Susan E. Erdman

Host lifestyle affects human microbiota on daily timescales

BackgroundDisturbance to human microbiota may underlie several pathologies. Yet, we lack a comprehensive understanding of how lifestyle affects the dynamics of human-associated microbial communities.ResultsHere, we link over 10,000 longitudinal measurements of human wellness and action to the daily gut and salivary microbiota dynamics of two individuals over the course of one year. These time series show overall microbial communities to be stable for months. However, rare events in each subjects’ life rapidly and broadly impacted microbiota dynamics. Travel from the developed to the developing world in one subject led to a nearly two-fold increase in the Bacteroidetes to Firmicutes ratio, which reversed upon return. Enteric infection in the other subject resulted in the permanent decline of most gut bacterial taxa, which were replaced by genetically similar species. Still, even during periods of overall community stability, the dynamics of select microbial taxa could be associated with specific host behaviors. Most prominently, changes in host fiber intake positively correlated with next-day abundance changes among 15% of gut microbiota members.ConclusionsOur findings suggest that although human-associated microbial communities are generally stable, they can be quickly and profoundly altered by common human actions and experiences.

CD4+ CD25+ Regulatory T Lymphocytes Inhibit Microbially Induced Colon Cancer in Rag2-Deficient Mice

Inflammatory bowel diseases, including ulcerative colitis and Crohns disease, increase the risk of colorectal cancer in humans. It has been recently shown in humans and animal models that intestinal microbiota and host immunity are integral in the progression of large bowel diseases. Lymphocytes are widely believed to prevent bacterially induced inflammation in the bowel, and lymphocytes are also critical in protecting against primary tumors of intestinal epithelia in mice. Taken together, this raises the possibility that lymphocytes may inhibit colon carcinogenesis by reducing bacterially driven inflammation. To examine the role of bacteria, lymphocytes, and inflammatory bowel disease in the development of colon cancer, 129/SvEv Rag-2-deficient and congenic wild-type mice were orally inoculated with a widespread enteric mouse bacterial pathogen, Helicobacter hepaticus, or sham-dosed with media only. H. hepaticus-infected Rag2-/-, but not sham-dosed Rag2-/- mice, rapidly developed colitis and large bowel carcinoma. This demonstrated a link between microbially driven inflammation and cancer in the lower bowel and suggested that innate immune dysregulation may have an important role in inflammatory bowel disease and progression to cancer. H. hepaticus-infected wild-type mice did not develop inflammation or carcinoma showing that lymphocytes were required to prevent bacterially induced cancer at this site. Adoptive transfer with CD4+ CD45RBlo CD25+ regulatory T cells into Rag-deficient hosts significantly inhibited H. hepaticus-induced inflammation and development of cancer. These results suggested that the ability of CD4+ T cells to protect against intestinal cancer was correlated with their ability to reduce bacterially induced inflammatory bowel disease. Further, regulatory T cells may act directly on the innate immune system to reduce or prevent disease. These roles for T cells in protection against colon carcinoma may have implications for new modes of prevention and treatment of cancer in humans.

Mast cells are an essential hematopoietic component for polyp development

It is generally agreed that most colon cancers develop from adenomatous polyps, and it is this fact on which screening strategies are based. Although there is overwhelming evidence to link intrinsic genetic lesions with the formation of these preneoplastic lesions, recent data suggest that the tumor stromal environment also plays an essential role in this disease. In particular, it has been suggested that CD34+ immature myeloid precursor cells are required for tumor development and invasion. Here we have used mice conditional for the stabilization of β-catenin or defective for the adenomatous polyposis coli (APC) gene to reinvestigated the identity and importance of tumor-infiltrating hematopoietic cells in polyposis. We show that, from the onset, polyps are infiltrated with proinflammatory mast cells (MC) and their precursors. Depletion of MC either pharmacologically or through the generation of chimeric mice with genetic lesions in MC development leads to a profound remission of existing polyps. Our data suggest that MC are an essential hematopoietic component for preneoplastic polyp development and are a novel target for therapeutic intervention.

