Anemia is frequently observed in cats and dogs with chronic kidney disease (CKD), typically presenting as normocytic, normochromic and non-regenerative. Prior reports indicate that approximately 30-65% of cats and 60% of dogs with CKD will develop anemia, with the prevalence and severity increasing alongside the progression of International Renal Interest Society (IRIS) stages (Table 1). [Chalhoub 2011, Elliott 1998, King JN, Lippi, King LG] Clinical manifestations of anemia in CKD patients include mucous membrane pallor, heart murmur, fatigue, listlessness, lethargy, weakness, pica, and loss of appetite.

The etiology of CKD associated anemia is multifactorial. The primary cause lies in the inadequate production of erythropoietin (EPO), a hormone crucial for regulating red blood cell (RBC) production. EPO stimulates erythropoiesis by binding to receptors on erythroid progenitors, promoting their differentiation into normoblasts and subsequently mature erythrocytes. [Chalhoub 2011] Other contributing factors to CKD-associated anemia include absolute and functional iron deficiency, inadequate nutrition, chronic inflammation or infection, medication effects, shortened red blood cell (RBC) lifespan, excessive blood sampling, spontaneous bleeding and parasite infestation (particularly fleas). [Chalhoub 2011, Borin-Criellenti]

Cats with CKD often exhibit relative iron deficiency and increased concentrations of hepcidin, a pivotal regulator in iron homeostasis. [Gest] It is well document that systemic inflammation occurs in association with CKD in people and early studies suggest that is also true in cats. [Begum, Gupta, Javard]. Hepcidin, which is upregulated in inflammatory conditions, induces iron sequestration within cells and limits GI absorption of iron. Cytokines also impede erythropoiesis during inflammatory states. While gastric ulceration is less frequent in companion animals compared to humans, chronic low-grade gastrointestinal hemorrhage due to mucosal fragility and uremic thrombocytopathia may contribute to anemia. [McLeland] Finally, the lifespan of RBCs may be shortened due to the adverse effects of uremic toxins. [Lau]

The production of RBCs in the bone marrow is governed by EPO, a hormone primarily synthesized by renal EPO-producing (REP) cells located in the corticomedullary interstitium. As CKD progresses, it is hypothesized that the number of active REP cells decreases, leading to reduced production of EPO; anemia is the result of insufficient stimulation of RBC progenitors in the bone marrow by EPO. Hypoxia-inducible factor (HIF), a master gene regulator that responds to reduced tissue oxygen concentrations, controls EPO production. [Spencer, Sugahara] During normoxia, HIFα is hydroxylated vial HIF-prolyl hydroxylases which use oxygen and 2-oxyglutarate as substrates. Following hydroxylation, HIFα is recognized by von Hippel-Lindau E3 ubiquitin ligase, resulting in the lysosomal degradation of HIFα. [Sugahara] However, during hypoxic states, HIFα remains unhydroxylated and translocates to the nucleus where it accumulates and combines with HIFβ to initiate EPO transcription. [Sugahara] Although the detailed mechanisms of EPO regulation are still being elucidated, recent studies suggest that impaired EPO production in CKD is not solely attributed to loss of functional renal tissue. [Dahl 2022 x2] One proposed theory involves microenvironmental relative hyperoxia surrounding REP cells, where nearby cell loss results in reduced oxygen utilization despite tissue hypoxia. [Dahl 2022 x2] Inflammation, a well-documented consequence of CKD in both humans and animals, can further suppress EPO effects and exacerbate anemia. [Gupta] Additionally, factors such as fibrosis mediators (TGF-β, NFκB, IL-6), FGF-23, and uremic toxins contribute to EPO suppression. [Lau, Shih, Afsar] Understanding the intricate regulation of HIF and EPO is crucial, especially considering therapeutic advances such as HIF-prolyl hydroxylase inhibitors (HIF-PHIs), which enhance endogenous EPO production by inhibiting HIFα hydroxylation even in normoxic conditions and possibly correcting the dysregulation of REP cells that occurs in CKD. [Locatelli]. These drugs, long utilized in human medicine and now emerging in veterinary care, represent a promising avenue for managing anemia associated with CKD [Charles].

