A protein kinase C α and ß inhibitor blunts hyperphagia to halt renal function decline and reduces adiposity in a rat model of obesity-driven type 2 diabetes.

Type 2 diabetes (T2D) and its complications can have debilitating, sometimes fatal consequences for afflicted individuals. The disease can be difficult to control, and therapeutic strategies to prevent T2D-induced tissue and organ damage are needed. Here we describe the results of administering a potent and selective inhibitor of Protein Kinase C (PKC) family members PKCα and PKCß, Cmpd 1, in the ZSF1 obese rat model of hyperphagia-induced, obesity-driven T2D. Although our initial intent was to evaluate the effect of PKCα/ß inhibition on renal damage in this model setting, Cmpd 1 unexpectedly caused a marked reduction in the hyperphagic response of ZSF1 obese animals. This halted renal function decline but did so indirectly and indistinguishably from a pair feeding comparator group. However, above and beyond this food intake effect, Cmpd 1 lowered overall animal body weights, reduced liver vacuolation, and reduced inguinal adipose tissue (iWAT) mass, inflammation, and adipocyte size. Taken together, Cmpd 1 had strong effects on multiple disease parameters in this obesity-driven rodent model of T2D. Further evaluation for potential translation of PKCα/ß inhibition to T2D and obesity in humans is warranted.

Development of a CD8 co-receptor independent T-cell receptor specific for tumor-associated antigen MAGE-A4 for next generation T-cell-based immunotherapy.

BACKGROUND: The cancer-testis antigen MAGE-A4 is an attractive target for T-cell-based immunotherapy, especially for indications with unmet clinical need like non-small cell lung or triple-negative breast cancer. METHODS: An unbiased CD137-based sorting approach was first used to identify an immunogenic MAGE-A4-derived epitope (GVYDGREHTV) that was properly processed and presented on human leukocyte antigen (HLA)-A2 molecules encoded by the HLA-A*02:01 allele. To isolate high-avidity T cells via subsequent multimer sorting, an in vitro priming approach using HLA-A2-negative donors was conducted to bypass central tolerance to this self-antigen. Pre-clinical parameters of safety and activity were assessed in a comprehensive set of in vitro and in vivo studies. RESULTS: A MAGE-A4-reactive, HLA-A2-restricted T-cell receptor (TCR) was isolated from primed T cells of an HLA-A2-negative donor. The respective TCR-T-cell (TCR-T) product bbT485 was demonstrated pre-clinically to have a favorable safety profile and superior in vivo potency compared with TCR-Ts expressing a TCR derived from a tolerized T-cell repertoire to self-antigens. This natural high-avidity TCR was found to be CD8 co-receptor independent, allowing effector functions to be elicited in transgenic CD4+ T helper cells. These CD4+ TCR-Ts supported an anti-tumor response by direct killing of MAGE-A4-positive tumor cells and upregulated hallmarks associated with helper function, such as CD154 expression and release of key cytokines on tumor-specific stimulation. CONCLUSION: The extensive pre-clinical assessment of safety and in vivo potency of bbT485 provide the basis for its use in TCR-T immunotherapy studies. The ability of this non-mutated high-avidity, co-receptor-independent TCR to activate CD8+ and CD4+ T cells could potentially provide enhanced cellular responses in the clinical setting through the induction of functionally diverse T-cell subsets that goes beyond what is currently tested in the clinic.

Cellular and functional actions of tofacitinib related to the pathophysiology of hibernoma development.

