Renal anemia is a major complication in chronic kidney disease (CKD), and is a problem that has yet to be overcome. This problem had been improved by the recombinant human erythropoietin (rhEPO), but some patients are relatively resistant to it and require higher doses, which is associated with increased adverse outcomes and mortality [1]. Furthermore, the consequence of higher demand for rhEPO to achieve target Hb level is costly. Some factors can be considered as the reasons of rhEPO resistance; blood loss, impaired hematopoiesis and inadequate dialysis. Moreover, there is a hypothesis that enhancement of the immune system activity in CKD patients is one of the important causes of resistance to EPO [2], and the use of anti-inflammatory drugs like pentoxifylline (PTX) may be useful as a methylxanthine derivate used for the management of chronic peripheral arterial disease that increases the blood flow to the microcirculation because of its potent hemorheological properties, which include preservation of the erythrocyte water and cation content and enhanced tissue oxygenation [3]. Also, anti-inflammatory effects, anti-apoptotic, anti-oxidant, and anti-TNF-α/anti-IFN-γ actions are important properties of PTX [4,5].

Although some non-randomized clinical trials [6,7] showed that PTX was able to increase the hemoglobin (Hb) levels in CKD patients significantly, there is no substantial evidence supporting the effectiveness of PTX for improving anemia control in hemodialysis (HD) patients [8].

Due to paucity of information on the PTX effect in HD population, we designed a double-blind randomized clinical trial study to determine the beneficial effect of PTX on anemia and prescription rhEPO dose in HD patients.

Participants and sampling

This double blinded clinical trial study was conducted in Haj Ebrahimi dialysis center affiliated with Shiraz University between March 2018 and June 2018. Participants were recruited from patients who were dialyzed three times a week for 4 hours by Fresenius 4008B machinery, bicarbonate dialysate and high flux polysulphone membrane. The inclusion criteria was the age of 18 years old or more, patients receiving 4-hour HD treatments 3 times per week at least for 3 months, Hb<11mg/dl in at least 2 consecutive tests in a month, Kt/V>1.2, transferrin saturation>20%, and no change in rhEPO dose for at least 1 month before starting the study. Exclusion criteria included intolerance of the drug in oral form and blood transfusion due to any reason in the recent 3 months, active infection, recent bleeding, and consumption of steroid androgen and theophylline. We collected 60 patients who met the study criteria that randomized into similar size groups of trial (PTX) and control as blocking randomization was used for allocation sequence.

Study protocol

We divided the patients into 2 groups, initially each group contained 30 members. Patients in the trial group received PTX (400 mg, Pentoxifylline®, extended release, FARABI, Pharmaceutical Company) twice a day for 12 weeks, whereas the control group received placebo (starch, magnesium, lactose, and talcum powder) twice a day for 12 weeks. The type and dose of the individualized drugs remained unchanged during the study. Also, all patients in both groups received rh-Erythropoietin 4000 IU solution for injection as a synthetic PD poietin (Pooyesh daru®; Tehran, Iran) and Nephrovit tablet (Nephrovit®; OSVAH, Tehran, Iran) as a previous regimen. All patients had an AV fistula with the membrane and similar dialysis prescription. Each patient was given an order number and received the medications in the corresponding prepacked bottles. Clinical investigators, laboratory personnel, and patients were all masked to the treatment assignment. Patients were observed closely during the course of the study as to the developing side effects, drug intolerance, and other complaints and signs. During the course of the study, 3 patients in the PTX group were excluded due to adverse events, whom were referred to specialist.

Data collection

A questionnaire was prepared for each patient including demographic data such as age, sex, and also questions about their underlying diseases, past history, and drug history. For all the patients, blood samples were taken to measure the plasma erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), and albumin (Alb), at the beginning and end of the study, which were immediately centrifuged and frozen at -70°C. Hb levels were measured monthly and rhEPO dose titration was done. The dose of rhEPO reduced about 25% from the baseline if Hb raised to more than 11.5 mg/dl, while the EPO dose was increased to 25% if Hb remained constantly less than 10 mg/dl.

Ethical consideration

The protocol was in accordance with the Declaration of Helsinki and the local ethics committee (Ethical code: CT-P-9374-7064). All patients provided informed consent before the study. It was registered at Clinical-Trials.gov with the code of IRCT-20100413003698N2.