Several studies have highlighted anemia as a significant predictor of poor survival in CKD. [King JN, King LG, Chakrabarti, Geddes] Even minor reductions in PCV, such as a median decrease from 35% to 31%, are associated with disease progression. [Chakrabarti] Anemia triggers physiological responses, including heightened release of norepinephrine, renin, angiotensin II, and aldosterone, which can increase cardiac workload and induce hypertension. [Hammond] Furthermore, anemia may contribute to left ventricular hypertrophy, predisposing patients to heart failure and fluid overload. [Chalhoub, Hammond, Wilson] Timely correction of anemia is crucial for resolving high-output heart failure secondary to anemia. [Hammond]

Anemia exacerbates CKD progression by compromising tissue oxygen delivery. Hypoxia plays a pivotal role in fibrosis formation and is, therefore, considered to be a critical mediator in CKD progression. [Elliott 2023, Nangaku, Spencer] Inflammatory and fibrotic changes that occur in CKD lead to interstitial expansion and increase the diffusion distance and compromise tubular cell access to peritubular capillary blood supply. Additionally, glomerular damage and vasoconstriction further reduce post-glomerular peritubular capillary blood flow. [Nangaku] Vascular endothelial growth factor (VEGF), essential for vascular health, may become depleted in CKD, contributing to capillary rarefaction, or the loss of peritubular capillaries, thus reducing the density of blood supply to the tubules. [Mayer, Habenicht, Lourenco, Paschall]  Hypoxia, developing as a result of both the compromised blood supply and the anemia, leads to fibrosis, tubular cell transdifferentiation, and activation of fibroblasts, thereby worsening the diseased state. [Nangaku, Spencer] Therefore, resolving anemia should be seen as a crucial therapeutic goal, especially in advanced CKD stages where fibrosis and capillary rarefaction are prominent. [Elliott 2023, Paschall, Chakrabarti, McLeland].  However, level 1 evidence that resolving anemia in CKD slows progression is currently lacking, even in human CKD.

Notably, moderate to severe anemia significantly impacts quality of life in feline CKD patients, manifesting as weakness, lethargy, and reduced appetite. Anemia (PCV <27%) correlates with lower health-related quality of life scores in cats with CKD. [Lorbach]

Transfusion

Blood transfusion has long been employed to address severe, symptomatic anemia (Table 2). Such conditions may occur in patients newly diagnosed with advanced CKD or acute on chronic crisis, those with untreated or inadequately managed CKD-related anemia, or those experiencing acute hemorrhage (e.g., gastrointestinal bleeding). Blood transfusion offers the advantage of promptly correcting anemia. However, potential drawbacks include the risk of hypersensitivity reactions or acute lung injury, volume overload from rapid administration, and shortened survival of red blood cells in a uremic environment. When transfusion is necessary, the objective is to restore the packed cell volume to the lower end of the normal range, followed by a longer-term therapy to maintain red blood cell mass.

Erythropoiesis-Stimulating Agents (ESAs)

Veterinarians have utilized ESAs, such as EPO analogues including epoetin and darbepoetin, for many years. Epoetin alfa (Epogen®, Amgen; Procrit®, Janssen), a human recombinant EPO, was associated with immunogenic reactions causing pure red cell aplasia in 20-25% of dogs and cats, leading to its discontinuation. Darbepoetin, another human recombinant EPO that is hyperglycosylated for extended half-life and reduced immunogenicity, is commonly employed in managing CKD associated anemia (CKD) (Figure 1). [Chalhoub 2012, Fiocchi] In a retrospective study, 25 cats with CKD associated anemia were treated with darbepoetin and available for 8 weeks of follow-up, 56% responded by achieving and maintaining a packed cell volume (PCV) of ≥ 25% by day 56, with a median PCV increase of 8%. [Chalhoub 2012] Most of these cats (21 out of 25) received a maximum dose of 1 ug/kg subcutaneously weekly at some point during treatment. Some of these cats also received blood transfusions and iron dextran as part of their management. Among responder cats, the median time to response was 21 days (range, 7-47 days). It is worth noting that these 25 cats were selected from 66 cases that received darbepoetin, the remainder were not available for full follow-up. In a similar study of 33 dogs with CKD treated with darbepoetin, 85% responded by reaching a PCV of 30% or higher, with a median maximum weekly dose of 0.8 ug/kg subcutaneously. [Fiocchi] The median time to response in dogs was 29 days (range, 6-106 days). Monitoring is required to prevent complications. In cats, 41% of cats were noted to be hypertensive during treatment and three of four cats experiencing seizures or acute neurologic events were hypertensive. However less than half of cats had blood pressure measured prior to treatment. Among dogs, hypertension occurred in 24 out of 25 dogs requiring treatment in 13 cases, while mild hyperkalemia affected 42%, not requiring intervention. Seizures were observed in 5 dogs, and pure red cell aplasia was suspected but not proven in one case. Darbepoetin is not FDA approved for use in dogs or cats. Recombinant EPO products for cats and dogs have been investigated in preliminary studies but it remains unlikely that these will become commercially available. [Randolph 2004 x 2]