Tofacitinib is an oral JAK inhibitor for the treatment of rheumatoid arthritis. In the 2-year carcinogenicity study with tofacitinib, increased incidence of hibernoma (a neoplasm of brown adipose tissue [BAT]) was noted in female rats at ≥30 mg/kg/day (≥41x human exposure multiples). Thus, signaling pathways within BAT were investigated by measuring BAT: weight, cell proliferation biomarkers, content of basal and prolactin-induced phosphorylated Signal Transducer and Activator of Transcription (STAT), and uncoupling protein 1 (UCP-1). The relationship between cardiovascular hemodynamics and plasma norepinephrine (NE) levels was also investigated. Tofacitinib administered to female rats at doses of 10, 30, or 75 mg/kg/day for 14 days increased BAT weight at 75 mg/kg/day and cell proliferation at ≥30 mg/kg/day. JAK inhibition, observed as lower pSTAT3 and pSTAT5 in BAT, was noted at ≥10 mg/kg/day, while lower activity of BAT was observed as lower UCP-1 protein at ≥30 mg/kg/day. In cultured brown adipocytes, prolactin-induced increase in pSTAT5 and pSTAT3 were inhibited by tofacitinib in a concentration-dependent manner. Tofacitinib lowered blood pressure, increased heart rate, and resulted in dose-dependent increases in circulating NE. Thus, JAK/STAT inhibition in BAT and sympathetic stimulation are two factors which might contribute to the genesis of hibernomas by tofacitinib in rats.

Regulatory Forum Commentary* Counterpoint: Dose Selection for rasH2 Mouse Carcinogenicity Studies.

Dose selection for the 6-month rasH2 mouse carcinogenicity studies depends heavily on the maximum tolerated dose (MTD) obtained from 1-month range-finding studies. A retrospective evaluation of range-finding studies and pivotal 6 month rasH2 mouse studies for 11 compounds demonstrated that the MTD based on at least a 10% decrease in body weight gain, mortality, and target organ toxicity in range-finding studies appropriately identified high doses for pivotal studies for 8 of 11 compounds. Two of the selected high doses were based on decreased body weight gain alone, while 7 were based on mortality at higher doses in shorter duration range-finding studies. High-dose selection was based on the maximum feasible dose for one study. The Center for Drug Evaluation and Research, U.S. Food and Drug Administration Executive Carcinogenicity Assessment Committee often suggested different doses than those proposed by the sponsor. High mortality was observed in only one pivotal study and the high dose was lowered during the course of that study.

Two-year carcinogenicity study in rats with a nonnucleoside reverse transcriptase inhibitor.

Administration of lersivirine, a nonnucleotide reverse transcriptase inhibitor, daily by oral gavage to Sprague-Dawley rats for up to 2 yr was associated with decreased survival, decreased body weights, and an increase in neoplasms and related proliferative lesions in the liver, thyroid, kidney, and urinary bladder. Thyroid follicular adenoma and carcinoma, the associated thyroid follicular hypertrophy/hyperplasia, hepatocellular adenoma/adenocarcinoma, altered cell foci, and hepatocellular hypertrophy were consistent with lersivirine-related induction of hepatic microsomal enzymes. Renal tubular adenoma and renal tubular hyperplasia were attributed to the lersivirine-related exacerbation of chronic progressive nephropathy (CPN), while urinary bladder hyperplasia and transitional cell carcinoma in the renal pelvis and urinary bladder were attributed to urinary calculi. Renal tubular neoplasms associated with increased incidence and severity of CPN, neoplasms of transitional epithelium attributed to crystalluria, and thyroid follicular and hepatocellular neoplasms related to hepatic enzyme induction have low relevance for human risk assessment.

Toxicities associated with 1-month treatment with propylthiouracil (PTU) and methimazole (MMI) in male rats.