Cancer-Associated Fibroblasts Drive the Progression of Metastasis through both Paracrine and Mechanical Pressure on Cancer Tissue

Neoplastic cells recruit fibroblasts through various growth factors and cytokines. These “cancer-associated fibroblasts” (CAF) actively interact with neoplastic cells and form a myofibroblastic microenvironment that promotes cancer growth and survival and supports malignancy. Several products of their paracrine signaling repertoire have been recognized as tumor growth and metastasis regulators. However, tumor-promoting cell signaling is not the only reason that makes CAFs key components of the “tumor microenvironment,” as CAFs affect both the architecture and growth mechanics of the developing tumor. CAFs participate in the remodeling of peritumoral stroma, which is a prerequisite of neoplastic cell invasion, expansion, and metastasis. CAFs are not present peritumorally as individual cells but they act orchestrated to fully deploy a desmoplastic program, characterized by “syncytial” (or collective) configuration and altered cell adhesion properties. Such myofibroblastic cohorts are reminiscent of those encountered in wound-healing processes. The view of “cancer as a wound that does not heal” led to useful comparisons between wound healing and tumorigenesis and expanded our knowledge of the role of CAF cohorts in cancer. In this integrative model of cancer invasion and metastasis, we propose that the CAF-supported microenvironment has a dual tumor-promoting role. Not only does it provide essential signals for cancer cell dedifferentiation, proliferation, and survival but it also facilitates cancer cell local invasion and metastatic phenomena. Mol Cancer Res; 10(11); 1403–18. ©2012 AACR.

CD4+CD25+ Regulatory Lymphocytes Induce Regression of Intestinal Tumors in ApcMin/+ Mice

Colorectal cancer in humans results from sequential genetic changes in intestinal epithelia commencing with inactivation of the APC tumor suppressor gene. Roles for host immunity in epithelial tumorigenesis are poorly understood. It has been previously shown that CD4+CD25+ lymphocytes inhibit colitis-associated epithelial tumors in Rag-deficient mice. Here we show that addition of CD4+CD25+ lymphocytes in ApcMin/+ mice reduces multiplicity of epithelial adenomas. Interleukin-10 was required in regulatory cells for therapeutic effect. Recipients of regulatory cells showed increased apoptosis and down-regulation of cyclooxygenase-2 within tumors coincident with tumor regression. These data suggest a role for regulatory lymphocytes in epithelial homeostasis in the ApcMin/+ mouse model of intestinal polyposis. Similarities with cancer of the breast, prostate, lung, and other sites raise the possibility of broader roles for regulatory lymphocytes in prevention and treatment of epithelial cancers in humans.

E2F4 Is Essential for Normal Erythrocyte Maturation and Neonatal Viability

The retinoblastoma protein (pRB) plays a key role in the control of normal development and proliferation through the regulation of the E2F transcription factors. We generated a mutant mouse model to assess the in vivo role of the predominant E2F family member, E2F4. Remarkably, loss of E2F4 had no detectable effect on either cell cycle arrest or proliferation. However, E2F4 was essential for normal development. E2f4-/- mice died of an increased susceptibility to opportunistic infections that appeared to result from craniofacial defects. They also displayed a variety of erythroid abnormalities that arose from a cell autonomous defect in late stage maturation. This suggests that E2F4 makes a major contribution to the control of erythrocyte development by the pRB tumor suppressor.

Gastroenteritis in NF-κB-Deficient Mice Is Produced with Wild-Type Camplyobacter jejuni but Not with C. jejuni Lacking Cytolethal Distending Toxin despite Persistent Colonization with Both Strains

ABSTRACT Campylobacter jejuni continues to be a leading cause of bacterial enteritis in humans. However, because there are no readily available animal models to study the pathogenesis of C. jejuni-related diseases, the significance of potential virulence factors, such as cytolethal distending toxin (CDT), in vivo are poorly understood. Mice deficient in NF-κB subunits (p50−/− p65+/−) in a C57BL/129 background are particularly susceptible to colitis induced by another enterohepatic microaerobe, Helicobacter hepaticus, which, like C. jejuni, produces CDT. Wild-type C. jejuni 81-176 and an isogenic mutant lacking CDT activity (cdtB mutant) were inoculated into NF-κB-deficient (3X) and C57BL/129 mice. Wild-type C. jejuni colonized 29 and 50% of the C57BL/129 mice at 2 and 4 months postinfection (p.i.), respectively, whereas the C. jejuni cdtB mutant colonized 50% of the C57BL/129 mice at 2 p.i. but none of the mice at 4 months p.i. Although the C57BL/129 mice developed mild gastritis and typhlocolitis, they had robust immunoglobulin G (IgG) and Th1-promoted IgG2a humoral responses to both the wild-type strain and the C. jejuni cdtB mutant. In contrast, 75 to 100% of the 3X mice were colonized with both the wild type and the C. jejuni cdtB mutant at similar levels at all times examined. Wild-type C. jejuni caused moderately severe gastritis and proximal duodenitis in 3X mice that were more severe than the gastrointestinal lesions caused by the C. jejuni cdtB mutant. Persistent colonization of NF-κB-deficient mice with the wild type and the C. jejuni cdtB mutant was associated with significantly impaired IgG and IgG2a humoral responses (P < 0.001), which is consistent with an innate or adaptive immune system defect(s). These results suggest that the mechanism of clearance of C. jejuni is NF-κB dependent and that CDT may have proinflammatory activity in vivo, as well as a potential role in the ability of C. jejuni to escape immune surveillance. NF-κB-deficient mice should be a useful model to further study the role of CDT and other aspects of C. jejuni pathogenesis.

