Animals ; Autophagy/genetics* ; Cholesterol/metabolism* ; Gene Expression Regulation ; Integrases/metabolism* ; Leydig Cells/metabolism* ; Male ; Mice, Knockout ; Multienzyme Complexes/metabolism* ; Phosphoproteins/metabolism* ; Primary Cell Culture ; Progesterone Reductase/metabolism* ; RNA Splicing Factors/metabolism* ; Scavenger Receptors, Class B/metabolism* ; Sequestosome-1 Protein/metabolism* ; Signal Transduction ; Sirtuin 1/genetics* ; Sodium-Hydrogen Exchangers/metabolism* ; Steroid 17-alpha-Hydroxylase/metabolism* ; Steroid Isomerases/metabolism* ; Testosterone/genetics*

HCO3(-) reabsorption in the renal tubules plays a critically important role in maintaining the global acid-base balance. Loss of HCO3(-) causes metabolic acidosis. Proximal renal tubule is the major site for HCO3(-) reabsorption, accounting for more than 80% of total HCO3(-) reabsorption along the nephron. Over the past more than half centuries, tremendous progresses have been made on understanding the molecular mechanisms underlying the HCO3(-) reabsorption in proximal tubules. The transepithelial movement of HCO3(-) involves the coordinated operation of machineries on both the apical and the basolateral membranes of the epithelial cells. On the apical domain, Na(+)-H(+) exchanger NHE3 and the vacuolar H(+)-ATPase are two major pathways mediating the apical uptake of HCO3(-)-related species. Taken together, NHE3 and H(+)-ATPase are responsible for about 80% of HCO3(-) reabsorption in the proximal tubule. The remaining 20% is likely mediated by pathways yet to be characterized. On the basolateral membrane, NBCe1 represents the only major known pathway mediating the extrusion of HCO3(-) coupled with Na(+) into the interstitial space. In the present article, we provide a historical view about the studies on the mechanisms of HCO3(-) reabsorption since 1940s. Moreover, we summarize the latest progresses emerging over the past decade in the physiological as well as pathological roles of acid-base transporters underlying the HCO3(-) reabsorption in proximal tubules.
Acidosis ; physiopathology ; Animals ; Bicarbonates ; metabolism ; Humans ; Kidney Tubules, Proximal ; physiopathology ; Sodium-Hydrogen Exchangers ; physiology ; Vacuolar Proton-Translocating ATPases ; physiology

BACKGROUNDRecent studies showed the central Na+/H+ exchanger type 3 (NHE3) has a close relationship with ventilation control. The objective of the study is to investigate the role of NHE3 in sleep apnea in Sprague-Dawley (SD) rats.

METHODSA sleep study was performed on 20 male SD rats to analyze the correlation between the sleep apneic events and total NHE3 protein content and inactive NHE3(pS552) in the brainstem measured by Western blotting. Another 20 adult male SD rats received 3 days of sleep and respiration monitoring for 6 hours a day, with adaption on the first day, 0.5% DMSO microinjection into the fourth ventricle on the second day, and AVE0657 (specific inhibitor of NHE3) microinjection on the third day. Rats were divided into two groups with injection of 5 µmol/L or 8 µmol/L AVE0657 before the sleep study. The effects of AVE0657 on sleep apnea and sleep structure of rats were analyzed through self-control.

RESULTSThe total post-sigh apnea index (TPSAI) and post-sigh apnea index in non-rapid eye movement (NREM) sleep (NPSAI) and total apnea index (AI) in NREM sleep (NAI) were negatively correlated with NHE3(pS552) protein contents in the brainstem (r = -0.534, -0.547 and -0.505, respectively, P < 0.05). The spontaneous apnea index in REM sleep (RSPAI) was positively correlated with the level of NHE3(pS552) protein expression in the brainstem (r = 0.556, P < 0.05). However, the sleep AI had no relationship with total NHE3 protein. Compared with the blank control and microinjection of 0.5% DMSO, 5 µmol/L AVE0657 significantly reduced the total AI and NPSAI (both P < 0.05) without a significant effect on sleep architecture. In contrast to blank control and microinjection of 0.5% DMSO, injection of 8 µmol/L AVE0657 significantly reduced the AI and PSAI in NREM and REM sleep (all P < 0.05).