Thionamides such as propylthiouracil (PTU) and methimazole (MMI) have been used for more than 50 years to treat the more common causes of thyrotoxicosis/hyperthyroidism such as Graves' disease. Serious adverse effects associated with thionamides in humans include idiosyncratic liver damage, agranulocytosis, aplastic anemia, and vasculitis. Both prospective and retrospective clinical studies with these drugs have failed to identify predictive biomarker for these adverse effects. To assess whether rat is a good model for predicting drug-related adverse events in the liver and in the bone marrow, we conducted a comprehensive study in male rats with multiple doses of PTU and MMI. As expected, euthyroid animals became hypothyroid along with several secondary changes associated with hypothyroidism. There were slight reductions in red blood cell parameters along with some marginal effects on the bone marrow elements. However, there was no evidence of significant neutropenia and liver injury in both PTU-treated and MMI-treated cohorts. MMI-related effects were noted in the seminiferous tubules of the testes. Overall, 1-month daily treatment of euthyroid rats with PTU or MMI resulted in hypothyroidism, minor bone marrow effects, and several secondary effects associated with hypothyroidism, but without any evidence of adverse effects reported in humans including liver injury and agranulocytosis.

Regulatory Forum commentary: alternative mouse models for future cancer risk assessment.

International regulatory and pharmaceutical industry scientists are discussing revision of the International Conference on Harmonisation of Technical Requirements for Registration of Pharmaceuticals for Human Use (ICH) S1 guidance on rodent carcinogenicity assessment of small molecule pharmaceuticals. A weight-of-evidence approach is proposed to determine the need for rodent carcinogenicity studies. For compounds with high human cancer risk, the product may be labeled appropriately without conducting rodent carcinogenicity studies. For compounds with minimal cancer risk, only a 6-month transgenic mouse study (rasH2 mouse or p53+/- mouse) or a 2-year mouse study would be needed. If rodent carcinogenicity testing may add significant value to cancer risk assessment, a 2-year rat study and either a 6-month transgenic mouse or a 2-year mouse study is appropriate. In many cases, therefore, one rodent carcinogenicity study could be sufficient. The rasH2 model predicts neoplastic findings relevant to human cancer risk assessment as well as 2-year rodent models, produces fewer irrelevant neoplastic outcomes, and often will be preferable to a 2-year rodent study. Before revising ICH S1 guidance, a prospective evaluation will be conducted to test the proposed weight-of-evidence approach. This evaluation offers an opportunity for a secondary analysis comparing the value of alternative mouse models and 2-year rodent studies in the proposed ICH S1 weight-of-evidence approach for human cancer risk assessment.

The rasH2 mouse model for assessing carcinogenic potential of pharmaceuticals.

A factor limiting widespread use of the transgenic rasH2 mouse model for carcinogenicity testing of pharmaceuticals is the paucity of published data on actual drug candidates in rasH2 mice. This report addresses this gap by highlighting rasH2 mouse study data for 10 pharmaceutical candidates. These results were compared with findings in the 2-year studies in Sprague-Dawley rats for the same 10 compounds. In the 6-month rasH2 studies, only 2 of the 10 compounds tested positive for carcinogenicity and these correlated with positive findings in the companion 2-year rat studies. One compound, sunitinib, produced gastroduodenal carcinoma in both sexes and increased hemangiosarcoma in spleen and uterus in female rasH2 mice; in rats it produced gastroduodenal carcinoma and increased pheochromocytoma (males only). The second compound, bazedoxifene, produced ovarian granulosa cell neoplasms in rasH2 mice and rats, and renal tubular neoplasms associated with increased chronic progressive nephropathy only in rats. The higher percentage of carcinogenicity positive rat bioassays could be attributed to rat-specific phenomena with little or low relevance to man. Thus, this article confirms previous reports that rasH2 mice develop rodent-specific neoplasms less frequently than rats and positive findings, when present, are accompanied by similar positive results in the rat.

Expression of TMPRSS4 in non-small cell lung cancer and its modulation by hypoxia.