Statistical analysis

Data were analyzed using SPSS-18 for windows. Chi-square was used to assess the relationship of two qualitative data and independent t-test and Pair t-Test were used to evaluate the correlation of a quantitative data. Also, we used Mann-Whitney U Test and Wilcoxon Test as nonparametric tests if appropriate. P-value <0.05 was considered significant.

We started with 60 patients that were divided into PTX and control groups. During the study, 3 patients in the PTX group were excluded due to adverse events: abdominal discomfort, gastrointestinal and conjunctival hemorrhage. So, we gathered 57 patients with a mean age of 54.1 ± 13.8 years old with dialysis duration of 21.4 ± 4.1 months. 52.6% (30 cases) of the patients were female. The mean KT/V at the initiation and termination of the study was 1.4 ± 0.8 and 1.3 ± 0.5, implying the efficient dialysis. The most common underlying cause was hypertension in 27 patients (47.4%) and diabetes mellitus in 12 patients (21.0%). The mean age in the PTX group was 49.1 ± 13.4 years old and 12 cases were female (40.7%), and the mean age in the control group was 52.6 ± 8.2 years old with 18 female cases (60.0%). As shown in (Tables 1 and 2), there was no difference in demographic data and laboratory tests of both groups at the beginning of the study, whereas at the end of the study there was a significant difference in the level of EPO dose, CRP and albumin between the PTX and control groups.

Hb levels increased significantly after treatment in both groups (9.9 ± 1.0 mg/dl vs. 10.7 ± 0.8 mg/dl, p=0.016 in the PTX group and 9.8 ± 0.8 mg/dl vs. 10.5 ± 1.0 mg/dl, p= 0.002 in the control group). Although the mean Hb change in the PTX group was more, it was not statistically significant (1.0 ± 0.9 vs. 0.7 ± 1.1, p=0.195).

EPO doses decreased following treatment in the PTX group (11000.0 ± 3140.0 IU vs. 9100.0±3400.0 IU, p=0.018), while rhEPO dose was not significantly changed in the control group (11130.0 ± 3180.0 IU vs. 10870.0 ± 3900.0 IU, p=0.690).

CRP levels, as an inflammatory index, significantly decreased in the PTX group (22.8 ± 15.3 vs. 16.5 ± 9.5, p=0.005) at the end of the study. However, there was no significant difference in the CRP levels in the control group (p=0.080).

Although the mean of ESR level demonstrated a decrescendo pattern in both groups, there was no significant difference (p>0.05). Serum Alb levels, as a nutritional factor, increased significantly after treatment in the PTX group (3.9 ± 0.4 vs. 4.1 ± 0.3, p=0.031) in contrast to lack of significant changes in the control group (p=0.062).

This study has provided some evidence to help the nephrologists whether PTX administration will effectively reduce the required dose of rhEPO in HD patients, possibly through decreasing the inflammation. Although the advent of rhEPO improves the treatment of anemia among CKD patients, higher rhEPO doses have been associated with adverse outcomes. As inflammation and increase in cytokines are contributing factors in the development of anemia in CKD patients, the selection of PTX as an adjuvant therapy with EPO may be reasonable, because it inhibits the monocyte production of tumor necrosis (TNF)-α and T cell production of interferon-gamma (IFNγ) [4,5].

The PTX effect on anemia in HD patients

The result of this study showed that the Hb level increased significantly after treatment in both groups. Although PTX consumption led to more Hb elevation in the PTX group, it was not significant. On the other hand, PTX consumption significantly decreased the rhEPO dose in the PTX group more than the control group. These results are in contrast with those of the studies reported by some investigators, showing no significant effect of PTX on Hb concentration and/or rhEPO dose in their clinical trials [9,10]. In addition, this study showed that the erythropoiesis-stimulating agent’s resistance index at the end of the 4-month study period were 15% lower in the PTX group compared to control group, although it was not statistically significant. Nonetheless, PTX might have had significant effects on the increase of Hb levels and/or decrease of rhEPO demand. Also, some of the studies recommended that PTX could be considered as a therapeutic method in this issue [6,7,9,11].

Ferrari et al. [7] demonstrated that PTX reduces the circulating interleukin (IL)-6, increases the transferrin saturation, decreases the serum ferritin and improves the HB in moderate to severe CKD. These changes suggest improving iron release, and it is hypothesized that PTX improves iron disposition possibly through modulation of hepcidin. However, PTX administration had no effect on the serum hepcidin levels in a clinical trial of HB patients exactly designed for investigating such possible effect [11].

