Discussion
Our current study suggested that inhalational anaesthetic sevoflurane enhanced ovarian cancer cell viability, proliferation, migration, and invasion. In contrast, intravenous anaesthetic propofol inhibited those cellular activities. These phenotypic observations could be associated with upregulated expressions of GLUT1, MPC1, GLUD1, p-Erk1/2, HIF-1α, CXCL12 and CXCR4, and downregulated PEDF expression in the sevoflurane group. Sevoflurane, but not propofol, promoted metabolism of ovarian cancer cells by enhancing the uptake of the metabolic substrates such as glucose and glutamine.
The lower concentration of glucose and higher concentration of pyruvate in the media were found after sevoflurane exposure compared to the naïve control, which were in agreement with observed upregulated expression level of GLUT1. The enhanced activity of GLUT1 may transport more glucose into the cytoplasm to convert to pyruvate, which explained the metabolic changes in the media after sevoflurane administration. In contrast, propofol downregulated GLUT1, which resulted in the less uptake of glucose from media, and hence the concentration of glucose was increased after propofol exposure. The cellular glucose uptakevia high level of GLUT1 expression was correlated with the malignancy of cancers (Leung, 2004; Pezzuto et al., 2020) and the overexpression of GLUT1 in cancer cells were essential for the high rate of glycolysis (Wright, 2020). Propofol was reported to downregulateGLUT1 gene in macrophages (Tanaka et al., 2010) and this was in line with the downregulation of GLUT1 proteins in the rat brain tissue under hypoxic preconditioning (Xiao et al., 2020). This is quite similar to cancer cell scenario, as, under high proliferative rate, cancer cells had inadequate oxygen supply and were actually under hypoxic condition (Mudassar, Shen, O’Neill, & Hau, 2020). These supported our findings that the malignancy of ovarian cancer cells was related to the expression of GLUT1 after anaesthetic administration.
The concentration of lactate was increased in both sevoflurane and propofol treatments, which might be resulted from the increased pyruvate in glycolysis and then generated more lactate. It was reported that sevoflurane and propofol both increased the lactate level in the blood of dogs (Söbbeler et al., 2018). Another study in mice also demonstrated that sevoflurane increased pyruvate and lactate levels (Horn & Klein, 2010). All these reports were in line with the findings of our current study.
It was found that both the expressions of MPC1 and GLUD1 were upregulated after sevoflurane exposure but were downregulated after propofol treatment. Besides, the concentration of glutamine in media was decreased after sevoflurane administration. MPC1, a member of mitochondrial carrier system, locates at the inner membrane of mitochondria and transports pyruvate into mitochondria from cytoplasm (Taylor, 2017). The expression of MPC1 is decreased in most tumour types especially those under a high rate of proliferation as related to the increased rate of glycolysis. This suggests that pyruvate likely shifts from the mitochondrial TCA cycle to cytoplasm glycolysis, which does not require oxygen supply as cancer cells are usually under hypoxic condition (Rauckhorst & Taylor, 2016). In addition, glutaminolysis compensates for the disturbed function of the TCA cycle due to less pyruvate intake and cancer cells, in turn, uptake glutamine and convert them into glutamate under the activation of GLUD1. The glutamate can be used by TCA cycle to restore the survival of cancer cells as the intermediates of TCA cycle are the source for synthesis of amino acids, proteins, fatty acids, lipids, carbon skeleton, and nucleic acids (Yoo, Yu, Sung, & Han, 2020). Evidence from other studies showed propofol might disturb the mitochondrial respiratory chain, which was related to TCA cycle (Berndt et al., 2018). It was also reported that unlike inhibitory effects of propofol, sevoflurane preserved the function of the mitochondrial respiratory chain in a myocardial ischaemic model (Lotz, Stumpner, & Smul, 2020). Our data demonstrated that after sevoflurane administration, the MPC1 and GLUD1 expressions were upregulated, which might enhance the activity of the TCA cycle to meet the demands of cancer survival and progression. The decreased concentration of glutamine in media suggested that the utilisation of glutamine was likely increased and glutaminolysis was then promoted. With the disturbed function of the TCA cycle after propofol exposure, amino acids that can be used in the TCA cycle were accumulated, such as asparagine and arginine (Pasini et al., 2018), which was consistent with our findings.
