It was widely accepted that the HFD is harmful to health and has an important role in shaping the composition of the gut microbiome, and the HFD was widely used in various animal models of pathogenic mechanism research of many diseases. However, there is no uniform standard for the compositions of feed using in HFD, which may cause different results in HFD experiments in different laboratories. Therefore, we designed this study in light of this situation to investigate the effects of fat sources in HFD on body weight and blood lipids as well as gut microbes to verify the fat sources have a significant impact on the experimental results. The result proved that our hypothesis is correct, and this difference does exist and will seriously affect the experimental results of HFD.
As a common animal source of fat, lard is widely used in the HFD experiment of animal models and food cooking of human, many researchers have used lard as a negative control in HFD intervention to highlight the health value of plant-originate fat or fish oil.27,28,29,30. While, in this study, although the median value has increased compared with the control group, the body weight increasing of lard intervention group was lower than most of the fat from various plant sources. Moreover, lard effects on live weight, CHOL, TRIG, HDL, and LDL showed similar results. These results indicated that the effects of lard on these indicators are not higher than that of other fats in the HFD. Further study of the HFD focus on related indicators, it may need to consider whether to use lard as a negative control carefully.
On the other hand, the impact of lard intervention on gut microbes is much more severe than other fats. The lard intervention had the most significant impact on the abundance of Firmicutes, Bacteroidetes, and Proteobacteria in the intestinal bacteria. We noticed that the composition of SFA in fat was the main driving element of the gut microbiome in this study, it makes sense because lard has a high content of SFA as about 40% in it. However, palm oil, which also has the highest content of SFA, did not show such severe effects on the diversity of the gut microbiome. Because the chemical composition of dietary fat is very complex, especially the oils derived from plants, which contain many kinds of metabolic products, so it is difficult to access the relationship between the diet fat components and the physiological indicators or gut microbiome, it is challenging to clarify the driving mechanism of these components on intestinal bacteria, and it needs further research.
Although almost all HFDs promote weight gain, and HFD cause hepatic steatosis and an increase of inflammatory cytokines in the liver 131, the different content of SFA and UFA can affect some results of HFD. In the study of Caesar et al., HFD with lard, which rich in saturated fatty acids, promotes the obesity process through intestinal bacteria, and promotes the occurrence of WAT inflammation through TLR signaling and CCL2 cytokines. However, fish oil rich in PUSA inhibits this process18. Moreover, fish oil does not raise liver cholesterol like lard17. This indicates that UFA has protective effects on the metabolism and inflammatory response caused by HFD. UFA mainly contains two types, PUFA and MUFA, and the effects of these two fats on metabolism are also different. Acute ingestion of a PUFA-rich HF meal induced a greater DIT in normal-weight women compared with SFA- or MUFA-rich HF meals32. In addition, the two affect the body metabolism at different rates, after a MUFA-rich meal, body show lower RER and greater DIT than PUFA HFD; However, after a 5-d high-fat diet, PUFA diet show greater change in metabolic responses, showing the metabolic adaptability of a PUFA-rich diet33. These indicate that the differences in the content of SFA, MUFA and PUFA will significantly affect the results of HFD interference experiments. The types of oils used in this study have great differences in the content of the three main fats, from the results of the correlation between the content of the three fats and body indicators or intestinal microbiota, we can see the differences among these dietary fats.
The correlation analysis between the composition of dietary fat and body weight shows that there is no significant correlation between body weight and the three main components (SFA, PUFA, MUFA). Although we have not counted the amount of food eaten by the mice during the experiment processes, because a considerable part of the feed was wasted during the teeth grinding34, we admit that food intake is important data in the HFD experiment, and we indeed observed that the food intake of sesame oil and palm oil treatment groups was slightly less than that of the other treatment groups during the experiment. There are indeed differences in intake between different dietary fat interventions, and we also agree that this difference may affect the indicators we measure. However, animal disease models based on HFD usually takes a long time (several months commonly) to induce the corresponding disease occurrence. Therefore, in the animal model experiment of HFD, it is unrealistic to control the intake of animals precisely, and most of them adopt free-eating. Our experimental method was designed according to the general experimental process of the HFD, and we believe that such results can guide the choice of fat types for the HFD. Regarding the influence of the same calorie intake on HFD, and the influence of specific molecular components in fats on HFD, more research is needed in the future to explain. The blood lipids contents are important indicators for clinical diagnosis, and the raising of blood lipids will significantly increase the probability of suffering from cardiovascular disease.35,36,37. We tested four important blood lipid indicators (CHOL, TRIG, HDL, LDL) and found that the different sources of dietary fat have different effects on these indicators. The results of olive oil treatment are not consistent with popular opinions that olive oil has always been considered a healthy dietary fat38. This may be because the fat concentration used in our experiment was too high, previous research shows that the effect of fat on the body is concentration-dependent. The analysis results from canola oil intervention data showed that the greatest benefits occurred when ~15% of the total energy intake was consumed39. It indicated that excessive intake of olive oil could also produce unhealthy results as the dietary fats, and it reminds us that when other ratio of HFD experiments, the intake fat may also be a critical factor that affects the results of the experiment.
