Golemis. PKD1 regulates susceptibility to ulcerative colitis and colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3459.

Tumor and stromal cell plasticity induced by different tumor microenvironments (TMEs) has a significant impact on tumorigenesis of colorectal carcinoma (CRC). Specifically the role of the immunological TME has been investigated intensively. The prognosis of CRC is influenced by the density and localization of infiltrating T cells. Moreover, the presence of an mRNA expression profile indicative of a type 1 adaptive immune response (high IFN-γ and IFN-induced gene expression, cytotoxic T cell (CTL) signature) represents a positive prognostic factor for patients with CRC. IFN-γ is a major mediator of the Th1 immune response, which is produced by NK, NKT, Th1 cells and CTL. The role of IFN-γ in anti-tumor immune response has been mainly attributed to the immune-modulatory activity of the cytokine, such as the recruitment and activation of cytotoxic T-cells or monocytes. However, the expression of the IFN-γ receptor is ubiquitous and studies of IFN-γ-induced genes expression in CRC revealed that many different cell types are stimulated by the cytokine within a tumor, including tumor cells and stromal cells such as macrophages/monocytes and endothelial cells. In vitro, IFN-γ shows anti-tumorigenic activities in CRC cell lines, activates monocytes and exerts potent anti-angiogenic effects on endothelial cells. Here we investigated the cell-type-specific impact of the response to IFN-γ on CRC development using mouse strains with conditional knock-out of the IFN-γ receptor in intestinal epithelial cells (Villin-Cre), T-cells (CD4-Cre), myeloid cells (CD11b-Cre) and endothelial cells (Tie2-Cre + BMT). Tumor growth was chemically induced by injection of azoxymethane combined with three cycles of treatment with dextran-sodium sulfate. This model recapitulates inflammation-induced carcinogenesis. Our results revealed increased tumor numbers and load when the IFN-γ response was blocked in epithelial cells, despite an initial attenuation of inflammation. Tumors that developed in Ifngr2-Villin-Cre mice showed an attenuated IFN-γ response, and a decrease of CD8+ T cell infiltration, cell death and hypoxia. Abrogation of the IFN-γ-response in endothelial cells also fostered tumor growth, which could be attributed to an increase of angiogenesis. Surprisingly, only a modest effect was seen in the T-cell-specific knock-out, in comparison to the myeloid-specific knock-out, which was associated with a strong increase of tumor numbers and load. The latter indicated that the anti-tumorigenic activity mediated by IFN-γ relies to a larger extent on the innate than on the adaptive immune response. Our study documents that the anti-tumorigenic activity of IFN-γ is based on direct effects on epithelial tumor cells and on the vasculature, as well as on the involvement of the innate immune response. Citation Format: Nathalie Britzen-Laurent, Wei Guo, Victoria Langer, Svetlana Khoziainova, Thomas Weisenburger, Thomas H. Winkler, Julia Straube, Maximilian J. Waldner, Christoph Becker, Elisabeth Naschberger, Lisa Skottke, Tripal Philipp, Sergei Grivennikov, Michael Stürzl. Role of IFN-gamma-activation of distinct tumor and stromal cell populations in colorectal carcinoma pathogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 5162.

Command and Control Innate lymphoid cells are vital for the development of gut-associated lymphoid tissues, maintenance of the epithelial barrier, and protection against intestinal microbes; their dysfunction can promote immune pathology. Immunoglobulin A (IgA) production is important for maintenance of the gut epithelial barrier and the composition of the gut microbiota. Through the generation of knockout mouse models, Kruglov et al. (p. 1243 ) were able to distinguish how soluble and membrane-bound lymphotoxins expressed by innate lymphoid cells in the gut specifically regulate IgA production and thereby control gut microbiota composition.

