Erythropoietin

Unlike other growth factors, erythropoietin is mainly synthesised by the peritubular endothelial cells of the kidney. Production is triggered by tissue hypoxia (lack of oxygen). Cells can sense hypoxia via mediators such as the transcription factor HIF (hypoxia-inducible factor). HIF activates genes vital in the adaptive response to hypoxia including the erythropoietin gene. Erythropoietin molecules bind to specific membrane receptors on primitive erythroid cells in the bone marrow and induce maturation. The increase in red cells released into the blood stops when normal oxygen transport is restored – this feedback circuit is illustrated in Figure 2.2.

Structure

The mature red cell is around 7.8 µm across and 1.7 µm thick. Its biconcave shape allows maximum flexibility and an umbrella shape is adopted to traverse the smallest capillaries which have diameters of only 5 µm. The ability of red cells to recover from the recurrent stresses of the turbulent circulation hinges on the design of the membrane.

The red cell membrane is composed of a collapsible lattice of specialised proteins (the ‘cytoskeleton’) and an outer lipid bilayer (Fig 2.3). The protein skeleton is responsible for maintaining red cell shape while the lipid bilayer provides a hydrophobic skin. The main skeletal proteins are spectrin, actin, proteins 4.1 and 4.2, and ankyrin. Spectrin is the most abundant and consists of alpha and beta chains wound around each other. Spectrin heterodimers can align at the ends to form tetramers (i.e. four chains). Spectrin tetramers are joined together by actin in association with protein 4.1. This flexible skeleton is attached to the rest of the membrane by ankyrin, which interacts with protein 4.2 to link the spectrin beta chain to the cytoplasmic end of the transmembrane protein band 3. The lipid bilayer consists mainly of a mixture of phospholipids and cholesterol. Cholesterol molecules are inserted between phospholipid molecules in such a way that they stiffen the membrane while still allowing a degree of fluidity between the bilayers.

Defects of both the red cell membrane proteins and lipids may lead to changes in red cell shape and premature destruction.

Metabolism

Red cells require an energy source to maintain their structure and also a mechanism for detoxification of oxidants. Energy is provided by the Embden–Meyerhof pathway, a sequence of biochemical reactions in which glucose is metabolised to lactate with the generation of two molecules of adenosine triphosphate (ATP). ATP maintains the osmotic pressure of the cell by driving sodium and calcium pumps in the membrane. It also provides energy for the cytoskeletal changes needed for recovery of cell shape. The Embden–Meyerhof pathway does not require oxygen as a substrate but a small amount of oxidative glycolysis occurs by the hexose monophosphate shunt (pentose phosphate pathway) in which glucose-6-phosphate is metabolised to generate nicotinamide adenine dinucleotide phosphate (NADPH). The hexose monophosphate shunt plays a vital role in oxygen detoxification and when oxidised substrates accumulate in the cell it increases activity several fold. Inherited deficiencies of red cell enzymes in either the Embden–Meyerhof pathway (e.g. pyruvate kinase) or the hexose monophosphate shunt (e.g. glucose-6-phosphate dehydrogenase) can lead to shortened red cell survival and haemolytic anaemia (see p. 29).

Haemoglobin and oxygen transport

The key function of red cells, to carry oxygen to the tissues and return CO₂ from the tissues to the lungs, depends on the specialised protein haemoglobin which is present in large amounts in mature cells. The normal adult haemoglobin molecule (HbA) contains four polypeptide chains (‘globin’ chains): the two alpha chains and two beta chains are often notated as α₂β₂. Combined with each of the polypeptide chains is a ‘haem’ molecule which contains ferrous iron (Fe²⁺) and protoporphyrin (Fig 2.4). The iron combines reversibly with oxygen and thus haem forms the oxygen-carrying part of the molecule. Other globin chains are formed by the fetus and the change from fetal to adult haemoglobin occurs in the first 3–6 months of life. However, the subunits designated γ and δ persist into later life and small amounts of fetal haemoglobin (HbF; α₂γ₂) and HbA₂ (α₂δ₂) are found in adults.

Haemoglobin is more than an inert carrier molecule. The individual globin chains interact with each other to facilitate the offloading of oxygen at lower oxygen saturations. The metabolite 2,3-diphosphoglyceride (2,3-DPG) generated in a side-arm of the Embden–Meyerhof pathway has an important role in the process, which results in a sigmoid-shaped oxygen dissociation curve (Fig 2.5). In anatomical terms haemoglobin has a high affinity for oxygen in the lungs and a much lower affinity in the tissues. The oxygen dissociation curve moves to the left when oxygen affinity increases; this occurs when H⁺ ion concentration is reduced or haemoglobin F (which cannot bind 2,3-DPG) raised. The curve moves to the right when oxygen affinity decreases; for instance when 2,3-DPG concentration rises or the abnormal sickle haemoglobin (HbS) is present. The P₅₀ level is defined as the partial pressure of oxygen at which haemoglobin is half saturated.

Ageing and death

Beyond 100 days red cells start to show features of ageing including a declining rate of glycolysis, reduced levels of ATP and membrane lipid, and a loss of flexibility. The terminal event is unclear but effete cells are removed from the circulation by the macrophages of the liver and spleen.

Most of the catabolised haemoglobin, particularly the iron, is reused (see also p. 24). The protoporphyrin of haem is metabolised to the yellow pigment bilirubin which is bound to albumin in the plasma. Bilirubin is conjugated in the liver to a water-soluble diglucuronide that is converted to stercobilin and stercobilinogen and excreted in the faeces. Some stercobilin and stercobilinogen are reabsorbed from the intestine and excreted in the urine as urobilin and urobilinogen.