LPS exposure during pregnancy is one of the adverse exposure factors in pregnancy that cannot be ignored. At present, no studies have focused on the effect of LPS exposure during pregnancy on maternal and offspring metabolism from the perspective of metabolomics. A model of low-dose LPS exposure during pregnancy causing glucose metabolism disorders in adult male offspring has been reported[7]. Metabolomics was used to analyze the metabolic changes of maternal serum and male fetal liver after LPS exposure during pregnancy. The results of this study showed that low-dose LPS exposure during pregnancy had a similar tendency to interfere with the glycerophospholipids metabolism and fatty acid metabolism of maternal and male offspring.
4.1 Effects of LPS exposure during pregnancy on maternal metabolites
Some researchers have suggested that inflammatory mediators activate the innate immune system to cause lipid remodeling of glycerolipids, glycerophospholipids, and prenols in majority of mammalian[32]. Most of previous studies have analyzed the effect of inflammation on the glycerophospholipids metabolism in RAW264.7 cells in mice[33–36]. Dennis et al. characterized lipidomic (glycerides, glycerophospholipids, sphingolipids, and fatty acids) responses of murine macrophage to inflammatory by LC/MS[32]. It reported that levels of most unsaturated free fatty acids were reduced at longer time points, whereas most free saturated fatty acids increased after treated with LPS. Furthermore, in the glycerophospholipids and glycerolipids, this trend was especially noticeable for the saturated and monounsaturated species in that the 32:0, 34:0, 34:1, 36:0, and 36:1 phosphatidic acid species increased by severalfold in response to the stimulus, whereas certain polyunsaturated species such as 38:4 phosphatidylinositol showed decreases of up to 50%[32]. She et al. identified 16 altered glycerophospholipids in LPS-treated RAW264.7 cells. Among the 16 glycerophospholipids, significant decreases in the levels of lysoPC (20:4), PC (36:4), PC (34:4), PC (35:4), PC (36:5), and PC (40:5) species were accompanied by relative increases in the levels of lysoPE (20:3), lysoPE (22:1), lysoPC (19:3), lysoPC (P−18:0), PC (O−32:3), PC (O−30:0), PC(O−32:0)species in the LPS-treated[33]. And other studies have similar results[36] [34]. These studies have suggested that LPS exposure leads to upregulation of glycerophospholipids containing saturated or monounsaturated fatty acids, and downregulation of glycerophospholipids containing polyunsaturated fatty acids. However, changes in glycerophospholipids caused by inflammation involve complex mechanisms and further research is needed.
In the present study, we found that fatty acid metabolism in maternal serum was significantly changed after LPS exposure, stearic acid, palmitic acid, oleic acid up-regulated, and 3-dehydroxycarnitine were up-regulated, while polyunsaturated fatty acid DHA was down-regulated. Stearic acid and palmitic acid are saturated fatty acids, oleic acid is a monounsaturated fatty acid, and DHA is an omega−3 polyunsaturated fatty acid. 3-dehydroxycarnitine is an acylcarnitine. 8,11,14-Eicosatrienoic acid, is an omega−9 polyunsaturated fatty acid. Dennis et al. reported that levels of most unsaturated free fatty acids were reduced at longer time points, whereas most free saturated fatty acids increased after treated with LPS [32]. These polyunsaturated fatty acids included linolenic acid, α-linolenic acid, DHA, stearidonic acid and 5,8,11,14,17-eicosapentaenoic acid, and saturated fatty acids included lauric acid (12:0), myristic acid (14:0), palmitic acid, stearic acid and oleic acid etc[32]. Another study also suggested that palmitic acid, palmitoleic acid, stearic acid, and oleic acid increased under LPS stimulation[33]. Presently, few studies have investigated the effects of inflammation on fatty acid metabolism, and more studies have focused on the pro-inflammatory or anti-inflammatory effects of fatty acids. Therefore, it is unclear whether LPS leads to saturated fatty acids or omega−9 polyunsaturated fatty acids, as well as to the down-regulation of polyunsaturated fatty acids[37–39]. Furthermore, it is controversial that whether monounsaturated fatty acids have anti-inflammatory effects[40]. Accordingly, these results suggests that inflammation may interact with fatty acids. Further research and verification are needed in the future.