Nitric oxide and TNF-alpha trigger colonic inflammation and carcinogenesis in Helicobacter hepaticus-infected, Rag2-deficient mice.

Recombinase-activating gene-2-deficient (Rag2−/−) mice lacking functional lymphocytes provide a useful model of chronic inflammatory bowel disease-emulating events in human colon cancer. Infection of Rag2−/− mice with Helicobacter hepaticus led to accumulation of macrophages and neutrophils in the colon, a process temporally related to up-regulation of tissue inducible nitric oxide synthase (iNOS) expression at the site of infection and increased nitric oxide (NO) production, as evidenced by urinary excretion of nitrate. Progressive development of increasingly severe inflammation, hyperplasia, dysplasia, and cancer accompanied these changes. Concurrent administration of an iNOS inhibitor prevented NO production and abrogated epithelial pathology and inhibited the onset of cancer. The presence of Gr-1+ neutrophils and elevated tumor necrosis factor-α (TNF-α) expression in colon were required for increased iNOS expression and cancer, whereas interleukin-10 (IL-10) down-regulated TNF-α and iNOS expression and suppressed cancer. Anti-inflammatory CD4+ regulatory lymphocytes also down-regulated iNOS and reduced cancer formation. Collectively, these results confirm essential roles for inflammation, increased TNF-α expression, and elevated NO production in colon carcinogenesis.

Innate Immune Inflammatory Response against Enteric Bacteria Helicobacter hepaticus Induces Mammary Adenocarcinoma in Mice

Inflammation associated with bacterial infections is a risk factor for cancers in humans, yet its role in breast cancer remains poorly understood. We have previously shown that innate immune inflammatory response against intestinal bacteria is sufficient to induce colon cancer. Here we report that infecting Rag2-deficient C57BL/6 Apc(Min/+) mice with an intestinal bacterial pathogen, Helicobacter hepaticus, significantly promotes mammary carcinoma in females and enhances intestinal adenoma multiplicity by a tumor necrosis factor alpha (TNFalpha)-dependent mechanism. The mammary and intestinal tumor development as well as the increase in proinflammatory mediators is suppressed by adoptive transfer of interleukin 10-competent CD4+CD45RB(lo)CD25+ regulatory (T(R)) cells. Furthermore, prior exposure of donor mice to H. hepaticus significantly enhances antitumor potency of their T(R) cells. Interestingly, these microbially experienced T(R) cells suppress tumorigenesis more effectively in recipient mice irrespective of their tumor etiology. These data suggest that infections with enteric pathogens enhance T(R)-cell potency and protect against epithelial cancers later in life, potentially explaining paradoxical increases in cancer risk in developed countries having more stringent hygiene practices. The possibility that dysregulated gut microbial infections in humans may lead to cancer in anatomically distant organs, such as breast, highlights the need for novel immune-based strategies in cancer prevention and treatment.

Cutting Edge: Typhlocolitis in NF-κB-Deficient Mice

Activation of inflammatory gene expression by the transcription factor NF-κB is a central pathway in many inflammatory disorders, including colitis. Increased NF-κB activity has been linked with development of colitis in humans and animal models, thus it was unexpected when NF-κB-deficient mice developed spontaneous typhlocolitis. To further characterize this finding, we induced typhlocolitis in rederived NF-κB-deficient mice using intragastric infection with Helicobacter hepaticus. At 6 wk postinfection (PI), severe colitis with increased type 1 cytokine expression was seen in infected mice that lacked the p50 subunit of NF-κB and were also heterozygous for the p65 subunit of NF-κB(p50−/−p65+/−). Mice lacking the p50 subunit alone (p50−/−) were less severely affected, and wild-type mice and p65+/− mice were unaffected. T cell development in NF-κB-deficient mice was normal. These data indicate that p50 and p65 subunits of NF-κB have an unexpected role in inhibiting the development of colitis.