Fat-rich diet prevents bladder stones | Tsinghua University School of Pharmacy Wang Zhao Research Group issued a report that fat-rich diet can reduce the risk of primary bladder stones

On September 8, 2021, the Wang Zhao research group of the School of Pharmacy of Tsinghua University made important progress in the relationship and mechanism between dietary fat components and the formation of primary bladder stones. The latest research results, entitled "Ample dietary fat reduced the risk of primary vesical calculi by inducing macrophages to engulf budding crystals in mice", were published online in Acta Pharmaceutica Sinica B (APSB), a well-known academic journal in the pharmaceutical field. This study revealed that in the primary bladder stone model, when the diet is rich in fat, bladder epithelial cells can secrete more chemokine 14(C-X-C motif chemokine ligand 14,CXCL14), and then recruit more macrophages, so that macrophages can better play the role of phagocytosis of the initial crystal germination of stone formation, and ultimately reduce the formation of primary bladder stones.

Ancient disease formation: What does it have to do with fat?
Primary bladder stone is an ancient disease, but its formation mechanism is not clear. Malnourished children in poor areas are the main patients with primary bladder stones.
With the increase of breastfeeding and dairy intake, the incidence of bladder stones decreased gradually.
In the past, researchers believed that protein in dairy products is the main factor to prevent the formation of bladder stones, but the role of other nutrients in dairy products such as fat has been ignored.
　　This article discusses the relationship between dietary fat composition and the formation of primary bladder stones and the molecular mechanism, in order to find effective interventions to reduce the incidence of the disease.

Construction of primary bladder stone model: fat-rich diet reduces disease incidence
First of all,The authors innovatively constructed a mouse model of primary bladder stones using DL-choline tartrate,Compared with the commonly used melamine and ethylene glycol models, this method is currently the only method to induce bladder stones without causing kidney damage.
In order to examine the effect of fat composition on the formation of primary bladder stones, the authors fed mice with fat-rich diet (HF), normal diet (NF) and low-fat diet (LF) for a period of 20 weeks.
As shown in Figure 1, the mice in the fat-rich diet group had no stones in the bladder, while the incidence of bladder stones in the normal and low-fat diet groups was 100 percent.It is suggested that fat-rich diet can reduce the occurrence of primary bladder stones in mice.

Figure 1 Fat-rich diet can reduce the incidence of primary bladder stones

Mechanism of action: how a fat-rich diet slows stone formation
　　When exploring the mechanism of fat-rich diet slowing down the formation of primary bladder stones in mice, the authors chose the time point of short-term 4-week dietary intervention.
By detecting the concentration of calcium and tartaric acid that affect the formation of stones, the levels of various inorganic salt ions, and the changes in the levels of calcium absorption hormones, it is inferred that changes in biological properties may affect the formation of stones, rather than physical and chemical changes.
The higher levels of free fatty acids in the urine and blood of the mice in the fat-rich diet group were involved in maintaining the normal microstructure of the bladder epithelium of the mice in this group.
The results of RNA transcriptome sequencing of the three groups of mouse bladder tissues were further analyzed by transcriptome sequencing,The results showed that the expression of CXCL14 in the bladder tissue of the mice in the fat-rich diet group (HF group) was significantly increased (Figure 2).

Figure 2 The expression of CXCL14 in bladder epithelial cells was significantly increased in the fat-rich diet group

Phase 1: Clarify the involvement of M2 macrophages
Macrophage activation can be divided into "classic"/M1 and "alternative"/M2, which play opposite roles in immune and inflammatory responses.
　　The authors further distinguished between M1 and M2 macrophages in each group of mouse bladder tissue by selecting specific markers for M1 and M2 macrophages.

Figure 3 Mouse bladder epithelial cells in the fat-rich diet group play a phagocytic role by recruiting more macrophages

The results of Figure 3 show that there is no significant difference in the number of M1 (CD68 /F4/80) macrophages in the bladder tissue of the three groups, while the number of M2 (CD163 /F4/80) macrophages in the HF group is significantly higher than that in the other two groups.
　　These results suggest that M2 macrophages may be mainly involved in the clearance process of crystals.

Phase II: Verification of the effect of fatty acids
To further validate the role of fatty acids,The authors cultured human bladder epithelial HCV-29 cells in vitro, by adding different concentrations of free fatty acids to the HCV-29 cell culture medium, and then determine the expression of CXCL14 in cells and culture medium supernatant.
The results show that (as shown in Figure 4), after adding additional free fatty acids for 24 hours, the expression of CXCL14 in the supernatant of HCV-29 cells and cell culture medium will increase significantly, and the increasing trend of CXCL14 is proportional to the amount of fatty acids added,Thus, it was further confirmed that the addition of fatty acids increased the expression of CXCL14 secreted by HCV-29 cells.

Figure 4 The addition of fatty acids makes HCV-29 cells secrete CXCL14 and then promote macrophages to play the role of phagocytosis of crystals

　　

In the primary bladder stone model, the crystals gradually accumulate in the mucosal layer of the bladder tissue.
In the fat-rich diet mode, due to the intake of more fat, on the one hand, the mucosal layer of bladder tissue can maintain its normal tissue structure, on the other hand, the increase of free fatty acid content in serum and urine promotes the secretion of more CXCL14 protein from bladder epithelium. CXCL14 protein can further induce local macrophages to differentiate into M2 type, and then play an effective role in removing crystals.
Under the low-fat diet mode, the integrity of the mucosal layer of bladder tissue is destroyed, and the accumulation of crystals is accelerated, which leads to the formation of primary bladder stones.
In summary, studies have shown that fat-rich diet can reduce the occurrence of primary bladder stones in mice. The main mechanism is that fat-rich diet can increase the level of fatty acids in the body, and induce macrophages to phagocytize the germination of crystals in the early stage of stone formation by recruiting macrophages to migrate to the bladder epithelial tissue, so as to prevent the formation of stones. The experimental results show that dietary fat is of great significance in maintaining bladder physiological function and preventing primary bladder stones. This study provides intervention methods and measures for clinical prevention of bladder stones.

Professor Wang Zhao of the School of Pharmacy of Tsinghua University is the corresponding author of this article, and Chen Huiling and Hu Kaiqiang, doctoral students of the School of Pharmacy, are the co-first authors. Laboratory members, doctoral students Liang Yaru, Gao Yuqi, Zeng Chenye, Xu Kang, Dr. Qiao Yi, Department of Urology, Peking Union Medical College Hospital, Associate Professor Qiu Ying, School of Medicine, Tsinghua University, and Associate Professor Liu Qingfei, School of Pharmacy, have made important contributions to this research. This study was supported and funded by the National Natural Science Foundation of China (81974503, 81871095), the Ministry of Science and Technology Key R & D Program (2018YFC2000304), the Overseas Key R & D Program (2016YFE0113700), and the Beijing Natural Science Foundation (7202096).

Institute of Biomedicine and Equipment
The Institute of Biomedicine and Equipment was established in 2016 with Professor Wang Zhao as its director. The team is committed to in-depth research in many fields such as life science, biomedicine, medical equipment, bio-related fluid mechanics and mechanical design, food safety and nutrition, etc. It also focuses on the development and application of ECMO blood pump, brain-strengthening instrument, sleep aid series products, household caries risk prediction and evaluation system, caries prevention ready-to-eat nutritious food, fruit and preservation of fruits and vegetable preservation, at the same time, it has obtained a number of related patents.

Reprinted | School of Pharmacy, Tsinghua University