CONCLUSIONSThe severity of sleep apnea was negatively correlated with central inactive NHE3. A specific inhibitor of NHE3 decreased the sleep AI. Thus, our results indicate that central NHE3 might be a molecular target for sleep apnea treatment, whose inhibitors may be potential therapeutic drugs for sleep apnea.

Animals ; Male ; Rats ; Rats, Sprague-Dawley ; Sleep Apnea Syndromes ; metabolism ; physiopathology ; Sleep, REM ; physiology ; Sodium-Hydrogen Exchanger 3 ; Sodium-Hydrogen Exchangers ; antagonists & inhibitors ; metabolism

This study was purposed to explore the changes of possible angiogenetic factors other than VEGF after inhibition of NHE1 and their related mechanisms. The K562 cells were treated by NHE1 specific inhibitor cariporide, the angiogenesis factors after inhibition of NHE1 were screened by using protein chip, the IL-8 expression level after cariporide treatment was detected by real-time quantitative PCR; the K562 cells with stable interference of NHE1 were constructed, the IL-8 expression level after interference of NHE1 was detected by real-time quantitative PCR; the p38 phosphorylation level in K562 cells treated with cariporide was detected by Western blot. After treatment of K562 cells with p38 inhibitor SB203580, the IL-8 expression level was decreased by real-time quantitative PCR. The results of protein chip showed that IL-8 expression decreased after cariporide treatment. Real-time quantitative PCR confirmed this inhibitory effect. The p38 phosphorylation level increased after cariporide treatment. The down-regulation of IL-8 expression induced by cariporide treatment was partially restored after K562 cells were treated with p38 inhibitor SB203580. It is concluded that the inhibition of NHE1 can inhibit IL-8 expression through up-regulation of p38 phosphorylation.
Cation Transport Proteins ; antagonists & inhibitors ; Down-Regulation ; Guanidines ; pharmacology ; Humans ; Imidazoles ; pharmacology ; Interleukin-8 ; metabolism ; K562 Cells ; Phosphorylation ; drug effects ; Pyridines ; pharmacology ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; antagonists & inhibitors ; Sulfones ; pharmacology ; p38 Mitogen-Activated Protein Kinases ; metabolism

Abstract:Subgroup J avian leukosis virus (ALV-J) infect cells by binding to the chNHE1 receptor protein of the host and causes tumors. The tumor incidence of the ALV-J-infected chickens was observed by histo pathology, and virus was isolated on DF-1 cell line. The ALV-J load and mRNA of chNHElreceptor protein were detected by real time PCR. The relationship between ALV-J load, chNHE1 receptor expression levels and tumor spectrum was analyzed. The results showed that the tumors induced by ALV-J in laying hens and local lines of chicken were different. No significant relationship was observed between ALV-J load and tumor spectrum. ALV-J load was positively correlated with mRNA expression of chNHE1. The mRNA expression of chNHE1 increased when the tumors occurred. Our results suggest the chNHE1 protein is not only the receptor of ALV-J infected host but also play an important role in the process of tumor development. This study provides a scientific basis for further studying of oncogenic mechanism of ALV-J.
Animals ; Avian Leukosis ; genetics ; metabolism ; virology ; Avian Leukosis Virus ; genetics ; physiology ; Chickens ; genetics ; metabolism ; Poultry Diseases ; genetics ; metabolism ; virology ; Receptors, Virus ; genetics ; metabolism ; Sodium-Hydrogen Exchangers ; genetics ; metabolism ; Viral Load

