In vivo characterization of minipig skin as a model for dermatological research using multiphoton microscopy.

Minipig skin is one of the most widely used non-rodent animal skin models for dermatological research. A thorough characterization of minipig skin is essential for gaining deeper understanding of its structural and functional similarities with human skin. In this study, three-dimensional (3-D) in vivo images of minipig skin was obtained non-invasively using a multimodal optical imaging system capable of acquiring two-photon excited fluorescence (TPEF) and fluorescence lifetime imaging microscopy (FLIM) images simultaneously. The images of the structural features of different layers of the minipig skin were qualitatively and quantitatively compared with those of human skin. Label-free imaging of skin was possible due to the endogenous fluorescence and optical properties of various components in the skin such as keratin, nicotinamide adenine dinucleotide phosphate (NAD(P)H), melanin, elastin, and collagen. This study demonstrates the capability of optical biopsy techniques, such as TPEF and FLIM, for in vivo non-invasive characterization of cellular and functional features of minipig skin, and the optical image-based similarities of this commonly utilized model of human skin. These optical imaging techniques have the potential to become promising tools in dermatological research for developing a better understanding of animal skin models, and for aiding in translational pre-clinical to clinical studies.

Characterization of cecal gene expression in a differentially susceptible mouse model of bacterial-induced inflammatory bowel disease.

BACKGROUND: A/JCr mice develop typhlitis in response to Helicobacter hepaticus infection, whereas C57BL/6 mice coexist with this bacterium in a "commensal" relationship and do not develop disease even during prolonged colonization. METHODS: To determine mechanisms that control this balance between responsiveness and nonresponsiveness, the mucosal response of A/JCr and C57BL/6 mice to acute H. hepaticus colonization was evaluated using genome-wide profiling. Transcription levels for a subset of gene discoveries were then evaluated longitudinally by semiquantitative real-time reverse-transcriptase polymerase chain reaction (RT-PCR) to identify changes in gene expression that occur during progression from the acute to chronic phase of colonization. To determine whether chronic mucosal inflammation in A/JCr mice was mediated through a Th1 mechanism, as was inferred from the gene expression data, mice with typhlitis were treated with neutralizing antibody targeting IL-12/23p40 or IFN-gamma and the response to treatment was determined by cecal lesion severity and transcription of disease-related genes. RESULTS: A/JCr mice had a biphasic expression of proinflammatory genes that corresponded with the acute and chronic phases of disease. In contrast, C57BL/6 mice exhibited a less robust acute transcriptional response that waned by day 30 postinoculation. Sustained upregulation of proinflammatory signals and responsiveness to anti-IL-12/23p40 and anti-IFN-gamma antibody suggests that inflammation in A/JCr mice was mediated through a Th1 mechanism. Prolonged upregulation of SOCS3 during the acute response to colonization suggests that C57BL/6 mice maintain mucosal homeostasis, at least in part by attenuating responsiveness to cytokine signaling. CONCLUSIONS: Collectively, these findings provide a foundation for understanding the immunological mechanisms that confer resistance or susceptibility to H. hepaticus-induced typhlitis.

Cross-Training Laboratory Animal Care Personnel in Physically Separate Animal Facilities at a Land-Grant Institution.

The University of Illinois at Urbana-Champaign maintains physically separated animal care facilities under centralized management by the Division of Animal Resources. As part of a land-grant institution, the animal care and use program operates several animal units in key locations for specific disciplines within the campus, all of which have the core mission to teach, conduct research, and engage in public service. Populations of research animals vary with the levels of research funding, the number of research investigators on staff, research direction, and animal availability. Accordingly, the requirement for animal care staffing in each unit may vary widely also. To best use the existing animal care staff and remain fiscally responsible, cross-training of staff was implemented to allow staff to travel from units with small animal populations to units with larger populations or short-term staffing shortages. Here we detail and describe the system we used to assess the needs for cross-training, identify the staff to train, and implement the training plan. We believe this information will assist other programs, particularly those with large or complex organization (for example, land-grant institutions) that experience similar fluctuations in animal use.

Sex influence on chronic intestinal inflammation in Helicobacter hepaticus-infected A/JCr mice.