Overexpression of TMPRSS4, a cell surface-associated transmembrane serine protease, has been reported in pancreatic, colorectal and thyroid cancers, and has been implicated in tumor cell migration and metastasis. Few reports have investigated both TMPRSS4 gene expression levels and the protein products. In this study, quantitative RT-PCR and protein staining were used to assess TMPRSS4 expression in primary non-small cell lung carcinoma (NSCLC) tissues and in lung tumor cell lines. At the transcriptional level, TMPRSS4 message was significantly elevated in the majority of human squamous cell and adenocarcinomas compared with normal lung tissues. Staining of over 100 NSCLC primary tumor and normal specimens with rabbit polyclonal anti-TMPRSS4 antibodies confirmed expression at the protein level in both squamous cell and adenocarcinomas with little or no staining in normal lung tissues. Human lung tumor cell lines expressed varying levels of TMPRSS4 mRNA in vitro. Interestingly, tumor cell lines with high levels of TMPRSS4 mRNA failed to show detectable TMPRSS4 protein by either immunoblotting or flow cytometry. However, protein levels were increased under hypoxic culture conditions suggesting that hypoxia within the tumor microenvironment may upregulate TMPRSS4 protein expression in vivo. This was supported by the observation of TMPRSS4 protein in xenograft tumors derived from the cell lines. In addition, staining of human squamous cell carcinoma samples for carbonic anhydrase IX (CAIX), a hypoxia marker, showed TMPRSS4 positive cells adjacent to CAIX positive cells. Overall, these results indicate that the cancer-associated TMPRSS4 protein is overexpressed in NSCLC and may represent a potential therapeutic target.

Spontaneous tumor incidence in rasH2 mice: review of internal data and published literature.

Alternate transgenic mouse models are accepted as replacements for the standard carcinogenicity mouse bioassay by regulatory agencies with a companion 2-year rat bioassay. The slower rate of industry acceptance of these shorter transgenic mouse cancer bioassays has been due to lack of historical data and diagnostic criteria, and the use of nonstandardized terminologies in published data. To address these issues, especially that of generating a large historical database, a retrospective analysis of the spontaneous tumor incidences in rasH2 mice from internally sponsored 6-month carcinogenicity studies was compared to the published literature. Incidences of common spontaneous tumors (incidences > 1%) observed in these studies were lung bronchiolo-alveolar adenomas (mean 3.9-9.9%; range 0-18%), lung bronchiolo-alveolar adenocarcinomas (mean 1.4-2.4%; range 0-5%), splenic hemangiosarcomas (mean 3.0-3.9%; range 0-17%), cutaneous squamous cell papillomas (mean 1.1-1.2%; range 0-4%), Harderian gland adenoma (mean 0.8-1.2%; range 0-4%), and hepatocellular adenomas (mean 1.8%; 0-9% in males only). The remarkable similarity in the tumor incidences in multiple rasH2 studies over a decade and the observed stability of the inserted human gene are important indicators of the minimal drift in this model. Overall, the historical control data for spontaneous neoplasms should assist in the interpretation of future rasH2 mouse studies.

An "Omics" based survey of human colon cancer.

Despite an increased understanding of the molecular pathogenesis of colorectal cancer (CRC) during the past two decades, reliable and robust biomarkers to enable screening, surveillance, and primary prevention of this disease are lacking. CRC diagnosis and therapy remain dependent upon descriptive classification and staging systems, based primarily on morphology and histology. The traditional approach of understanding complex biological systems by studying smaller, discrete units of the whole system has been less fruitful for understanding complex diseases. The implicit assumption of traditional methods, that a single or even only a few factors, play a dominant role in a complex disease might be inadequate when studying multifactorial diseases such as cancer. The burgeoning field of systems biology adopts a holistic approach, wherein the integration of individual parts of the system is sought. The cornerstone of a systems biology approach has been the development of a variety of high-throughput "omics" sciences, including genomics, transcriptomics, proteomics, and metabolomics. This review will focus on the "omics" literature in the field of sporadic human CRC and present examples of how a systems approach has been extremely useful in understanding concepts that would have been difficult to develop using traditional methods.

Obesity and non-insulin-dependent diabetes mellitus in Swiss-Webster mice associated with late-onset hepatocellular carcinoma.