This study was aimed to investigate whether the inhibition of NHE1 activity and intracellular acidification can reverse resistance of leukemia cells to the imatinib and to explore downstream signal molecule networks of BCR/ABL in the cells of chronic myelocytic leukemia (CML) patients. The mRNA and protein expression of P-glycoprotein (Pgp) and the drug accumulation were assayed after acidifying the primary leukemia cells of patients or K562/DOX and K562/G01 cells. The effects of intracellular acidification of primary leukemia cells on the phosphorylation level changes of ERK1/2 and p38 MAPK were analyzed by Western blot. The results showed that the intracellular concentration of drugs in the advanced patients increased and the sensitivity of K562/DOX and K562/G01 cells to imatinib was enhanced after intracellular acidification or treatment with NHE1 inhibitor cariporide. With downregulation of intracellular pH, the phosphorylation of p38 MAPK decreased in advanced patients and the phosphorylation of ERK1/2 increased within 3 min and then decreased after 30 min. SB203580, the specific inhibitor of p38 MAPK, displayed a synergistic effect with the inhibitor of NHE1 to downregulate the mRNA and protein expression of Pgp. It is concluded that the inhibiton of NHE1 can significantly decrease the protein expression of Pgp in K562/DOX and K562/G01 cells, increase the accumulation of Rhodamine123 and doxorubicin in the cells of advanced patients and enhance the sensitivity of cells to imatinib in which the p38 MAPK signal transduction pathways involves.
ATP-Binding Cassette, Sub-Family B, Member 1 ; metabolism ; Benzamides ; pharmacology ; Cation Transport Proteins ; antagonists & inhibitors ; metabolism ; Drug Resistance, Neoplasm ; drug effects ; Enzyme Inhibitors ; pharmacology ; Gene Expression Regulation, Leukemic ; Humans ; Imatinib Mesylate ; Imidazoles ; pharmacology ; K562 Cells ; MAP Kinase Signaling System ; Piperazines ; pharmacology ; Pyridines ; pharmacology ; Pyrimidines ; pharmacology ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; antagonists & inhibitors ; metabolism ; p38 Mitogen-Activated Protein Kinases ; antagonists & inhibitors ; metabolism

OBJECTIVETo investigate the effect of adenosine and its agonist on hypoxia-induced right ventricular hypertrophy (RVH) and explore the underlying mechanism.

METHODSFifty-six rats were randomly divided into normoxia group, hypoxia group, and treated hypoxia groups (with different treatments with adenosine, A1 receptor agonist CPA, A2 receptor agonist NECA, CPA plus A1 receptor inhibitor DPCPX, or NECA plus A2B receptor inhibitor MRS1754). The rats except for those in normoxia group were exposed to normobaric chronic hypoxia (9.5%-10.5% oxygen) for 21 days, and the corresponding treatments were administered since the 7th day of hypoxia till day 21 via implantable capsule with a pressure pump. After the treatments, the right ventricles were then removed and weighed for evaluation of hypertrophy, and the expressions of NHE-1 and CnAβ mRNA in the myocardial tissue were detected using RT-PCR.

RESULTSAfter a 21-day hypoxia, the rats showed significantly increased RV/(LV+S) ratio (0.369∓0.033) and RV/BW ratio (0.75∓0.095) compared to those in normoxia group (0.271∓0.010 and 0.59∓0.039, respectively; P<0.001), adenosine treatment group (0.281∓0.022 and 0.65∓0.077, respectively; P<0.001, P=0.025), hypoxia with CPA group (0.313∓0.021 and 0.66∓0.067, respectively P<0.001), and hypoxia with NECA group(0.333∓0.019, and 0.68∓0.074, respectively P<0.001). The NHE-1 and CnAβ mRNA levels in hypoxia group were significantly higher than those in normoxia group, adenosine treatment group, hypoxia with CPA group, and hypoxia with NECA group(P<0.001).

CONCLUSIONAdenosine and its agonist can inhibit hypoxia-induced RVH in rats through the NHE-1/CaN signal pathway.