The colon tumor microenvironment (TME) is becoming increasingly recognized as a complex but central player in the development of many cancers. Previously, we identified an oncogenic role for the mRNA-binding protein IMP1 (IGF2BP1) in the epithelium during colon tumorigenesis. In the current study, we reveal the contribution of stromal IMP1 in the context of colitis-associated colon tumorigenesis. Interestingly, stromal deletion of Imp1 (Dermo1Cre;Imp1LoxP/LoxP, or Imp1ΔMes) in the azoxymethane/dextran sodium sulfate (AOM/DSS) model of colitis-associated cancer resulted in increased tumor numbers of larger size and more advanced histological grade than controls. In addition, Imp1ΔMes mice exhibited a global increase in pro-tumorigenic microenvironment factors, including enhanced inflammation and stromal components. Evaluation of purified mesenchyme from AOM/DSS-treated Imp1ΔMes mice demonstrated an increase in hepatocyte growth factor (HGF), which has not been associated with regulation via IMP1. Genetic knockdown of Imp1 in human primary fibroblasts confirmed an increase in HGF with Imp1 loss, demonstrating a specific, cell-autonomous role for Imp1 loss to increase HGF expression. Taken together, these data demonstrate a novel tumor-suppressive role for IMP1 in colon stromal cells and underscore an exquisite, context-specific function for mRNA-binding proteins, such as IMP1, in disease states.

The tumor microenvironment offers multiple targets for cancer therapy, including pro-tumorigenic inflammation. Natural compounds represent an enormous source of new anti-inflammatory and anticancer agents. We previously showed that the styryl lactone goniothalamin (GTN) has promising antiproliferative and anti-inflammatory activities. Because inflammation is a major driver of colorectal cancer (CRC), we therefore evaluated the therapeutic and preventive potentials of GTN in colitis, colitis-associated cancer (CAC) and spontaneous CRC. First, in a simplistic model of inflammation in vitro, GTN was able to inhibit cytokine production in bone marrow-derived macrophages induced by lipopolysaccharide. Next, in dextran sulfate sodium (DSS) induced-colitis model, mice treated with GTN displayed restored tissue architecture, increased cell proliferation in the colonic crypts and reduced epithelial damage. Moreover, colon tissue from GTN-treated mice had significantly less expression of the inflammatory genes interleukin 1β (IL-1β), tumor necrosis factor α (TNF-α), interleukin 6 (IL-6), S100A9, interleukin 23A (IL-23A), IL-22 and IL-17A In the azoxymethane/DSS model of CAC, GTN reduced tumor multiplicity, load and size. Additionally, GTN suppressed production of IL-6, IL-17 and TNF-α in tumor tissue, as well as abrogated stromal immune cell activation and nuclear translocation of NF-κB. Finally, in a tamoxifen inducible model of sporadic CRC, GTN-treated mice had significantly fewer tumors and decreased levels of IL-17A, IL-6, S100A9 and TNF-α protein within the tumors. These results suggest that GTN possesses anti-inflammatory and antitumor activities and represents a preventive and therapeutic agent modulating the inflammatory environment in the colon during colitis as well as CAC and CRC development.

Innate immunity controls pathogen replication and spread. Yet, certain pathogens, such as Hepatitis C Virus (HCV), escape immune elimination and establish persistent infections that promote chronic inflammation and related diseases. Whereas HCV regulatory proteins that attenuate antiviral responses are known, those that promote inflammation and liver injury remain to be identified. Here, we show that transient expression of HCV RNA-dependent RNA polymerase (RdRp), NS5B, in mouse liver and human hepatocytes results in production of small RNA species that activate innate immune signaling via TBK1-IRF3 and NF-κB and induce cytokine production, including type I interferons (IFN) and IL-6. NS5B-expression also results in liver damage.

