Human skeletal muscle and erythrocyte proteins involved in acid-base homeostasis: adaptations to chronic hypoxia

Abstract

Chronic hypoxia is accompanied by changes in blood and skeletal muscle acid-base control. We hypothesized that the underlying mechanisms include altered protein expression of transport systems and the enzymes involved in lactate, HCO3- and H+ fluxes in skeletal muscle and erythrocytes. Immunoblotting was used to quantify densities of the transport systems and enzymes. Muscle and erythrocyte samples were obtained from eight Danish lowlanders at sea level and after 2 and 8 weeks at 4100 m (Bolivia). For comparison, samples were obtained from eight Bolivian natives. In muscle membranes there were no changes in fibre-type distribution, lactate dehydrogenase isoforms, Na+,K+-pump subunits or in the lactate-H+ co-transporters MCT1 and MCT4. The Na+-H+ exchanger protein NHE1 was elevated by 39 % in natives compared to lowlanders. The Na+-HCO3- co-transporter density in muscle was elevated by 47-69 % after 2 and 8 weeks at altitude. The membrane-bound carbonic anhydrase (CA) IV in muscle increased in the lowlanders by 39 %, whereas CA XIV decreased by 23-47 %. Levels of cytosolic CA II and III in muscle and CA I and II in erythrocytes were unchanged. The erythrocyte lactate-H+ co-transporter MCT1 increased by 230-405 % in lowlanders and was 324 % higher in natives. The erythrocyte inorganic anion exchanger (Cl--HCO3- exchanger AE1) was increased by 149-228 %. In conclusion, chronic hypoxia induces dramatic changes in erythrocyte proteins, but only moderate changes in muscle proteins involved in acid-base control. Together, these changes suggest a hypoxia-induced increase in the capacity for lactate, HCO3- and H+ fluxes from muscle to blood and from blood to erythrocytes.

Figures

Figure 1. Muscle and blood pH regulation

Summary of membrane-bound transport systems (circles) and enzymes (squares) involved in the H+ fluxes between muscle and blood. Carbonic anhydrase (CA) I, II, III, IV and XIV represent cytosolic and membrane-bound CA isoforms present in muscle and erythrocytes. NHE1: Na+–H+ exchanger isoform 1. MCT1 and MCT4: monocarboxylate transporter (lactate-H+ co-transporter) isoform 1 and 4. NBC: Na+-HCO3− co-transporter. AE1: Cl−-HCO3− exchanger 1.

Figure 2. Histochemical properties of muscle samples

The bars to the left represent means (±s.e.m., n = 8) of the relative occurrence of the three myosin heavy chain isoforms (I, IIA and IIX) in the muscle samples. The bars to the right represent the percentage (of total LDH) of the lactate dehydrogenase heart-form (LDH-H). White bars: muscle samples obtained at sea level. Light grey bars: muscle samples obtained after 2 weeks at altitude. Dark grey bars: muscle samples obtained after 8 weeks at altitude. Black bars: muscle samples from Bolivian natives.

Figure 3. Muscle membrane transporters

The bars represent the mean density (n = 8, ±s.e.m.) of various membrane-bound transporter proteins. The values are calculated relative to the density in membrane samples obtained from lowlanders before the stay at altitude stay (sea level samples; white bars). Light grey bars: samples obtained after 2 weeks at altitude. Dark grey bars: samples obtained after 8 weeks at altitude. Black bars: samples from Bolivian natives. Abbreviations (from left to right): MCT1: monocarboxylate (lactate-H+) transporter 1. MCT4: monocarboxylate transporter 4. α1: Na+,K+-pump subunit α1. α2: Na+,K+-pump subunit α2. β1: Na+,K+-pump subunit β1. Na/H: Na+–H+ exchanger protein NHE1. NBC: Na+-HCO3− co-transporter. * Significantly different from sea-level values (P < 0.05).

Figure 4. Muscle CAs

The CA isoforms CA II and CA III were quantified in the cytosolic fraction, whereas the isoforms CA IV and CA XIV were quantified in the membrane fraction. The mean values are calculated relative to the density in muscle samples obtained at sea level. Symbols as in Fig. 3. *Significantly different from sea-level values (P < 0.05).

Figure 5. Erythrocyte enzymes and transporters

CA isoforms CA I and CA II were quantified in the cytosolic fraction of blood and calculated relative to the haemoglobin concentration. The glucose transporter GLUT1, the monocarboxylate transporter isoform MCT1 and the inorganic anion exchanger AE1 were quantified in erythrocyte membranes (arbitrary density units per milligram of membrane protein). Symbols as in Fig. 3. Note different y-axes. * Significantly different (P < 0.05) from sea-level samples.