Aiming at cancer, a momentous problem in the history of human medicine, scientists around the world are constantly exploring new ways of treatment from various angles. Early stage, people have done a lot of research on solid tumor itself, most of which are committed to inhibiting tumor proliferation and migration, improving autoimmunity and drug treatment. However, with the deepening of research, we came to know the concept of tumor microenvironment (TME)[1]. This new field has been developed so far, it is no stranger to the typical states of hypoxia, acidity, high content of lactate, abundant blood vessels and infiltrating immune cells in the local environment around the tumor. In addition, the characteristics of acidic microenvironment have attracted even more researchers' curiosity. In order to sustain the high proliferation of cancer cells, anaerobic glycolysis and increased oxidative phosphorylation are selected for rapid energy supply which engenders a large amount of CO2 and H+[2]. As a result, the pH value in the area near cancer cells is lower than that in normal tissues. Previous studies have shown that acidic cellular environment can lead to DNA strand breakage and histone deacetylation, thereby increasing genetic instability and increasing the probability of cell carcinogenesis[3, 4]. Not only that, the acidic environment also assists cancer cells avoid the attack of immune cells[5]. But this does not signify that cancer cells are eosinophilic, it is just that they can adapt to the acidic environment better than normal cells. For cancer cells, excessive H+ produced by metabolic preference in cancer cells will also restrain the enzymes of glucose metabolism, thus restricting the proliferation of itself[6]. However, its cunning lies in that cancer cells are equipped with a more efficient system to release H+ outside the cell and maintain intracellular balance: Na+/H+-exchangers, Na+,HCO3− -cotransporters, and H+-ATPases [7, 8].
As an important transporter for maintaining cell acid-base equilibrium, Na+/H+ exchanger exists in all apparatus of mammals[9]. Just like its name, it is responsible for absorbing extracellular Na+ in the form of active transport, discharging excess H+ produced by metabolism and other biochemical reactions, and maintaining the pH requirements of various biological reactions in the intracellular environment[10]. So far, 13 subtypes of Na+ / H+ ion pumps have been found, belonging to a large family (NHE), named NHE1-5 (SLC9A1-5), NHE6-9 (SLC9A6-9), NHE10, NHE11 (SLC9C1, C2) and NHA1, NHA2 (SLC9B1, B2)[11, 12]. They are distributed in different regions of cells and engage in similar work. In addition to their common functions of intracellular alkalization and cell volume control, individual members of NHE family have also been revealed to be closely related to the occurrence and development of common inflammatory diseases and even tumors[13]. NHE1, which is almost only located outside the plasma membrane, has been studied more. It has been found that it is anchored with the cytoskeleton to participate in cell migration and invasion[14]. Abnormal NHE1 mRNA levels have been detected in pathological samples of a variety of digestive system tumors[15]. Animal and cytological experiments have also proved that it does make outstanding contributions to life activities such as cell proliferation and apoptosis, but the explicit mechanism and clinical practice have not received conclusions and support to verify these fingdings[16]. In NHE family, except NHE3 and NHE7-9, there are few studies on other family members for reference. Among them, NHE3 has a small number of reports that it plays a role in enteric diseases[17]. NHE7 affects the development of pancreatic cancer[18] and breast cancer[19] by changing the acid-base balance of cells, and the abnormal expression of NHE8, NHE9 in colon cancer suggesting that it may have an important impact on the development and prognosis of cancer[20, 21]. In general, the physiological function of NHE family in cancer has yet to be explored.
As is known to all ,the physiological functions of the nervous system and digestive system are extremely dependent on hydrogen ion exchange[22]. We found that there had been a great quantity studies on the regulation of NHE family on the occurrence and development of glioma[23]. Nevertheless, there are few studies in colorectal cancer (CRC). CRC remains a global public hygiene problem as usual, leading to high cancer-related mortality. According to statistics, CRC accounts for about 10% of global cancer diagnosis and cancer-related deaths every year. Moreover, on the basis of the latest research statistics, CRC has become the second deadliest cancer in the world[24, 25]. Although surgery, chemotherapy and radiotherapy are quite effective for patients in stage I and II, follow-up observation shows that the probability of 5-year survival has increased to 66%. Unfortunately, we all know that the insurmountable fatal point of CRC lies in the invasion and metastasis of cancer cells. However, modern clinical medicine has failed to reach the level of limiting its metastasis. The 5-year survival rate of patients with advanced colorectal cancer is only about 13%[26]. Consequently, it is urgent to find new approach to prevent and cure CRC.
Our study is inspired by the acidic characteristics of tumor microenvironment. Based on the NHE family that regulates the pH value inside and outside the cell, combined with the H + dependent physiological function of the intestine. Using the available data resources, we can comprehensively develop new ideas for the defense and treatment of colorectal cancer. At first, we used the data from TCGA to analyze the mRNA expression of 10 subtypes of NHE family in COAD relative to the adjacent cancer, as well as the clinicopathological related effects (since SLC9B1, SLC9C1 and C2 are only expressed in male testis, they are not within the scope of this study). Immediately, we found 90 proteins interacting with NHE family members through STRING, and speculated that NHE family members may participate in the reaction mechanism. Certainly, whether NHE family members affect the number and function of infiltrated immune cells and stem cells in the tumor microenvironment, we also made a full analysis using TCGA and TIMER databases. Then we utilized Cytoscape software to structure the ceRNA network of NHE family members. The screened ceRNA may become a new method for cancer treatment. In addition, we found the correlation between DNA methylation in COAD and NHE family members through DNMIVD, and obtained IHC images of NHE family members in COAD and normal colon tissues in HPA. The cure and survival rate are the most direct criteria for the research and treatment of any disease. Therefore, we made a survival analysis related to COAD for NHE family. In addition to COAD, we also used R-package "ggpubr" to understand the expression of NHE family in other cancers. The drug sensitivity of NHE family is a test for its future clinical application, so we studied the data obtained from cellminertm with "limma", "ggplot2" and "ggpubr". Finally, all our results and graphs were completed by R software (version 4.0.5), and the p value of t-test < 0.05 was considered statistically significant.