Section of Experimental Animal Models, Department of Veterinary and Animal Sciences, Ridebanevej 9, 1871, Frederiksberg C, Denmark.
GVG Genetic Monitoring GmbH, Deutscher Platz 5, 04103, Leipzig, Germany.
Sci Rep. 2022 Aug 12;12(1):13767. doi: 10.1038/s41598-022-17242-8.
Group sizes in an animal study are calculated from estimates on variation, effect, power and significance level. Much of the variation in glucose related parameters of the diet-induced obese (DIO) mouse model is due to inter-individual variation in gut microbiota composition. In addition, standard tandem repeats (STRs) in the non-coding DNA shows that inbred mice are not always homogenic. C57BL/6NTac (B6NTac) mice from Taconic and C57BL/6NRj (B6NRj) mice from Janvier Labs were fed a high calorie diet and treated with liraglutide. The fecal microbiota was sequenced before high-calorie feeding (time 1) and after diet-induced obesity instantly before liraglutide treatment (time 2) and mice were divided into clusters on the basis of their microbiota. Although liraglutide in both sub-strains alleviated glucose intolerance and reduced body weight, in a one-way ANOVA a borderline reduction in glycosylated hemoglobin (HbA1c) could only be shown in B6NTac mice. However, if the microbiota clusters from time 1 or time 2 were incorporated in a two-way ANOVA, the HbA1c effect was significant in B6NTac mice in both analyses, while this did not change anything in B6NRj mice. In a one-way ANOVA the estimated group size needed for a significant HbA1c effect in B6NTac mice was 42, but in two-way ANOVAs based upon microbiota clusters of time 1 or time 2 it was reduced to 21 or 12, respectively. The lowering impact on glucose tolerance was also powered by incorporation of microbiota clusters of both times in both sub-strains. B6NRj had up to six, while B6NTac had maximum three alleles in some of their STRs. In B6NRj mice in 28.8% of the STRs the most prevalent allele had a gene frequency less than 90%, while this was only 6.6% in the B6NTac mice. However, incorporation of the STRs with the highest number of alleles or the most even distribution of frequencies in two-way ANOVAs only had little impact on the outcome of data evaluation. It is concluded that the inclusion of microbiota clusters in a two-way ANOVA in the evaluation of the glucose related effects of an intervention in the DIO mouse model might be an efficient tool for increasing power and reducing group sizes in mouse sub-strains, if these have a microbiota, which influences these parameters.
在动物研究中，群组大小是根据变异、效应、功效和显著性水平的估计值来计算的。饮食诱导肥胖（DIO）小鼠模型中葡萄糖相关参数的大部分变异是由于肠道微生物群落组成的个体间变异所致。此外，非编码 DNA 中的标准串联重复（STR）表明，近交系小鼠并不总是同质的。Taconic 的 C57BL/6NTac（B6NTac）小鼠和 Janvier Labs 的 C57BL/6NRj（B6NRj）小鼠都喂食高热量饮食，并接受利拉鲁肽治疗。在高卡路里喂养前（时间 1）和饮食诱导肥胖后立即接受利拉鲁肽治疗前（时间 2）对粪便微生物群进行测序，并根据其微生物群将小鼠分为聚类。尽管两种亚系的利拉鲁肽都缓解了葡萄糖不耐受并降低了体重，但在单向方差分析中，只能在 B6NTac 小鼠中显示糖化血红蛋白（HbA1c）的边缘降低。然而，如果将时间 1 或时间 2 的微生物群聚类纳入双向方差分析，则在这两种分析中，B6NTac 小鼠的 HbA1c 效应均具有统计学意义，而在 B6NRj 小鼠中则没有任何改变。在单向方差分析中，B6NTac 小鼠中 HbA1c 效应具有统计学意义的估计组大小为 42，但在基于时间 1 或时间 2 的微生物群聚类的双向方差分析中，分别降至 21 或 12。在两种亚系中，通过纳入两次的微生物群聚类，对葡萄糖耐量的降低作用也具有功效。B6NRj 有多达 6 个，而 B6NTac 有多达 3 个等位基因在某些 STR 中。在 B6NRj 小鼠中，28.8%的 STR 中最常见的等位基因的基因频率低于 90%，而在 B6NTac 小鼠中则为 6.6%。然而，在双向方差分析中纳入具有最多等位基因或频率分布最均匀的 STR 对数据评估结果的影响很小。结论是，如果 DIO 小鼠模型中干预的葡萄糖相关效应的评估中纳入微生物群聚类，则对于具有影响这些参数的微生物群的近交系小鼠亚系，可能是增加功效和减少组大小的有效工具。