In activated macrophages, the anti-inflammatory cytokine IL-10 inhibits expression of molecules that propagate inflammation in a manner that depends on transcription factor STAT3. Expression of IL-10 is regulated posttranscriptionally by the RNA-binding protein tristetraprolin (TTP), which destabilizes IL-10 mRNA in activated macrophages. Using LPS-activated bone marrow-derived murine macrophages, we demonstrate that TTP is a negative regulator of the IL-10/STAT3 anti-inflammatory response. LPS-stimulated TTP-deficient macrophages overproduced IL-10, contained increased amounts of activated STAT3, and showed reduced expression of inflammatory cytokines, including cytokines encoded by TTP target mRNAs. Thus, in LPS-stimulated TTP-deficient macrophages, increased IL-10/STAT3 anti-inflammatory control was dominant over the mRNA stabilization of specific TTP targets. The TTP gene promoter contains a conserved STAT3 binding site, and IL-10 induces STAT3 recruitment to this site. Correspondingly, STAT3 was required for efficient IL-10–induced TTP expression. Hence, by inducing TTP expression, STAT3 activates a negative regulatory loop that controls the IL-10/STAT3 anti-inflammatory response.

Inhibition of programmed cell death is considered to be a major aspect of tumorigenesis. Indeed, several key oncogenic transcription factors, such as NF-κB and STAT3, exert their tumor-promoting activity at least in part through upregulation of survival genes. However, many cancers develop in response to chronic tissue injury, in which the resulting cell death increases the tumorigenic potential of the neighboring cells. In this review, we discuss a resolution to this paradox based on cell death-mediated induction of tumor promoting inflammatory cytokines, which enhance cell survival and trigger compensatory proliferation in response to tissue injury.

In a mouse model of ErbB-driven mammary tumours, Tan et al. find a role for RANKL (receptor activator of nuclear factor-κB ligand) in the formation of lung metastases. RANKL is produced by regulatory T cells infiltrating the primary tumours, and acts through its receptor RANK, which is expressed on the cancer cells. Targeting RANKL may therefore prove useful in reducing breast cancer metastases. In a mouse model of Erbb2-driven mammary tumours, this study finds a role for RANKL in the formation of lung metastases. RANKL is produced by regulatory T cells infiltrating the primary tumours and acts via its receptor RANK expressed on the cancer cells. Targeting RANKL may therefore prove useful in reducing breast cancer metastases. Inflammatory mechanisms influence tumorigenesis and metastatic progression even in cancers whose aetiology does not involve pre-existing inflammation or infection, such as breast and prostate cancers1. For instance, prostate cancer metastasis is associated with the infiltration of lymphocytes into advanced tumours and the upregulation of two tumour-necrosis-factor family members: receptor activator of nuclear factor-κB (RANK) ligand (RANKL) and lymphotoxin2. But the source of RANKL and its role in metastasis have not been established. RANKL and its receptor RANK control the proliferation of mammary lobuloalveolar cells during pregnancy3 through inhibitor of nuclear factor-κB (IκB) kinase-α (IKK-α)4, a protein kinase that is needed for the self-renewal of mammary cancer progenitors5 and for prostate cancer metastasis2. We therefore examined whether RANKL, RANK and IKK-α are also involved in mammary/breast cancer metastasis. Indeed, RANK signalling in mammary carcinoma cells that overexpress the proto-oncogene Erbb2 (also known as Neu)6, which is frequently amplified in metastatic human breast cancers7,8, was important for pulmonary metastasis. Metastatic spread of Erbb2-transformed carcinoma cells also required CD4+CD25+ T cells, whose major pro-metastatic function was RANKL production. Most RANKL-producing T cells expressed forkhead box P3 (FOXP3), a transcription factor produced by regulatory T cells, and were located next to smooth muscle actin (SMA)+ stromal cells in mouse and human breast cancers. The dependence of pulmonary metastasis on T cells was replaceable by exogenous RANKL, which also stimulated pulmonary metastasis of RANK+ human breast cancer cells. These results are consistent with the adverse impact of tumour-infiltrating CD4+ or FOXP3+ T cells on human breast cancer prognosis9,10 and suggest that the targeting of RANKL–RANK can be used in conjunction with the therapeutic elimination of primary breast tumours to prevent recurrent metastatic disease.