4.2 Effects of LPS exposure during pregnancy on male offspring metabolome
Fatty acids (FAs) are one of the most essential substances for fetal intrauterine growth. They are involved in many energy and metabolic processes, including cell membrane, retina and nervous system growth. Fatty acid deficiency and maternal-fetal metabolic disorders can lead to fetal malnutrition and premature delivery. Importantly, it may cause metabolic and cardiovascular diseases in adulthood[41]. As the most important metabolic organ of human beings, the liver is also an important site of fatty acid metabolism. This study found that LPS exposure during pregnancy caused changes in liver glycerophospholipids and fatty acid metabolism in male fetal liver, which may be a risk factor for the development of glucose metabolism disorders in adulthood. It is now clear that fatty acids influence a range of other diseases, including metabolic diseases such as type 2 diabetes, inflammatory diseases, and cancer. Studies have suggested that saturated fatty acids, especially even saturated fatty acids (lauric acid, myristic acid, palmitic acid, and stearic acid) increase total cholesterol and low-density lipoprotein cholesterol concentrations, as well as increase body inflammation and insulin resistance. Therefore, exposure to high levels of saturated fatty acids is associated with a higher risk of coronary heart disease, cardiovascular disease, and T2D[40]. Linoleic acid (18:2ω−6) is an omega−6 polyunsaturated fatty acid that lowers blood cholesterol and low-density lipoprotein cholesterol[43, 44]. Omega−3 polyunsaturated fatty acids, mainly including α-linolenic acid (18:3ω−3), DHA (22:6ω−3), EPA (20:5ω−3), docosapentaenoic acid (DPA, 22:5ω−3) has a wide range of biological effects, involving membranes, signal transduction, gene expression and lipid mediators, plays an important role in controlling inflammation, reducing blood triglyceride and adipocyte differentiation [45–47]. Therefore, omega−3 polyunsaturated fatty acids play an important role in reducing the risk of diabetes[48]. At the same time, EPA and DHA are essential for fetal growth and development. DHA plays a central role in the structure and function of the eye and brain [40]. Epoxy eicosatrienoic acid is a metabolite of arachidonic acid and has a variety of biological functions, and it is closely related to islet cell function, insulin resistance, blood glucose homeostasis and progression of T2D[49–51]. In this study, we found that stearic acid was up-regulated, while the omega−3 polyunsaturated fatty acids DHA, EPA, α-linolenic acid and epoxy eicosatrienoic acid were down- regulated. This suggests that intrauterine LPS exposure leads to an increase in risk factors for T2D and protective factors decrease, which may be one of the causes of disorders of glucose metabolism in adulthood. Stearic acid, alpha-linolenic acid, DHA, EPA, and epoxyeicosatrienoic acid may be early markers of glucose metabolism disorders in adult male offspring of intrauterine LPS exposure. At the same time, intrauterine LPS exposure led to a significant decrease in DHA, which is essential for fetal growth and development. This suggests that intrauterine LPS exposure may have adverse effects on the normal growth and development of the fetus and may affect the normal development of the eye or brain. In addition, the study also found that ceramide carnitine, isobutyryl-L-carnitine and stearyl carnitine were significantly up-regulated. Ceramide carnitine, isobutyryl-L-carnitine and stearyl carnitine belong to the acylcarnitine. Acetylcarnitine is produced by mitochondrial matrix enzymes from carnitine and acetyl-CoA, which are products of fatty acid beta oxidation and glucose oxidation. Forty-five acylcarnitines were analyzed in the IRAS study and found to be associated with higher concentrations of carnitine insulin resistance[31]. Rui et al found that acylcarnitine, especially acetylcarnitine C2, is a marker for diabetic patients[52]. These studies suggest that upregulation of acylcarnitine may be associated with the risk of developing diabetes. Based on this, we hypothesized that abnormal up-regulation of ceramide carnitine, isobutyryl-L-carnitine and stearyl carnitine may be an early metabolic imprint of glucose metabolism disorder in adult male offspring caused by intrauterine LPS exposure.
Recently, more studies have focused on the relationship between glycerophospholipids and T2D. Maria A et al. investigated the proportions of fatty acids in plasma phospholipids, as predictors for the worsening of T2D [53]. In the glycerophospholipids fraction, saturated fatty acids, such as myristic acid (14:0), stearic acid (18:0) and palmitic acid (16:0) ratio were associated with decreased insulin sensitivity. In addition, glycerophospholipids with a high DHA ratio are associated with increased insulin sensitivity[53]. For insulin secretion, linoleic acid (18:2n−6) is associated with increased insulin secretion in glycerophospholipids[53]. Wang et al. evaluated the relationship between phospholipids of different fatty acids and the incidence of diabetes by following 2909 people. It was found that after adjusting for factors such as age, gender, baseline body mass index, waist-to-hip ratio, alcohol intake and parental diabetes history, The incidence of diabetes is positively correlated with the ratio of cholesterol in blood and saturated fatty acids in glycerophospholipids. In cholesterol esters, the incidence of diabetes is positively correlated with the ratio of palmitic acid (16:0), palmitoleic acid (16:1n−7) and gamma-linolenic acid (20:3n−6), with linoleic acid ( The ratio of 18:2n−6) is negative. In glycerophospholipids, the incidence of diabetes is positively correlated with the ratio of palmitic acid (16:0) and stearic acid (18:0) [54]. Other studies have found similar results[55–57]. In summary, the relationship between glycerophospholipids and the onset of diabetes is related to the type and proportion of fatty acids contained therein, probably because glycerophospholipids are a reservoir of free fatty acids. In this study, it was found that saturated fatty acid chains, especially glycophospholipids containing (16:0) and (18:0) chains, were up-regulated. This result suggests that abnormal changes in glycerophospholipids are another possible mechanism of glucose metabolism disorders in adult male offspring of intrauterine LPS. At the same time, these results suggest that the metabolic changes of maternal and progeny in the model of LPS exposure to male progeny glucose metabolism are mainly related to the composition and proportion of fatty acids. Therefore, we should pay attention to the changes of fatty acids during pregnancy.
Studies have found that many adverse exposures during pregnancy may lead to an increased risk of glucose metabolism in adult offspring. One study have found that bile acids and tryptophan metabolism are novel pathways involved in metabolic abnormalities in bisphenol A -exposed pregnant mice and male offspring [11]. In this study, we find that maternal serum and male fetal liver showed similar changes in glycerophospholipid and fatty acid metabolism. That indicates the abnormal glycerophospholipids and fatty acid metabolism may be the mechanism of glucose metabolism disorder in male offspring.
Despite these findings, our studies have some limitations. First, this study only characterize the alterations of maternal and male offspring metabolome after LPS exposure, but we did not explore the underlying mechanism of these changes. Second, this study did not detect the metabolites in amniotic fluid and could not explain the relationship between maternal and offspring metabolic changes. Although the detailed exploration of glycerophospholipids and fatty acid metabolism pathways is beyond the scope of this study, future analysis, including studies on the input and/or transport of fetal fatty acids, will be potentially important in elucidating the mechanism of LPS-exposured changes in glycerophospholipids and fatty acid metabolism.