Hypoxia-inducible factor prolyl hydroxylase inhibitors (HIF-PHI)

An HIF-PHI for treating CKD associated anemia in cats, molidustat (VarenzinTM-CA1, Elanco), received conditional approval from the FDA in 2023. As previously mentioned, HIF-PHIs such as molidustat inhibit prolyl hydroxylase activity even under normoxic conditions, leading to sustained HIFα levels and continuous EPO production. [Flamme, Beck, Haase, Hirota, Gupta] In addition these drugs have been shown to suppress hepcidin and upregulate iron metabolism and transport in rats and people and are as effective in patients with systemic inflammation as those without. [Li]

In a study involving healthy cats, molidustat administered orally at 5 mg/kg daily resulted in a significant increase in hematocrit relative to baseline and significantly higher than placebo-treated cats by day 14 (PCV at day 14: 54.4% vs. 40.3%). [Boegel] Treatment was halted by day 23 due to hematocrit levels exceeding 60%, which gradually returned to baseline over the following 5 weeks. EPO concentrations peaked significantly 6 hours after molidustat administration but returned to baseline within 24 hours. Vomiting occurred more frequently in the treated group compared to the placebo group.

In a pilot study involving client-owned cats with CKD and a hematocrit below 28%, 15 cats received 5 mg/kg molidustat daily while 6 received placebo. [Charles] Treated cats showed higher hematocrit levels than placebo cats by day 21. However, only 50% of molidustat-treated cats met the criteria for treatment success, defined as a 4% increase in hematocrit or a 25% increase from baseline, over the 28-day study period. Among these, hematocrit increased from 23.6% ± 3.23 at day 0 to 27.8% ± 5.25 at day 28. Eight cats continued molidustat treatment after the initial study, at doses ranging from 2.5 to 5 mg/kg daily. By day 56, 75% of these cats were considered treatment successes. Vomiting was the most commonly reported adverse event, occurring in 6 of 15 treated cats, with varying frequencies.

Molidustat's FDA conditional approval prohibits off-label use. The recommended regimen includes daily treatment for 28 days followed by a 7-day discontinuation period, with potential for repeated cycles if necessary (Figure 2). While healthy cats showed risk of polycythemia without treatment breaks, this has not been observed in cats with CKD. [Boegel, Charles]

Gene Therapy

Recently, researchers evaluated the use of an adeno-associated virus vectored (AAV)-based gene therapy that enables the expression of feline EPO in 23 cats. [Vaden] A single intramuscular injection resulted in the desired increase in packed cell volume (PCV) within 28-70 days for 86% of the cats. However, polycythemia developed in 3 cats. While the therapy was generally well tolerated, some cats experienced hypertension and encephalopathy, similar to that described in association with other ESAs. Further investigation of this treatment is necessary to fully understand its effectiveness and safety profile, including whether repeat doses are necessary to maintain a satisfactory response.