Helicobacter hepaticus is a bacterial pathogen of mice that has been reported to cause chronic intestinal inflammation in A/JCr, germfree Swiss Webster, and immunodeficient mice. To the authors' knowledge, the influence of sex on development of chronic intestinal inflammation in H. hepaticus-infected mice has not been investigated. The purposes of the study reported here were to determine whether severity of intestinal inflammation differs between male and female A/JCr mice chronically infected with H. hepaticus and to characterize the mucosal immune response in these mice. The cecum of male and female A/JCr mice infected with H. hepaticus for 1 month and 3 months was objectively evaluated histologically for intestinal disease. Also, semi-quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) analysis was done to measure interferon-gamma (IFN-gamma), tumor necrosis factor-alpha (TNF-alpha), interleukin 4 (IL-4), IL-10, macrophage inflammatory protein-1alpha (MIP-1alpha), interferon-inducible protein of 10 kDa (IP-10), and monokine induced by gamma interferon (MIG) mRNA values in the cecal tissue of these mice. Significant differences in cecal lesion scores were not present at 1 month after infection. However, infected female mice had significantly up-regulated expression of cecal IL-10, MIP-1alpha, IP-10, and MIG mRNA compared with that in uninfected females, and expression of IL-10 and MIP-1alpha was significantly greater than that detected in infected male mice (P < or = 0.05). At 3 months after infection, cecal lesion scores were significantly (P < or = 0.05) increased in female and male mice compared with uninfected controls, and infected female mice had significantly (P < or = 0.05) higher cecal lesion scores than did infected male mice. In addition, infected females had significant (P < or = 0.05) increases in cecal IFN-gamma, TNF-alpha, IL-10, MIP-1alpha, IP-10, and MIG mRNA values compared with values in uninfected females and infected males, and male mice had significant (P < or = 0.05) increases in cecal TNF-alpha and IL-10 mRNA values compared with those for male control mice. These data indicate that, in H. hepaticus-infected A/JCr mice, females develop more severe intestinal inflammation than do males, and the chronic mucosal inflammation is polarized toward a Th1 response that is not down-regulated by increased activity of IL-10. We propose that H. hepaticus-infected A/JCr mice will serve as a good animal model with which to study the influence of sex on bacterial-induced mucosal inflammation.

Cage change intervals for opossums (Monodelphis domestica) in individually ventilated cages.

The opossum Monodelphis domestica is the most commonly used marsupial in biomedical research. At our institution, these opossums are housed in polycarbonate (35.6 cm × 25.4 cm × 17.8 cm) individually ventilated cages. Previous studies of the cage microenvironment of rodents housed in individually ventilated cages have demonstrated that the cage-change frequency could be extended from 7 to 14 d, without detriment to the animals' wellbeing. We sought to determine whether the cage change frequency for opossums housed in individually ventilated cages could be extended to 14 d. Opossums were placed into 3 experimental groups: singly housed males, singly housed females, and females housed with litters. The 14-d testing period was repeated twice, with temperature, relative humidity, and ammonia levels tested on days 0, 7, and 14. Acceptable ranges for the cage microenvironment were based on standards followed by our institution for housing rodents: temperature between 22 to 26 °C, relative humidity between 30% to 70%, and ammonia less than 25 ppm. Throughout both 14-d testing periods, temperature, relative humidity, and ammonia levels for singly housed male and singly housed female opossums were within acceptable ranges. However, ammonia levels exceeded the recommended 25 ppm on day 7 of both testing periods for female opossums housed with litters. In summary, the cage-change frequency for a singly housed opossum in an individually ventilated cage can be extended to 14 d.

Analysis of gene expression in ceca of Helicobacter hepaticus-infected A/JCr mice before and after development of typhlitis.

The inflammatory bowel diseases, Crohn's disease and ulcerative colitis, are chronic inflammatory disorders of the gastrointestinal tract. The causes of these diseases remain unknown; however, prevailing theories suggest that chronic intestinal inflammation results from a dysregulated immune response to ubiquitous bacterial antigens. While a substantial body of data has been amassed describing the role of the adaptive immune system in perpetuating and sustaining inflammation, very little is known about the early signals, prior to the development of inflammation, that initiate and direct the abnormal immune response. To this end, we characterized the gene expression profile of A/JCr mice with Helicobacter hepaticus-induced typhlitis at month 1 of infection, prior to the onset of histologic disease, and month 3 of infection, after chronic inflammation is fully established. Analysis of the gene expression in ceca of H. hepaticus infected mice revealed 25 up-regulated and 3 down-regulated genes in the month-1 postinoculation group and 31 up-regulated and 2 down-regulated genes in the month-3 postinoculation group. Among these was a subset of immune-related genes, including interferon-inducible protein 10, monokine induced by gamma interferon, macrophage-induced protein 1 alpha, and serum amyloid A1. Semiquantitative real-time reverse transcriptase PCR confirmed the increased expression levels of these genes, as well as elevated expression of gamma interferon. To our knowledge, this is the first report profiling cecal gene expression in H. hepaticus-infected A/JCr mice. The findings of altered gene expression prior to the development of any features of pathology and the ensuing chronic disease course make this an attractive model for studying early host response to microbe-induced inflammatory bowel disease.