Genetic mutations resulting in obesity and type 2 diabetes mellitus (T2D) are described for both inbred and outbred mice. However, no known mouse model completely recapitulates human T2D and its comorbidities. We identified a cohort of obese, male, outbred Swiss-Webster (SW) mice as polyuric, polydipsic, glucosuric, and hyperglycemic. Prevalence of glucosuria in the SW colony reached 60% (n=70) in males 8 weeks to 6 months of age. Despite severe obesity in some females, no females were diabetic. Pathologic findings in affected males included cachexia, dilated gastrointestinal tracts with poor muscular tone, pancreatic islet degeneration and atrophy with compensatory metaplasia and/or neogenesis, bacterial pyelonephritis, membranous glomerulopathy, and late-onset hepatic tumors with macrosteatosis, microsteatosis, and hydropic change in aged males. Serum insulin correlated with blood glucose in a nonlinear pattern, suggestive of islet exhaustion. Circulating leptin levels showed a weak inverse correlation with glucose. Diabetic males were bred with obese colony females to produce 20 male and 20 female offspring. Prevalence of diabetes in male offspring was 80% (16/20) with a median age of onset of 18 weeks. By contrast, no diabetic females were identified, despite being significantly more obese than males. Male predominance is likewise a feature of T2D in humans. To our knowledge, this is the first documentation of hepatocellular carcinoma and islet metaplasia and/or neogenesis in a spontaneous outbred mouse model of T2D. The SW availability and histopathologic features represent a promising new model for the study of T2D.

Diarrhea as a cause of mortality in a mouse model of infectious colitis.

BACKGROUND: Comparative characterization of genome-wide transcriptional changes during infection can help elucidate the mechanisms underlying host susceptibility. In this study, transcriptional profiling of the mouse colon was carried out in two cognate lines of mice that differ in their response to Citrobacter rodentium infection; susceptible inbred FVB/N and resistant outbred Swiss Webster mice. Gene expression in the distal colon was determined prior to infection, and at four and nine days post-inoculation using a whole mouse genome Affymetrix array. RESULTS: Computational analysis identified 462 probe sets more than 2-fold differentially expressed between uninoculated resistant and susceptible mice. In response to C. rodentium infection, 5,123 probe sets were differentially expressed in one or both lines of mice. Microarray data were validated by quantitative real-time RT-PCR for 35 selected genes and were found to have a 94% concordance rate. Transcripts represented by 1,547 probe sets were differentially expressed between susceptible and resistant mice regardless of infection status, a host effect. Genes associated with transport were over-represented to a greater extent than even immune response-related genes. Electrolyte analysis revealed reduction in serum levels of chloride and sodium in susceptible animals. CONCLUSION: The results support the hypothesis that mortality in C. rodentium-infected susceptible mice is associated with impaired intestinal ion transport and development of fatal fluid loss and dehydration. These studies contribute to our understanding of the pathogenesis of C. rodentium and suggest novel strategies for the prevention and treatment of diarrhea associated with intestinal bacterial infections.

Genetic deletion of mPGES-1 suppresses intestinal tumorigenesis.

Elevated levels of prostaglandin E(2) (PGE(2)) are often found in colorectal cancers. Thus, nonsteroidal anti-inflammatory drugs, including selective cyclooxygenase-2 (COX-2) inhibitors, are among the most promising chemopreventive agents for colorectal cancer. However, their long-term use is restricted by the occurrence of adverse events believed to be associated with a global reduction in prostaglandin production. In the present study, we evaluated the chemopreventive efficacy of targeting the terminal synthase microsomal PGE(2) synthase 1 (mPGES-1), which is responsible for generating PGE(2), in two murine models of intestinal cancer. We report for the first time that genetic deletion of mPGES-1 in Apc-mutant mice results in marked and persistent suppression of intestinal cancer growth by 66%, whereas suppression of large adenomas (>3 mm) was almost 95%. This effect occurred despite loss of Apc heterozygosity and beta-catenin activation. However, we found that mPGES-1 deficiency was associated with a disorganized vascular pattern within primary adenomas as determined by CD31 immunostaining. We also examined the effect of mPGES-1 deletion on carcinogen-induced colon cancer. The absence of mPGES-1 reduced the size and number of preneoplastic aberrant crypt foci (ACF). Importantly, mPGES-1 deletion also blocked the nuclear accumulation of beta-catenin in ACF, confirming that beta-catenin is a critical target of PGE(2) procarcinogenic signaling in the colon. Our data show the feasibility of targeting mPGES-1 for cancer chemoprevention with the potential for improved tolerability over traditional nonsteroidal anti-inflammatory drugs and selective COX-2 inhibitors.

