Glucose transport across lagomorph jejunum epithelium is modulated by AMP-activated protein kinase under hypoxia

Abstract

The gastrointestinal epithelium possesses adaptation mechanisms to cope with huge variations in blood flow and subsequently oxygenation. Since sufficient energy supply is crucial under hypoxic conditions, glucose uptake especially must be regulated by these adaptation mechanisms. Therefore, we investigated glucose transport under hypoxic conditions. Jejunal epithelia of rabbits were incubated in Ussing chambers under short-circuit current conditions. Hypoxia was simulated by gassing with 1% O instead of 100% O. The activity of sodium-coupled glucose transporter-1 (SGLT-1) was assessed by measuring the increase of short circuit current ( I) after the addition of 2 mM glucose to the mucosal buffer solution. We observed decreased activity of SGLT-1 after hypoxia compared with control conditions. To investigate underlying mechanisms, epithelia were exposed to agonists and antagonists of AMP-activated protein kinase (AMPK) before assessment of SGLT-1-mediated transport and the pAMPK/AMPK protein ratio. Preincubation with the antagonist restored SGLT-1 activity under hypoxic conditions to the level of control conditions, indicating an involvement of AMPK in the downregulation of SGLT-1 activity under hypoxia, which was confirmed in Western blot analysis of pAMPK/AMPK. Transepithelial flux studies using radioactively labeled glucose, ortho-methyl-glucose, fructose, and mannitol revealed no changes after hypoxic incubation. Therefore, we could exclude a decreased transepithelial glucose transport rate and increased paracellular conductance under hypoxia. In conclusion, our study hints at a decreased activity of SGLT-1 under hypoxic conditions in an AMPK-dependent manner. However, transepithelial transport of glucose is maintained. Therefore, we suggest other transport mechanisms, especially glucose transporter 1 and/or 2 to substitute SGLT-1 under hypoxia. NEW & NOTEWORTHY To our knowledge, this is the first approach to simulate hypoxia and study its effects in the jejunal epithelium using the Ussing chamber technique. We were able show that AMPK plays a role in the downregulation of SGLT-1 and that there seems to be an upregulation of other glucose transport mechanisms in the apical membrane of lagomorph jejunum epithelium under hypoxia, securing the epithelial energy supply and thus integrity.

The Fast Lane of Hypoxic Adaptation: Glucose Transport Is Modulated via A HIF-Hydroxylase-AMPK-Axis in Jejunum Epithelium

“…Kles and Tappenden [7] reported that the transepithelial transport of glucose was impaired under hypoxia, while glutamine transport was not. In accordance with that, it has been shown that the activity of the sodium-coupled glucose transporter SGLT1 was diminished under hypoxia in jejunum epithelium [5,8,9]. This downregulation appears to be a targeted and reasoned adaptation rather than a sign of cellular failure.…”

“…However, for the epithelium’s survival under challenging circumstances such as a sudden onset of hypoxia, an adaptation within minutes is necessary. Recent research provided evidence for a fast adaptation to hypoxia taking place on the protein rather than on the gene expression level [5,6]. This quick reaction might at least bridge the critical time until HIF-mediated adaptation mechanisms are set to work and could, thus, be a critical point for early therapeutic intervention.…”

“…This downregulation appears to be a targeted and reasoned adaptation rather than a sign of cellular failure. Accordingly, the total transepithelial transport of glucose is sustained in spite of the missing import via SGLT1 [5]. Some authors postulate a recruitment of glucose transporter 2 (GLUT2) to the apical membrane in order to enhance the energy-independent uptake of glucose [9,10,11].…”

“…While the exact transport systems involved in this adaptation are still under discussion and need further investigation, it is also not clear yet by which intracellular mechanism this fast adaptation is mediated. Our own previous work indicated an involvement of the AMP-activated protein kinase (AMPK) in the modulation of transepithelial glucose transport in the lagomorph jejunum epithelium [5]. An activation of AMPK under hypoxia has also been postulated in human airway epithelial cells, HeLa cells, as well as prostate carcinoma cells [12,13,14].…”

Activation of AMPK under Hypoxia: Many Roads Leading to Rome

“…An activation of AMPK under hypoxia has been demonstrated in various tissues and cell types, ranging from brain [32][33][34], liver [35] and skeletal muscle [36][37][38] to cardiomyocytes [39,40], alveolar epithelial cells [41,42], intestinal epithelial cells [43,44] and fibroblasts [45]. Interestingly, an inhibition of AMPK activation under hypoxia was reported after long-term hypoxia (in this case 8% oxygen for 12 d) [46].…”