Except glucose and glutamine, another “mirror change” of metabolites between sevoflurane and propofol administration was isopropanol. It was reported that the level of isopropanol was increased in the exhaled breath of lung cancer patients, and it had been regarded as a potential biomarker for lung cancer diagnosis (Chien et al., 2017). It seemed the level of isopropanol had some correlations with cancer malignancy, which was consistent with the findings of this study that sevoflurane enhanced the malignancy of ovarian cancer cells and increased the level of isopropanol, while propofol inhibited the malignancy of ovarian cancer cells and decreased the level of isopropanol. Isopropanol can be reversibly converted to acetone (Beauchamp, Valento, & Kim, 2016; Li, Liu, Liu, Cheng, & Duan, 2017), which may also contribute to the increased level of acetone in the media of the propofol group.
The levels of glycerol and fatty acids were increased in the propofol group. Through β-oxidation of fatty acids, acetyl-CoA is generated and used in the TCA cycle (Y. Liu, 2006). Thus, the changes of glycerol and fatty acids in the propofol group were another evidence that the mitochondria function and TCA cycle was inhibited or disturbed by propofol treatment. The glycerol and fatty acids might also come from cell membrane degradation and phospholipids broke down into them. From an earlier study, it was found that propofol affected the membrane ultrastructure of HeLa cells that the surface roughness of cellular membrane was decreased in a dose-dependent manner (Zhang et al., 2016).
In the current study, the expression level of PEDF was decreased after sevoflurane administration but increased after propofol treatment. In human retinal pigment epithelium, the expression of GLUT1 was increased under hypoxia condition that resulted in the increased uptake of glucose, which led to a decrease of PEDF expression (Calado, Alves, Simão, & Silva, 2016). Another study also reported that the overexpression of the PEDF gene in mice was related to the reduction of glucose uptake and decreased expression of GLUT1 (Calado, Diaz-Corrales, & Silva, 2016). These reports were consistent with our findings that sevoflurane increased the GLUT1 expression and glucose uptake, which led to a downregulated expression of PEDF. However, an opposite effect was found with propofol treatment.
In the current study, Erk1/2 signalling pathway was induced after sevoflurane exposure but inhibited after propofol exposure. There was evidence that the increased expression of PEDF was related to the inhibition of Erk1/2 signalling pathway in a diabetic model (Dong et al., 2019), which was in line with our results. HIF-1α is a transcriptional factor that can be regulated by a variety of signalling pathways, and Erk1/2 pathway is one of them (R. M. Liu, Xu, Chen, Feng, & Xie, 2020). In cancer cells, the HIF-1α is overexpressed, which regulates tumour survival-related genes, such as CXCL12 andCXCR4 (Gola et al., 2020; Xue et al., 2020). It was in line with the results of the current study that sevoflurane upregulated Erk1/2 signalling pathway, and HIF-1α, CXCL12 and CXCR4 expressions, while propofol downregulated these molecular entities.
Our study has some limitations. Firstly, the causal relationship between cellular signalling changes and metabolic alterations induced by anaesthetics remains unknown. However, it is very likely that, for example, sevoflurane promotes cancer cell survival and development due to survival cellular signalling pathway activation whereby more energy substrates use up for cell proliferation and growth. Secondly, our cultured cell study may not be relevant to human. Therefore, the implications of our current study may be limited. However, in some clinical studies, breast, colonic and rectal cancer patients were anaesthetised with inhalational anaesthetics sevoflurane or desflurane, or intravenous anaesthetic propofol during surgery and the survival rate of propofol anaesthetised patients were significantly higher than those with inhalational anaesthesia (Enlund et al., 2014; Wu et al., 2018). Laboratory data including the one reported here and retrospective clinical data all point to that sevoflurane might be a risk factor for cancer patients, while propofol may be beneficial to cancer patients for their surgery. Therefore, clinical studies are urgently needed to evaluate anaesthesia regimens for cancer patients to optimise the surgical outcomes.