Infestation of wild-caught American bullfrogs (Rana catesbeiana) by multiple species of metazoan parasites.

The American bullfrog (Rana catesbeiana) is an aquatic, carnivorous member of the family Ranidae that is used extensively in physiology education programs and in various physiology, toxicology, sensorineural, and genetics research. Eleven bullfrogs purchased from a vendor distributing wild-caught frogs for use in a physiology research protocol were emaciated but otherwise showed no apparent clinical signs of illness. Necropsies performed on selected emaciated frogs indicated heavy infestation with multiple species of endoparasites. Identified helminths included Gorgodera amplicava, Haematolechus breviplexus, Clinostomum spp, Contracaecum spp, Cosmocercoides dukae, and Eustrongyloides spp. Grossly, parasitized bullfrogs showed encysted trematode larvae within skeletal muscle, nematode impaction of the intestinal tract, and lack of coelemic fat stores. Histopathologic lesions were restricted primarily to the gastrointestinal tract and consisted of parasitic granulomas associated with Contracaecum spp. The parasitic lesions may have been associated with the poor body condition of the bullfrogs. Food crickets maintained in-house were negative for parasite larvae or ova. Heavy parasitism of wild-caught bullfrogs may confound research protocols and markedly impair animal health. We encourage researchers to purchase laboratory-bred and -reared bullfrogs and to routinely monitor the parasite status of colony frogs.

Agammaglobulinemia and Staphylococcus aureus botryomycosis in a cohort of related sentinel Swiss Webster mice.

Sentinel mouse seroconversion to infectious agents is critical for laboratory animal facility disease monitoring. We report spontaneous emergence of non-sex-linked agammaglobulinemia with B-cell deficiency and cutaneous Staphylococcus aureus granulomatosis (botryomycosis) in a cohort of related Swiss Webster sentinel mice. Our experience reinforces the importance of immunocompetency validation in surveillance programs.

Rapid reversal of interleukin-6-dependent epithelial invasion in a mouse model of microbially induced colon carcinoma.

Chronic inflammation of mucosal surfaces renders them increasingly susceptible to epithelial cancers both in humans and mice. We have previously shown that anti-inflammatory CD4(+)CD45RB(lo)CD25(+) regulatory (Treg or T(R)) lymphocytes down-regulate inflammation and block development of bacteria-triggered colitis and colorectal cancer (CRC) in 129/SvEv Rag2-/- mice. Interestingly, T(R) cells collected from Interleukin (IL)-10-deficient cell donors not only failed to suppress carcinogenesis but instead promoted invasive mucinous colonic carcinoma with a strong gender bias expressing in male mice. We found we show that peritoneal invasion in this model is dependent on pleiotropic cytokine IL-6. Mucinous carcinoma arose rapidly and consistently after treatment with IL10-/- T(R) cells, which were found to express Foxp3+ and localize throughout tumor tissue. Carcinogenesis was rapidly reversible with transfer of wild type IL10-competent T(R) cells. Likewise, treatment with IL10-Ig fusion protein was sufficient to revert the lesions histologically, and restore inflammatory cytokine and oncogene expression to base line levels. These studies indicate an essential role for IL 6 in this CRC phenotype. Furthermore, immune-competent T(R) cells were important not only for preventing pathology but also for constructive remodeling of bowel following tumorigenic microbial insults. These data provide insights into etiopathogenesis of inflammation-associated epithelial invasion and maintenance of epithelial homeostasis.

