Time to Integrate to Nest Test Evaluation in a Mouse DSS-Colitis Model

<div><p>Severity assessment in laboratory animals is an important issue regarding the implementation of the 3R concept into biomedical research and pivotal in current EU regulations. In mouse models of inflammatory bowel disease severity assessment is usually undertaken by clinical scoring, especially by monitoring reduction of body weight. This requires daily observance and handling of each mouse, which is time consuming, stressful for the animal and necessitates an experienced observer. The time to integrate to nest test (TINT) is an easily applicable test detecting disturbed welfare by measuring the time interval mice need to integrate nesting material to an existing nest. Here, TINT was utilized to assess severity in a mouse DSS-colitis model. TINT results depended on the group size of mice maintained per cage with most consistent time intervals measured when co-housing 4 to 5 mice. Colitis was induced with 1% or 1.5% DSS in group-housed WT and <i>Cd14</i>-deficient mice. Higher clinical scores and loss of body weight were detected in 1.5% compared to 1% DSS treated mice. TINT time intervals showed no dose dependent differences. However, increased clinical scores, body weight reductions, and increased TINT time intervals were detected in <i>Cd14</i>^<i>-/-</i> compared to WT mice revealing mouse strain related differences. Therefore, TINT is an easily applicable method for severity assessment in a mouse colitis model detecting CD14 related differences, but not dose dependent differences. As TINT revealed most consistent results in group-housed mice, we recommend utilization as an additional method substituting clinical monitoring of the individual mouse.</p></div

Clinical colitis scoring.

<p>Clinical colitis scoring.</p

Group size effect on TINT reliance.

<p>TINT time intervals determined in untreated WT mice on three consecutive days. A group size of 4 to 5 mice per cage resulted in consistent time intervals.</p

Intestinal inflammation induced by DSS-treatment.

<p>Hematoxylin and eosin staining of colon tissue obtained from (A-D) wild-type and (E-H) <i>Cd14</i>-deficient mice treated with 1% DSS for seven days (C-D and G-H, respectively). Untreated controls (A-B and E-F) did not show any signs of inflammation. Colitis was characterized by the presence of mixed cell infiltrates, hyperplasia, abnormal crypt architecture, edema and erosions (see boxed magnifications D and H). Original magnification 5x and 10x. Histological score quantifying the alterations observed in the colon (J).</p

Clinical disease activity score after DSS treatment.

<p>Assessment of severity in controls (A, B) and DSS treated mice (1% DSS C, D and 1.5% DSS E, F) determined by an overall clinical disease activity score (A, C, E) and specifically by the change in body weight (B, D, F). Untreated controls exhibited low clinical scores (A) and a steady body weight (B). Mice treated with 1% (C, D) or 1.5% DSS (E, F) demonstrated increasing clinical scores and loss of body weight. <i>Cd14</i>^<i>-/-</i> mice showed significantly higher clinical scores and a significantly higher reduction of body weight than WT mice.</p

Histology score.

<p>Histology score.</p

Colitis severity assessment by utilizing TINT.

<p>TINT time intervals determined in controls (A) and 1% (B) as well as 1.5% DSS treated mice (C). TINT time intervals were significantly increased in 1% DSS treated <i>Cd14</i>^<i>-/-</i> mice compared to WT mice on day 7 post colitis-induction (B).</p

Data_Sheet_1_Grimace scale, burrowing, and nest building for the assessment of post-surgical pain in mice and rats—A systematic review.docx

Several studies suggested an informative value of behavioral and grimace scale parameters for the detection of pain. However, the robustness and reliability of the parameters as well as the current extent of implementation are still largely unknown. In this study, we aimed to systematically analyze the current evidence-base of grimace scale, burrowing, and nest building for the assessment of post-surgical pain in mice and rats. The following platforms were searched for relevant articles: PubMed, Embase via Ovid, and Web of Science. Only full peer-reviewed studies that describe the grimace scale, burrowing, and/or nest building as pain parameters in the post-surgical phase in mice and/or rats were included. Information about the study design, animal characteristics, intervention characteristics, and outcome measures was extracted from identified publications. In total, 74 papers were included in this review. The majority of studies have been conducted in young adult C57BL/6J mice and Sprague Dawley and Wistar rats. While there is an apparent lack of information about young animals, some studies that analyzed the grimace scale in aged rats were identified. The majority of studies focused on laparotomy-associated pain. Only limited information is available about other types of surgical interventions. While an impact of surgery and an influence of analgesia were rather consistently reported in studies focusing on grimace scales, the number of studies that assessed respective effects was rather low for nest building and burrowing. Moreover, controversial findings were evident for the impact of analgesics on post-surgical nest building activity. Regarding analgesia, a monotherapeutic approach was identified in the vast majority of studies with non-steroidal anti-inflammatory (NSAID) drugs and opioids being most commonly used. In conclusion, most evidence exists for grimace scales, which were more frequently used to assess post-surgical pain in rodents than the other behavioral parameters. However, our findings also point to relevant knowledge gaps concerning the post-surgical application in different strains, age levels, and following different surgical procedures. Future efforts are also necessary to directly compare the sensitivity and robustness of different readout parameters applied for the assessment of nest building and burrowing activities.</p