Iron plays a critical role in oxygen transport, cellular respiration, and DNA synthesis and repair due to its electron donation and acceptance capabilities. Despite its abundance, the body meticulously regulates iron absorption and storage to prevent the formation of harmful reactive oxygen species from iron excess. The concentration of iron in the bloodstream is governed by three primary mechanisms: absorption of dietary iron in the duodenum and upper jejunum, recycling of iron by the reticuloendothelial system, and release of iron from hepatocytes. [Lane]

Ingested iron typically enters enterocytes in its ferrous form (Fe2+). Ferric iron (Fe3+), when ingested, is first converted to ferrous iron by the ascorbate-dependent hemoprotein duodenal cytochrome b (DcytB) before being absorbed. [Lane, Anderson] Certain dietary compounds can bind to iron, hindering its absorption. Once absorbed, most iron that isn't immediately needed is stored within enterocytes as ferritin until the cell is shed and the iron is excreted in feces. When required, iron crosses the enterocyte basolateral membrane via ferroportin, binding to plasma transferrin for transport to various tissues. The proteins involved in iron uptake from the gut, its transport in blood and uptake into the tissues are upregulated by HIF in response to hypoxic states.  Iron is stored predominantly as ferritin or hemosiderin in organs such as the liver, spleen, bone marrow, duodenum, and skeletal muscles. However, the largest source of iron in the body is found within heme, encompassing hemoglobin, myoglobin, and hemoproteins. Iron is lost from the body through the shedding of enterocytes, uroepithelial cells, and epidermal cells.

Iron regulatory proteins optimize iron supply during deficiency while restraining absorption and promoting storage when iron stores are sufficient. Crucially, hepcidin, an acute phase reactant protein produced in the liver, downregulates the system when iron stores are replete. Hepcidin inhibits intestinal iron absorption and intracellular iron transport, including release from macrophages and hepatocytes, thereby sequestering iron. Reduced hepcidin concentrations occur in conditions such as anemia, hypoxia, or iron deficiency, whereas increased concentrations are associated with adequate or increased body iron stores, systemic inflammatory states, or reduced glomerular filtration rate (due to hepcidin's renal elimination and metabolism). [Zaritsky, Babitt]

CKD often disrupts iron homeostasis through mechanisms such as impaired dietary iron absorption and reticuloendothelial cell iron blockade, resulting in functional iron deficiency (FID) despite adequate iron stores; hepcidin is a major contributor to these events. [Babitt] CKD patients may also experience absolute iron deficiency (AID) due to increased iron losses from conditions like platelet dysfunction induced chronic hemorrhage or frequent blood sampling. Blood trapping in the dialysis apparatus may contribute to reduced iron stores in dialysis patients. Administration of an ESA may exacerbate iron depletion by enhancing erythropoiesis, creating a mismatch between iron supply and demand. Distinguishing FID induced by an ESA from anemia of chronic disease can be challenging.

Traditional methods of assessing iron stores are problematic in CKD patients. [Batchelor, Gaweda, Gafter-Gvili] Serum iron concentrations do not reliably reflect total body iron. Ferritin is a better indicator of total body iron in health it is also an acute phase reactant that can be increased in inflammation thereby masking AID. Transferrin concentrations, gauged through total iron binding capacity (TIBC), may appear normal or high in AID but because it is a negative acute phase reactant it decreases during inflammation, potentially obscuring AID. Percent saturation or transferrin saturation (TSAT) is the ratio of serum iron to TIBC, or the percentage of transferrin that is occupied by iron.  TSAT indicate iron deficiency when low but cannot differentiate between AID and IFD. Because increased TSAT correlates with iron overload, TSAT can help guide iron therapy by indicating when iron should not be administered. ESAs can strip iron from transferrin faster than it can be mobilized, reducing TSAT. Although patients with FID would be expected to have low serum transferrin and high to normal ferritin, none of these parameters consistently differentiate between AID and FID in patients with CKD associated anemia. Semiquantitative assessment of iron stores in bone marrow or liver with Prussian blue staining remains the gold standard despite its invasiveness. Animals with AID would have an absence or iron fragments in the bone marrow while those with FID would be expected to have adequate iron stores. 

