Krüppel-like factor 5 is a crucial mediator of intestinal tumorigenesis in mice harboring combined ApcMin and KRASV12 mutations

<p>Abstract</p> <p>Background</p> <p>Both mutational inactivation of the adenomatous polyposis coli (<it>APC</it>) tumor suppressor gene and activation of the <it>KRAS </it>oncogene are implicated in the pathogenesis of colorectal cancer. Mice harboring a germline <it>Apc</it>^{<it>Min </it>}mutation or intestine-specific expression of the <it>KRAS</it>^<it>V</it>12gene have been developed. Both mouse strains develop spontaneous intestinal tumors, including adenoma and carcinoma, though at a different age. The zinc finger transcription factor Krüppel-like factor 5 (KLF5) has previously been shown to promote proliferation of intestinal epithelial cells and modulate intestinal tumorigenesis. Here we investigated the <it>in vivo </it>effect of <it>Klf5 </it>heterozygosity on the propensity of <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12double transgenic mice to develop intestinal tumors.</p> <p>Results</p> <p>At 12 weeks of age, <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12mice had three times as many intestinal tumors as <it>Apc</it>^{<it>Min </it>}mice. This increase in tumor number was reduced by 92% in triple transgenic <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-mice. The reduction in tumor number in <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-mice was also statistically significant compared to <it>Apc</it>^{<it>Min </it>}mice alone, with a 75% decrease. Compared with <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12, tumors from both <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-and <it>Apc</it>^{<it>Min </it>}mice were smaller. In addition, tumors from <it>Apc</it>^{<it>Min </it>}mice were more distally distributed in the intestine as contrasted by the more proximal distribution in <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12and <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-mice. Klf5 levels in the normal-appearing intestinal mucosa were higher in both <it>Apc</it>^{<it>Min </it>}and <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12mice but were attenuated in <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-mice. The levels of β-catenin, cyclin D1 and Ki-67 were also reduced in the normal-appearing intestinal mucosa of <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-mice when compared to <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12mice. Levels of pMek and pErk1/2 were elevated in the normal-appearing mucosa of <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12mice and modestly reduced in Apc^Min/<it>KRAS</it>^<it>V</it>12/<it>Klf5</it>^+/-mice. Tumor tissues displayed higher levels of both Klf5 and β-catenin, irrespective of the mouse genotype from which tumors were derived.</p> <p>Conclusions</p> <p>Results of the current study confirm the cumulative effect of <it>Apc </it>loss and oncogenic <it>KRAS </it>activation on intestinal tumorigenesis. The drastic reduction in tumor number and size due to <it>Klf5 </it>heterozygosity in <it>Apc</it>^<it>Min</it>/<it>KRAS</it>^<it>V</it>12mice indicate a critical function of KLF5 in modulating intestinal tumor initiation and progression.</p

Krüppel-like factor 5 is an important mediator for lipopolysaccharide-induced proinflammatory response in intestinal epithelial cells

Lipopolysaccharide (LPS) is a bacterially-derived endotoxin that elicits a strong proinflammatory response in intestinal epithelial cells. It is well established that LPS activates this response through NF-κB. In addition, LPS signals through the mitogen-activated protein kinase (MAPK) pathway. We previously demonstrated that the Krüppel-like factor 5 [KLF5; also known as intestine-enriched Krüppel-like factor (IKLF)] is activated by the MAPK. In the current study, we examined whether KLF5 mediates the signaling cascade elicited by LPS. Treatment of the intestinal epithelial cell line, IEC6, with LPS resulted in a dose- and time-dependent increase in KLF5 messenger RNA (mRNA) and protein levels. Concurrently, mRNA levels of the p50 and p65 subunits of NF-κB were increased by LPS treatment. Pretreatment with the MAPK inhibitor, U0126, or the LPS antagonist, polymyxin B, resulted in an attenuation of KLF5, p50 and p65 NF-κB subunit mRNA levels from LPS treatment. Importantly, suppression of KLF5 by small interfering RNA (siRNA) resulted in a reduction in p50 and p65 subunit mRNA levels and NF-κB DNA binding activity in response to LPS. LPS treatment also led to an increase in secretion of TNF-α and IL-6 from IEC6, both of which were reduced by siRNA inhibition of KLF5. In addition, intercellular adhesion molecule-1 (ICAM-1) levels were increased in LPS-treated IEC6 cells and this increase was associated with increased adhesion of Jurkat lymphocytes to IEC6. The induction of ICAM-1 expression and T cell adhesion to IEC6 by LPS were both abrogated by siRNA inhibition of KLF5. These results indicate that KLF5 is an important mediator for the proinflammatory response elicited by LPS in intestinal epithelial cells

Translocation as a Population Restoration Technique for Northern Bobwhites: A Review and Synthesis

Northern bobwhite (Colinus virginianus) abundance has declined precipitously for decades across much of the species range, to the point of widespread local, regional, and statewide extirpation. Because of successful translocations of other gallinaceous birds, bobwhite enthusiasts increasingly call for use of the approach. Consequently, the National Bobwhite Technical Committee (NBTC), on behalf of state agencies, requested a review and recommendation by the NBTC Science Subcommittee. Thus, our paper is co-authored by invited experts and includes reviews of peer-reviewed publications, manuscripts in these proceedings, state agency reports, experience by co-authors, and a survey of perspectives on translocations by state wildlife agency members of the NBTC. We discuss the state of science on key aspects of bobwhite conservation, offer best management practices (BMPs) for using translocation as a potential bobwhite restoration technique, and suggest ways to reduce uncertainty about implementation. We note that although conservationists operate on a relatively solid foundation of improving bobwhite abundance via increased quantity, connectivity, and quality of habitat, population restoration success to- date is relatively rare and unpredictable. Similarly, some past translocations have been unreliable with an abundance of failures and inadequate experimental designs. We conclude that because of major uncertainties regarding habitat, population phenomena (e.g., Allee effect) and restoration techniques, outcomes of translocations remain unpredictable; thus, future efforts must be a part of sound and rigorous peer-reviewed research. To improve scientific efforts, we recommend the following BMPs for future translocations: (1) target bobwhite abundance should be \u3e800 post-translocation which will likely necessitate ≥600 ha of suitable and accessible habitat while a larger (e.g., \u3e800 ha) area will be needed in areas with lower carrying capacity and when sites are highly fragmented or isolated, (2) personnel should identify and avoid stressors to bobwhites in all phases of the translocation process (i.e., capture, holding, transportation, and release), (3) source populations should be disease free and from similar environments and latitude; preferably from the nearest suitable source, (4) conspecifics should be present on recipient sites (5) birds should be released just before the breeding season (i.e., March or April), and (6) the translocation should incorporate robust short- and long-term bird (i.e., abundance and/or density) and habitat monitoring efforts (i.e., the Coordinated Implementation Program (CIP) of the National Bobwhite Conservation Initiative (NBCI)). In conclusion, we note that translocation of bobwhites is not a panacea for broad scale restoration of bobwhites; however, the technique should remain at the forefront of bobwhite science, taking into account knowledge of the species’ life history and ecology, so that a practical and reliable solution can be developed. We recognize this paper is just the beginning of vigorous debate, testing of concepts, and on-the ground implementation of successful bobwhite conservation