Cirrhosis is the end result of chronic liver disease. Hepatic stellate cells (HSC) are believed to be the major source of collagen-producing myofibroblasts in cirrhotic livers. Portal fibroblasts, bone marrow-derived cells, and epithelial to mesenchymal transition (EMT) might also contribute to the myofibroblast population in damaged livers. Fibroblast-specific protein 1 (FSP1, also called S100A4) is considered a marker of fibroblasts in different organs undergoing tissue remodeling and is used to identify fibroblasts derived from EMT in several organs including the liver. The aim of this study was to characterize FSP1-positive cells in human and experimental liver disease. FSP1-positive cells were increased in human and mouse experimental liver injury including liver cancer. However, FSP1 was not expressed by HSC or type I collagen-producing fibroblasts. Likewise, FSP1-positive cells did not express classical myofibroblast markers, including αSMA and desmin, and were not myofibroblast precursors in injured livers as evaluated by genetic lineage tracing experiments. Surprisingly, FSP1-positive cells expressed F4/80 and other markers of the myeloid-monocytic lineage as evaluated by double immunofluorescence staining, cell fate tracking, flow cytometry, and transcriptional profiling. Similar results were obtained for bone marrow-derived and peritoneal macrophages. FSP1-positive cells were characterized by increased expression of COX2, osteopontin, inflammatory cytokines, and chemokines but reduced expression of MMP3 and TIMP3 compared with Kupffer cells/macrophages. These findings suggest that FSP1 is a marker of a specific subset of inflammatory macrophages in liver injury, fibrosis, and cancer.

Secondary lymphoid organs provide a unique microenvironment for generation of immune responses. Using a cell type–specific conditional knockout approach, we have dissected contributions of tumor necrosis factor (TNF) produced by B cells (B-TNF) or T cells (T-TNF) to the genesis and homeostatic organization of secondary lymphoid organs. In spleen, lymph nodes and Peyer patches, the cellular source of TNF, and its molecular form (soluble versus membrane-bound) appeared distinct. In spleen, in addition to major B-TNF signal, a complementary T-TNF signal contributed to the microstructure. In contrast, B-TNF predominantly controlled the development of follicular dendritic cells and B-cell follicles in Peyer patches. In lymph nodes, cooperation between TNF expressed by B and T cells was necessary for the maintenance of microarchitecture and for generation of an efficient humoral immune response. Unexpectedly, soluble but not membrane TNF expressed by B cells was essential for the organization of the secondary lymphoid organs. Thus, the maintenance of each type of secondary lymphoid organ is orchestrated by distinct contributions of membrane-bound and soluble TNF produced by B and T lymphocytes.

Inflammatory responses play decisive roles at different stages of tumor development, including initiation, promotion, malignant conversion, invasion, and metastasis. Inflammation also affects immune surveillance and responses to therapy. Immune cells that infiltrate tumors engage in an extensive and dynamic crosstalk with cancer cells, and some of the molecular events that mediate this dialog have been revealed. This review outlines the principal mechanisms that govern the effects of inflammation and immunity on tumor development and discusses attractive new targets for cancer therapy and prevention.

Colitis-associated cancer (CAC) is the most serious complication of inflammatory bowel disease. Proinflammatory cytokines have been suggested to regulate preneoplastic growth during CAC tumorigenesis. Interleukin 6 (IL-6) is a multifunctional NF-kappaB-regulated cytokine that acts on epithelial and immune cells. Using genetic tools, we now demonstrate that IL-6 is a critical tumor promoter during early CAC tumorigenesis. In addition to enhancing proliferation of tumor-initiating cells, IL-6 produced by lamina propria myeloid cells protects normal and premalignant intestinal epithelial cells (IECs) from apoptosis. The proliferative and survival effects of IL-6 are largely mediated by the transcription factor Stat3, whose IEC-specific ablation has profound impact on CAC tumorigenesis. Thus, the NF-kappaB-IL-6-Stat3 cascade is an important regulator of the proliferation and survival of tumor-initiating IECs.

