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Urine pp 25-32 | Cite as

Comparison of Urinary Proteomes Among Three Animal Models

  • Na Ni
  • Jianqiang Wu
Chapter

Abstract

Biomarkers are the monitorable changes associated with physiological or pathological changes. Urine is not regulated by the homeostatic mechanism and can reflect multiple changes in the body. Animal models can simulate human disease processes, monitor disease changes, and provide clues for early diagnosis. In this chapter, clues are provided for the dominant model animals associated with disease selection by comparing the urine proteome of rats, guinea pigs, and golden hamsters. The peptides were cleaved by membrane digestion and analyzed by LC-MS/MS. The number of urine proteins in the three different animals was different, and also different in every system of the body. This provides a basis for selecting the best animal models for different diseases.

Keywords

Urine proteomics LC-MS/MS Animal model Rat Guinea pig Golden hamster 

3.1 Introduction

The most important finding for biomarkers is to look for detectable changes associated with physiological and pathophysiological processes (Gao 2013). Urine is not regulated by the homeostatic mechanism and can reflect changes in metabolism in the body, making it easy to collect. Therefore, urine is a good biological source for finding disease markers (Gao 2015).

However, human clinical urine samples are affected by a variety of physiological or pathological effects, such as age, gender, diet, exercise, and drugs (Wu and Gao 2015). Animal models are the most effective way to find the causal relationship between diseases for the following reasons: (1) using animal models to reduce the effects of genetic and environmental factors on urine proteomics; (2) helping to identify biomarkers at each stage, including early diagnosis; (3) because clinical patient care is unavoidable, animal models can avoid the effects of drugs on the proteome. Therefore, the use of animal models for the study of urine protein markers is more economical and controllable, which is conducive to early observation of diseases and monitoring of related pathophysiological changes during disease progression (Zhao et al. 2014).

In research activities, rats are commonly used to establish disease animal models and conduct proteomic research, such as rat kidney disease model (Carter et al. 2016; Rosner 2009), rat coronary artery disease model (Paapstel et al. 2016; Zimmerli et al. 2008; Mullen et al. 2011), and rat bladder cancer model (Zhu et al. 2016; Vrooman and Witjes 2008). However, rats are not the dominant model animals for all diseases. Different diseases should choose their corresponding dominant model animals. Therefore, it is more necessary to study the superior models and proteomes of other animals. The hamsters commonly used as experimental animals are mainly the golden hamster and the Chinese hamster. The golden hamster has 38 inbred lines, 17 mutant lines, and 38 distant crosses. The hamster is mainly used in biomedical research in oncology research, reproductive physiology and family planning research, vascular physiology and microcirculation research, nutrition research, and infectious disease research. The hamster is the main biological material for the study of rabies virus and Japanese encephalitis virus and its vaccine production (Cui et al. 2014). Guinea pigs, also known as the squirrel, are widely used in immunology, nutrition, physiology, toxicology, and infectious disease research.

In this chapter, we analyze the difference in urinary proteins between rats, golden hamsters, and guinea pigs to provide basic data for medical experiments and provide a basis for the selection of dominant animal models.

3.2 Results and Analysis

Urine proteins were precipitated with ethanol and resuspended in lysis buffer. Urine proteins were digested with trypsin using the FASP method. Peptide mixtures were desalted and analyzed with LC-MS/MS using ABSCIEX Triple-TOF5600 mass spectrometer. The urinary protein of rat, golden hamster, and guinea pig was compared by SDS-PAGE. The differences were as follows: the rats differed significantly from others, while the golden hamsters and guinea pigs were more similar. The rat urinary proteins were mainly concentrated in 35–15 kDa, while the expressions between 35–25 kDa of guinea pig and golden hamster urinary proteins were weak (Fig. 3.1).
Fig. 3.1

SDS-PAGE analysis of urinary protein sample: R1-3, samples from rats; P1-3, samples from guinea pigs; H1-3, samples from golden hamsters. (Ni et al. 2018)

The results of urinary protein identification in rats, golden hamsters, and guinea pigs were searched by Mascot and then compared with human homologous proteins by UniProt and converted into adult homologous proteins. The comparison of three animal-human homologous proteins showed that there were 340 human homologous proteins in the urine of guinea pigs, 266 human homologous proteins in rat urine, and 366 human homologous proteins in the urine of golden hamsters. As shown in Fig. 3.2, there are 88 trusted proteins in all three species, while 86 proteins are unique to rats, 141 proteins are unique to guinea pigs, and 160 proteins are unique to golden hamsters. As shown in Fig. 3.3, the similarity of guinea pig and golden hamster urinary protein is greater.
Fig. 3.2

Rat, guinea pig, and golden hamster urine protein comparison Venn diagram. The purple part represents 86 proteins that are unique to rats, the yellow part represents 141 proteins that are unique to guinea pigs, and the green part represents 160 proteins that are unique to golden hamsters. (Ni et al. 2018)