Many studies proved that HFD affects the development of many diseases through the gut microbiome40,41. Our results showed that the sources of dietary fat could affect both the composition and diversity of the gut microbiota after HFD intervention. It is noteworthy that intervention with camellia oil and canola oil can increase the diversity of the gut microbiome, and in contrast, lard reduces it. Regarding the effects of a HFD, this difference is sufficient to produce the opposite conclusion. Therefore, in the HFD experiment, the source of the fat used should be carefully considered.
We noticed that canola oil does not seem to affect intestinal bacteria after the HFD intervention, and the body/liver weight or bloody lipid indicators data also showed canola oil has milder effects than other dietary fats. The previous reports show that canola oil can decrease triglyceride, post-intervention grading of fatty liver was reduced significantly42; Canola oil containing omega-3 PUFAs may confer cardiovascular protection by improving endothelial function and lowering LDL-cholesterol.43. Meta-analyses results also show that canola oil potentially improves cardiovascular risk factors compared with saturated fat, sunflower, and olive oil39. Although excessive intake of fats and oils is generally harmful to health, using canola oil in cooking may minimize the risk of harm. Of course, this result is unfavourable for most HFD experiments, but on the other hand, this has important reference significance for choice of dietary fats for cooking. Although it is not rigorous to conclude that the health value of canola oil is better than other categories based on the data of this research, it can at least partially explain that it is a better choice, and more comprehensive nutrition research should carry out in the future.
We also noticed that the abundance of Akkermansia muciniphila Changed significantly after HFD treatment, which indicated that it is very sensitive to HFD. A. muciniphila is a common human gut bacteria, promotes intestinal barrier function and produces nutrients that feed other gut bacteria44,45. It has been getting much attention because several studies have linked A. muciniphila to many metabolic and disease development processes, such as obesity46type 2 diabetes47heart disease48SCFAs production49Crohn’s disease50Ulcerative Colitis51Appendicitis52. We found that the abundance of A. muciniphila was negatively correlated with CHOL, HDL-C, and LDL-C in mice with dietary fat intervention, high-lighting Akkermansia As a potential mediator of the improved body mass and bloody lipids phenotype of mice with dietary intervention. This finding is in agreement with previous findings linking A. muciniphilia with protection to diet-induced obesity53. Even we can not conclude that the microbiota of canola oil intervention is more healthy than lard intervention group without further researches evidence, but the Firmicutes: Bacteroidetes of canola oil group is the smallest among the groups, and the A. mucinphila of canola oil group is higher than most others, that indicated that canola oil might be a healthy dietary fat for cooking from our bacterial community data.
Previous studies have shown that Helicobacteraceae increases in mice and humans after consumption of diets rich in saturated fats of animal origin, Helicobacter species could infect organs of the gastrointestinal tract, including stomach, intestine, and liver54,55. Helicobacter rodentium can trigger the development of IBD in mutated mice56. Our results showed that dietary oil intervention increased the abundance of Helicobacterit was closely related to H. pyloriwhich infections can cause severe stomach diseases in human57,58,59. It implied that HFD might increase the risk of H. pylori infection.
We compared the metabolic pathways of canola oil and lard groups, and found that of the 31 metabolic pathways with the most significant differences (p < 0.001), 22 of them were up-regulated in rapeseed oil, while only 9 were up-regulated in lard. This may be due to the higher diversity of intestinal bacteria treated with canola oil. Most of the up-regulated pathways are related to fatty acid synthesis and energy metabolism, indicating that the intestinal bacteria treated with canola oil can synthesize more fatty acid-related products, which has certain benefits to the host. While the lard treated group promotes the generation of some harmful substances, such as PWY490-3, promote the reduction of nitrate to nitrite, and excessive nitrite is harmful to the host; and the OANTIGEN-PWY pathway, which is included in the synthesis of O-antigens, and O-antigens is the part of the LPS that can induce some immune responses or cause immune diseases. Although the specific metabolic pathways corresponding to microbes are still unclear, the difference between the microbes and the metabolic pathway can be predicted to determine the corresponding regulatory relationship between them, thus laying a foundation for further study on dietary fats in HFD or cooking.