Enterohepatic Helicobacter species are prevalent in mice from commercial and academic institutions in Asia, Europe, and North America.

The discovery of Helicobacter hepaticus and its role in hepatitis, hepatocellular carcinoma, typhlocolitis, and lower-bowel carcinoma in murine colonies was followed by the isolation and characterization of other Helicobacter spp. involved in enterohepatic disease. Colonization of mouse colonies with members of the family Helicobacteriaceae has become an increasing concern for the research community. From 2001 to 2005, shipments of selected gift mice from other institutions and mice received from specified commercial vendors were screened for Helicobacter spp. by culture of cecal tissue. The identities of the isolates were confirmed by genus-specific PCR, followed by species-specific PCR and restriction fragment length polymorphism analysis. Sequencing of the 16S rRNA gene was performed if the species identity was not apparent. The survey included 79 mice from 34 sources: 2 commercial sources and 16 research sources from the United States and 1 commercial source and 15 research sources from Canada, Europe, or Asia. Helicobacter spp. were cultured from the ceca of 62 of 79 mice. No Helicobacter spp. were found in mice from advertised Helicobacter-free production areas from two U.S. vendors. Multiple Helicobacter spp. were found in mice from one vendor's acknowledged Helicobacter-infected production area. The European commercial vendor had mice infected with novel Helicobacter sp. strain MIT 96-1001. Of the U.S. academic institutions, 6 of 16 (37%) had mice infected with Helicobacter hepaticus; but monoinfection with H. bilis, H. mastomyrinus, H. rodentium, and MIT 96-1001 was also encountered, as were mice infected simultaneously with two Helicobacter spp. Non-U.S. academic institutions had mice that were either monoinfected with H. hepaticus, monoinfected with seven other Helicobacter spp., or infected with a combination of Helicobacter spp. This survey indicates that 30 of 34 (88%) commercial and academic institutions in Canada, Europe, Asia, Australia, and the United States have mouse colonies infected with Helicobacter spp. Mice from 20 of the 34 institutions (59%) were most commonly colonized with H. hepaticus alone or in combination with other Helicobacter spp. These results indicate that a broad range of Helicobacter spp. infect mouse research colonies. The potential impact of these organisms on in vivo experiments continues to be an important issue for mice being used for biomedical research.

Development of fatal colitis in FVB mice infected with Citrobacter rodentium.

Citrobacter rodentium is the causative agent of transmissible murine colonic hyperplasia. The disease is characterized by severe but temporary epithelial hyperplasia with limited inflammation in the descending colon of adult mice on a variety of genetic backgrounds. The natural history of infection with this murine pathogen has been characterized in outbred Swiss Webster (SW) mice but not in the cognate inbred FVB strain. In contrast to subclinical infection in SW mice, 12-week-old FVB mice developed overt disease with significant weight loss and mortality beginning by 9 days postinoculation (dpi). By 21 dpi, more than 75% of infected FVB mice died or had to be euthanized, whereas no mortality developed in SW mice. Mortality in FVB mice was fully prevented by fluid therapy. Fecal shedding of bacteria was similar in both groups through 9 dpi; however, a slight but significant delay in bacterial clearance was observed in FVB mice by 12 to 18 dpi. SW mice developed hyperplasia with minimal inflammation in the descending colon. FVB mice developed epithelial cell hyperproliferation, severe inflammation with erosions and ulcers, and epithelial atypia by 6 dpi in the descending colon. In the majority of surviving FVB mice, colonic lesions, including epithelial atypia, were reversible, although a small percentage (5 to 7%) exhibited chronic colitis through 7 months postinoculation. The existence of susceptible and resistant lines of mice with similar genetic backgrounds will facilitate the identification of host factors responsible for the outcome of infection and may lead to the development of novel strategies for preventing and treating infectious colitis.