Chronic inflammation is considered as the main factor of morbidity and mortality in HD patients that has arisen from the immune system response to uremia. It can cause immunosuppression and chronic activation of the immune system condition [12-17] Which may generate or worsen anemia by limitation of the pro-erythropoietic EPO effect through TNF-α and pro-inflammatory cytokines. TNF-α inhibits erythropoiesis in vitro [16] through an indirect mechanism requiring IFN-gamma [17]. In rheumatoid arthritis patients with anemia of chronic disease, administration of an anti–TNF-α antibody provided a significant rise in hemoglobin levels [18]. Anti–TNF therapy should be administered parenterally and has been associated with severe adverse consequences. PTX is administered orally and has a low incidence of serious side effects which makes it a more reasonable adjuvant therapy for treatment of renal anemia compared to anti-TNF antibody [19].

During the current study, 3 patients of the PTX group were excluded due to adverse events. The most common side effects of the drug are gastrointestinal ones as nausea and vomiting [20] Advanced research in this area is warranted.

The PTX effect on CRP and ESR in HD patients

It is established that inflammatory factors, such as CRP rise in ESRD patients [21]. In the current study, all parameters related to Hb procedure, like dialysis solution, type of membrane, vascular access, and KT/V which affect inflammation [22] were the same in both groups. However, the patients were evaluated throughout the study for signs and symptoms of infection to eliminate the confounding factors in evaluation of CRP. According to our results, PTX could significantly decrease the CRP level compared to the control group which was the same as previous studies in HD patients [9,15].

PTX inhibits phosphodiesterase, causing an increase in intracellular cyclic adenosine monophosphate activity and down-regulation of pro-inflammatory cytokine synthesis. This drug inhibits the synthesis of the inflammation markers at a transcriptional level. In addition, PTX prevents proliferation of the peripheral blood mononuclear cell, adherence to the cell matrix and endothelium and T and natural killer cell cytotoxicity [23-25].

The PTX effect on Alb as a nutritional index in HD patients

Serum Alb levels increased significantly after treatment in the PTX group compared to the control group like other studies. Malnutrition followed by decreased serum Alb and inflammation, which is often observed in HD, which can cause the activation of oxidative stress and inflammatory response and also cause renal anemia [1,12,15,16]. PTX increase the serum Alb levels by its anti-inflammatory effect. Also, the correlation of low serum Alb with mortality in CKD patients is partly linked to its association with systemic inflammation [13,14].

Enhancement of proinflammatory cytokines such as TNF-α and IL-6 in HD patients may suppress the appetite and induce catabolism, [26,27] contributing to a wasting illness. Bologa et al. [28] found a significant correlation between plasma TNF-α and IL-6, and the degree of hypoalbuminemia and dyslipoproteinemia in this population.

Vaidyanathapuram et al. [29] indicated that single nucleotide polymorphisms in the promoter region of the proinflammatory cytokines IL-6, TNF-α and the regulatory monokine IL-10 are strongly associated with indices of comorbidity and function, as well as serum albumin in ESRD patients on long-term HD. Several immune function mechanisms are involved in this condition, including activation of oxidative stress, inflammatory response and the dialysis measure itself. As a result, PTX by prohibiting inflammatory markers may lead to improvement in the serum albumin and nutritional state of patients.

Study limitations

The most important limitations of this study were short research duration and single center accomplishment. Therefore, larger and longer studies are recommended.

According to the results of the present study and other researches in this field, it has been seen that the use of PTX causes a reduction in the required dose of EPO and could be helpful in treatment of renal anemia. However, it is controversial as a therapeutic strategy in CKD anemia. In addition, CRP levels as an inflammatory index were significantly decreased in the PTX group. We recommend that more trials with larger sample size should be done to clarify the PTX role in improvement of CKD anemia.

The authors would also like to thank Shiraz University of Medical Sciences, Shiraz, Iran and also Center for Development of Clinical Research of Nemazee Hospital and Dr. Nasrin Shokrpour for editorial assistance.

The authors report no conflict of interest.

This article is based on a thesis written by Pegah Aghajanzade and supported financially by Shiraz University of Medical Sciences with Grant No 49-7064.

MP: Contributed to design and analysis, drafted the manuscript, finally approved it, revised the manuscript, and accepts accountability for the overall work. PA and MR: Contributed to design and analysis, revised the manuscript, finally approved it, and accepts accountability for the overall work. LM Contributed to data collection, revised the manuscript, finally approved it, and accepts accountability for the overall work.