Adenosine ; agonists ; pharmacology ; Animals ; Hypertrophy, Right Ventricular ; metabolism ; Hypoxia ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Signal Transduction ; drug effects ; Sodium-Hydrogen Exchangers ; metabolism

This study was aimed to investigate the expression of Na(+)/H(+) exchanger 1 (NHE1) in K562 and HL-60 cells undergoing DNA damage induced by etoposide and to elucidate the regulating mechanism. Real-time quantitative PCR (RQ-PCR) and Western blot methods were used to determine the expression of NHE1 in K562 cells after the treating with etoposide. Meanwhile, the flow cytometry was used to detect the apoptosis of leukemic cells. The luciferase reporter vector containing NHE1 promoter was constructed to measure relative luciferase activity after treating with different etoposide concentrations. The results showed that the mRNA and protein of NHE1 increased in accordance with apoptosis ratio in HL-60 cells after treated with etoposide (p < 0.05), but no such obvious increase in K562 cells. Treatment with NHE1 specific inhibitor could block etoposide induced alkalization and reduce the apoptosis ratio of HL-60 cells. The expression pattern and apoptosis alteration was not similar in K562 cells. Relative luciferase activity of reporter vector containing NHE1 promoter however increased in K562 cells after treated with etoposide. It is concluded that the expression of NHE1 is up-regulated in the process of apoptosis of HL-60 cells induced by etoposide and depends on the pHi increasing caused by NHE1 up-regulation which is not found in K562 cells although the transcriptional activity increased.
Apoptosis ; Cation Transport Proteins ; metabolism ; DNA Damage ; Etoposide ; HL-60 Cells ; Humans ; K562 Cells ; Promoter Regions, Genetic ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; metabolism

This study was purposed to investigate the effect of hypoxia microenvironment on K562 leukemic cell differentiation, and characteristics of NHE1 involvement in this process. The K562 cells were treated with hypoxia-mimical agent CoCl₂ or under actual hypoxia culture, and the specific NHE1 inhibitor Cariporide was used to inhibit NHE1 activity. The fluorescent probe BCECF was used for pH(i) measurements. Gene expression was analyzed by RT-PCR. The morphological characteristics was determined by Wright's staining. Signaling pathways were detected by Western blot using phosphospecific antibodies. The results indicated that the hypoxia or mimetic hypoxia favored K562 cells differentiation with up-regulation of C/EBPα. Moreover, treatment with Cariporide under hypoxia synergistically enhanced leukemia cell differentiation. Treatment with Cariporide increased levels of phosphorylated ERK5 and P38 mitogen-activated protein kinase (MAPK). It is concluded that the hypoxia or mimetic hypoxia can induce the differentiation of K562 cells, the inhibition of NHE1 activity can promote the hypoxia-induced K562 cell differentiation. The enhancement of hypoxia-induced K562 differentiation by Cariporide via MAPK signal pathway suggests a possible therapeutic target of NHE1 under hypoxia microenvironment in the treatment of leukemias.
Cation Transport Proteins ; metabolism ; Cell Differentiation ; Cell Hypoxia ; Humans ; K562 Cells ; MAP Kinase Signaling System ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; metabolism

This study was aimed to investigate the role of Na(+)/H(+) exchanger 1 (NHE1) in apoptosis of HL-60 cells induced by etoposide. Real-time quantitative PCR (RQ-PCR) and Western blot methods were used to determine the expression of NHE1 in HL-60 cells after the treatment with etoposide. Meanwhile, laser scanning confocal microscopy was used to test the intracellular pH (pHi) of HL-60 cells. Cell apoptosis was measured by DNA fragmentation and terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) assay. The results showed that etoposide induced cell apoptosis after treatment for 24 hours. The expression level of NHE1 mRNA increased by 2.848 +/- 0.886 times after treatment with etoposide for 12 hours (p < 0.01), and the expression of NHE1 protein was also up-regulated (p < 0.01). The pHi of HL-60 cells increased from 7.11 to 7.46 after treatment with etoposide for 24 hours. Treatment with cariporide could block etoposide-induced alkalinisation and enhance the apoptosis HL-60 cells. It is concluded that the expression of NHE1 is up-regulated in process of apoptosis of HL-60 cells induced by etoposide and the apoptosis depends on the pH increase caused by NHE1 higher expression.
Apoptosis ; drug effects ; Cation Transport Proteins ; genetics ; metabolism ; DNA Fragmentation ; Etoposide ; pharmacology ; Gene Expression Regulation, Leukemic ; HL-60 Cells ; Humans ; RNA, Messenger ; genetics ; Sodium-Hydrogen Exchanger 1 ; Sodium-Hydrogen Exchangers ; genetics ; metabolism