Metastatic progression depends on genetic alterations intrinsic to cancer cells as well as the inflammatory microenvironment of advanced tumours. To understand how cancer cells affect the inflammatory microenvironment, we conducted a biochemical screen for macrophage-activating factors secreted by metastatic carcinomas. Here we show that, among the cell lines screened, Lewis lung carcinoma (LLC) were the most potent macrophage activators leading to production of interleukin-6 (IL-6) and tumour-necrosis factor-alpha (TNF-alpha) through activation of the Toll-like receptor (TLR) family members TLR2 and TLR6. Both TNF-alpha and TLR2 were found to be required for LLC metastasis. Biochemical purification of LLC-conditioned medium (LCM) led to identification of the extracellular matrix proteoglycan versican, which is upregulated in many human tumours including lung cancer, as a macrophage activator that acts through TLR2 and its co-receptors TLR6 and CD14. By activating TLR2:TLR6 complexes and inducing TNF-alpha secretion by myeloid cells, versican strongly enhances LLC metastatic growth. These results explain how advanced cancer cells usurp components of the host innate immune system, including bone-marrow-derived myeloid progenitors, to generate an inflammatory microenvironment hospitable for metastatic growth.

Lymphotoxin-beta (LTbeta) is a key regulator of immune system development, but also affects late stages in hair development. In addition, high expression of LTbeta at an early stage in epidermis hinted at a further function in hair follicle induction or epithelial development. We report that hair follicles were normally induced in LTbeta(-/-) skin, but the periderm detached from the epidermis earlier, accompanied by premature appearance of keratohyalin granules. Expression profiling revealed dramatic down-regulation of a gene cluster encoding periderm-specific keratin-associated protein 13 and four novel paralogs in LTbeta(-/-) skin prior to periderm detachment. Epidermal differentiation markers, including small proline-rich proteins, filaggrins and several keratins, were also affected, but transiently in LTbeta(-/-) skin at the time of abnormal periderm detachment. As expected, Tabby mice, which lack the EDA gene, the putative upstream regulator of LTbeta in skin, showed similar though milder periderm histopathology and alterations in gene expression. Overall, LTbeta shows a primary early function in periderm differentiation, with later transient effects on epidermal and hair follicle differentiation.

Tumor necrosis factor (TNF) is critical and non-redundant to control Mycobacterium tuberculosis infection and cannot be replaced by other proinflammatory cytokines. Overproduction of TNF may cause immunopathology, while TNF neutralization reactivates latent and chronic, controlled infection, which is relevant for the use of neutralizing TNF therapies in patients with rheumatoid arthritis.

Acute and chronic airway inflammations caused by environmental agents including endotoxin represent an increasing health problem. Local TNF production may contribute to lung dysfunction and inflammation, although pulmonary neutrophil recruitment occurs in the absence of TNF. First, we demonstrate that membrane-bound TNF is sufficient to mediate the inflammatory responses to lipopolysaccharide (LPS). Secondly, using cell type-specific TNF-deficient mice we show that TNF derived from either macrophage/neutrophil (M/N) or T lymphocytes have differential effects on LPS-induced respiratory dysfunction (enhanced respiratory pause, Penh) and pulmonary neutrophil recruitment. While Penh, vascular leak, neutrophil recruitment, TNF, and thymus- and activation-regulated chemokine/CCL17 (TARC) expression in the lung were reduced in M/N-deficient mice, T cell-specific TNF-deficient mice displayed augmented Penh, vascular leak, neutrophil influx, increased CD11c+ cells and expression of TNF, TARC and murine CXC chemokines KC/CXCL1 in the lung. In conclusion, inactivation of TNF in either M/N or T cells has differential effects on LPS-induced lung disease, suggesting that selective deletion of TNF in T cells may aggravate airway pathology.