Fig. 3.3

The heat map of three kinds of animal urine protein. From left to right are: rats, golden hamsters, and guinea pigs. The expression of protein in golden hamster and guinea pig is close. (Ni et al. 2018)

All proteins identified in the urine of rats, golden hamsters, and guinea pigs were analyzed by PANTHER database, and the urine proteins of rats, golden hamsters, and guinea pigs were compared in terms of molecular function, biological processes, cellular components, and pathways (Fig. 3.4). The analysis showed that the urinary proteins involved in the three species were different in biological processes and pathways. They had different molecular functions and different cellular components. For example, guinea pigs and golden hamsters have more urinary proteins involved in the calcium signaling pathway, the integrin signaling pathway, and the Wnt signaling pathway than in rats.
Fig. 3.4

Gene ontology analysis. There were significant differences in the expression of three animal proteins. (a) Molecular function. (b) Biological process. (c) Cellular component. (d) Pathway. (Ni et al. 2018)

The urinary proteins of rat, guinea pig, and golden hamster were compared with the Human Protein Atlas database. The results showed that the protein expression of the three species were different in the urinary system, digestive system, nervous system, respiratory system, immune system, and endocrine system, and the expression of urinary protein in golden hamsters is much higher than others. The expression of urinary protein in rats is the lowest (Fig. 3.5).
Fig. 3.5

The Human Protein Atlas result between rats, guinea pigs, and golden hamsters. In different organs, the urine protein expression of three animals was significantly different. Among them, the highest expression is of golden hamsters, followed by guinea pigs, and finally rats. (Ni et al. 2018)

3.3 Research Prospects

At this stage, rats are commonly model animals, and reports of studies using guinea pigs and golden hamsters are rare. This study compared the urine proteins of rats, golden hamsters, and guinea pigs by proteomic methods in order to lay the data foundation for medical experiments and provide clues for the selection of dominant animal models.

In this study, the urine proteins of rats, guinea pigs, and golden hamsters were found to be very different. It can be seen from SDS-PAGE that the three kinds of animal have different high-abundance proteins and different abundance inhibition, and the protein expression of guinea pigs and golden hamsters is relatively similar. Therefore, the use of rats alone to establish disease animal models is not suitable for studying all diseases. The expression of urinary protein in rats, guinea pigs, and golden hamsters was different in each organ of every system, and the biological processes involved in them were also different. The expression of rats was the least. It indicated that golden hamsters and guinea pigs are relatively more dominant in the establishment of human disease models. For example, in our data, it can be seen that in the liver and pancreas, the expression of urinary protein in golden hamsters is high. Now in the medical field, the mortality of intrahepatic cholangiocarcinoma and pancreatic cancer is high, the early diagnosis is difficult, and the prognosis is poor. It is particularly important to look for urine protein markers for both diseases. By consulting the relevant literature, it has been reported that golden hamsters are dominant model animals for intrahepatic cholangiocarcinoma and pancreatic cancer (Kawaura et al. 2011; Takahashi et al. 2011). In the skin and lungs, although the urinary protein expression of guinea pigs is not as high as that of golden hamsters, the urine of golden hamsters is less and thicker. On the urine samples, guinea pigs are more suitable. The early diagnosis and differential diagnosis of skin allergies, skin blemishes, and chronic obstruction pulmonary disease are difficult, and the treatment is poor. Therefore, we can find the urine protein markers for differential diagnosis and monitor the disease progression and disease efficacy. It has been reported that the guinea pig is the dominant model of chronic obstructive pulmonary disease, skin irritation, skin dermatophytes, and other diseases (Basketter 2016; Cambier et al. 2017; Ramírez-Ramírez et al. 2017). Through the above research, the selection of disease animal models can be guided. For example, in the selection of liver and pancreatic disease models, we can give priority to golden hamsters. In disease models such as skin diseases, we can give priority to guinea pigs and combine the disease and animal characteristics and previous reports to correctly select the dominant model.

In summary, the use of guinea pigs, golden hamsters, and rats to select their corresponding disease models can improve the efficiency and accuracy of urine protein marker screening, and have more prospect in the search for urine protein markers for early diagnosis of human diseases.

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Copyright information

© Springer Nature Singapore Pte Ltd. 2019

Authors and Affiliations

  • Na Ni
    • 1
  • Jianqiang Wu
    • 2
  1. 1.Institute of Molecular Medicine and Cancel, Department of Biochemistry and Molecular Biology, College of Basic Medical SciencesChongqing Medical UniversityChongqingChina
  2. 2.Medical Research Center, Peking Union Medical College HospitalChinese Academy of Medical Sciences and Peking Union Medical CollegeBeijingChina

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