Novel biomarkers proposed for assessing iron stores in CKD include soluble transferrin receptor, percentage of hypochromic red blood cells, reticulocyte hemoglobin content, hepcidin, and plasma neutrophil gelatinase-associated lipocalin (NGAL) and offer potential insights into iron stores beyond traditional assays. [Batchelor] Reticulocyte hemoglobin concentration was found to add some value to the diagnosis of iron deficiency diagnosed by TSAT in cats, whereas the MCV of reticulocyte and standard erythrocyte indices did not contribute to the diagnosis. [Betting]

Iron supplementation is pivotal in managing CKD-associated anemia and has demonstrated benefits in physical, cognitive, and immune functions in humans with CKD. [Hanna] However, optimal management of iron therapy remains uncertain. Oral iron preparations are widely available, with iron citrate possibly offering additional benefits as a phosphate binder. Oral iron can provoke gastrointestinal upset, anorexia, and alter the microbiome, promoting the growth of potentially pathogenic bacteria while reducing the population of protective lactobacilli and bifidobacteria. [Anderson] Parenteral iron (intravenous for humans, intramuscular for dogs and cats) bypasses poor absorption and gastrointestinal side effects but carries a small risk of iron overload. In people, IV iron therapy may be more effective than oral therapy, but the comparable safety remains debatable. [Shepshelovich] The use of IV iron has not be studied in dogs and cats.

Strategies aimed at modulating hepcidin may enhance iron absorption from the gut and mobilization within the body. [Begum] Inhibiting molecules involved in hepcidin production, such as IL-6 antibodies, TNF-alpha and TGF-gamma inhibitors, or pentoxifylline, could potentially optimize iron availability. Direct hepcidin antagonists and hepcidin-ferroportin stabilizers are under evaluation for therapeutic use in people. HIF-PH inhibitors down-regulated hepcidin, thereby allowing more iron absorption for the gastrointestinal tract and release from the reticuloendothelial system and could improve iron metabolism in cats with anemia and facilitate oral iron therapy. However, this theoretical benefit of HIF-PH inhibitors remains to be studied in feline CKD patients.

Due to the risks and cost associated with previously available treatments for anemia, treatment has often been postponed until anemia reaches severe or symptomatic levels. However, considering evidence linking even mild anemia with disease progression, future research should investigate whether earlier treatment of anemia could help preserve renal function. The emergence of new drugs for anemia allows for more personalized therapy. Darbepoetin has demonstrated success in most treated patients. HIF-PH inhibitors also hold promise as a valuable approach to halt the progression of mild anemia in cats. The significance of functional iron deficiency is gaining recognition, with new biomarkers potentially offering more precise assessments of iron status in anemic patients. Advances in understanding hepcidin regulation may lead to the development of more effective strategies for iron replacement.

Note that these studies defined anemia as a hematocrit of <25% for cats and <37% for dogs. The true incidence of anemia is likely higher if defined as a hematocrit below the reference range.

IRIS stage 2

Cats 4.8%

Dogs 47%

IRIS stage 3

Cats 17.7%

Dogs 71%

IRIS stage 4

Cats 53.3%

Dogs 82%

Blood transfusion: Replace RBCs

                               Packed RBCs: 6-10 ml/kg IV, whole Blood: 12-20 ml/kg IV

Darbepoetin: Hyperglycosylated human recombinant erythropoietin.

                      Starting dose: 0.45-1 mcg/kg SQ weekly

                      Maintenance dose: 0.45-1 mcg/kg SQ q 2-3 weeks

Feline erythropoietin gene therapy:   Adeno-associated virus vectored gene therapy.

                                                             In development.

                                                             Given by IM injection.

Molidustat: Hypoxia inducible factor – prolyl hydroxylase inhibitor                                

                   5 mg/kg PO daily for 28 days, stop for 7 days

                   Repeat cycle as required for conditional approval period.

Absolute iron deficiency

Serum iron: reduced

Transferrin: Normal to increased

Percent saturation: reduced

Ferritin: reduced

Bone marrow iron stores: reduced

Hepcidin: reduced

Functional iron deficiency

Serum iron: reduced

Transferrin: normal to decreased

Percent saturation: normal to decreased

Ferritin: normal to increased

Bone marrow iron stores: increased

Hepcidin: increased

Caveats

Serum iron does not reflect total body iron.

Transferrin is indirectly measured by total iron binding capacity (TIBC).

The presence of inflammation can obscure a diagnosis of absolute iron deficiency.

Assays for the presence of ferritin and hepcidin are not readily available for dogs and cats.

Iron staining of bone marrow, while semi quantitative, is the gold standard for assessment of iron stores.