Cardiac antiapoptotic and proproliferative effect of recombinant human erythropoietin in a moderate stage of chronic renal failure in the rat

Abstract

Objective:Recombinant human erythropoietin (rhEPO) therapy under circumstances of moderate chronic renal failure (CRF), with yet lower kidney and heart lesion, may have a protective cardiac effect beyond the correction of anemia, whose mechanism deserves better elucidation, namely by clarifying the impact on gene expression profile of markers of apoptosis, inflammation, proliferation, angiogenesis, and lesion/stress in the heart.Materials and Methods:Four groups of rats were studied over a period of 15 weeks (n=7 each): control—without surgery and without drug treatment; rhEPO—treated with 50 IU/kg/week of rhEPO—beta; CRF—submitted to partial nephrectomy (3/4); CRF + rhEPO—CRF with rhEPO treatment after the 3rd week of surgery. The heart was collected in order to evaluate the gene expression, by real-time qPCR, of markers of apoptotic machinery, inflammation/immunology, proliferation/angiogenesis, and lesion/stress.Results:The main findings obtained were (a) CRF rats have demonstrated overexpression of EPO-R in the heart without changes on EPO expression, together with overexpression of Bax/Bcl2 ratio, PCNA, and IL-2; (b) rhEPO therapy on the heart of the rats with CRF induced by partial 3/4 nephrectomy promoted nonhematopoietic protection, demonstrated by the apoptosis prevention, viewed by the Bax/Bcl2 balance, by the promotion of proliferation, due to PCNA increment, and by the immunomodulatory action, expressed by a trend to prevent the IL-2 increment.Conclusion:In this model of moderate CRF, rhEPO treatment showed important cardiac nonhematopoietic effects, expressed mainly by the antiapoptotic and the proproliferative action, suggesting that early rhEPO therapy in moderate stages of CRF might have further therapeutic benefits.

Early cardiac changes in a rat model of prediabetes: brain natriuretic peptide overexpression seems to be the best marker

“…The yield from isolation was from 0.5 to 3 μg; RIN values were 6.0–9.0 and purity (A260/A280) was 1.8–2.0. Reverse transcription and relative quantification of gene expression were performed as previously described [25]. Real-time PCR reactions were performed using the following primer sequences for Bax, Bcl2, brain natriuretic peptide (BNP), connective tissue growth factor (CTGF), intercellular adhesion molecule 1 (ICAM-1), IL-6, inducible nitric oxide synthase (iNOS), Pro-collagen type III, superoxide dismutase (SOD), TGF-β1, TNF-α, thrombospondin 1 (TSP-1), tribbles 3 (TRB3), vascular cell adhesion molecule 1 (VCAM-1) and VEGF, which were normalized in relation to the expression of beta-actin as an internal control: Bax Forward: 5 ′ -CCAAGAAGC TGAGCGAGTGTCTC-3 ′ and Bax Reverse: 5 ′ -AGTTGC CATCAGCAAACATGTCA-3 ′ ; Bcl-2 Forward: 5 ′ -GGAG CGTCAACAGGGAGATG-3 ′ and Bcl-2 Reverse: 5 ′ -GA TGCCGGTTCAGGTACTCAG-3 ′ ; BNP Forward: 5 ′ -G GGCTGTGACGGGCTGAGGTT-3 ′ and BNP Reverse: 5 ′ -AGTTTGTGCTGGAAGTAAGA-3 ′ ; CTGF Forward: 5 ′ -CGTAGACGGTAAAGCAATGG-3 ′ and CTGF Reverse: 5 ′ -AGTCAAAGAAGCAGCAAACAC-3 ′ ; ICAM-1 Forward: 5 ′ -TTCAACCCGTGCCAGGC-3 ′ and ICAM-1 Reverse: 5 ′ -GTTCGTCTTTCATCCAGTTAGTCT-3 ′ ; IL-6 Forward: 5 ′ -ACCACTTCACAAGTCGGAGG-3 ′ and IL-6 Reverse: 5 ′ -ACAGTGCATCATCGCTGTTC-3 ′ ; iNOS Forward: 5 ′ -CAGAAGCAGAATGTGACCATCAT-3 ′ and iNOS Reverse: 5 ′ -CGGAGGGACCAGCCAAATC-3 ′ ; Pro-collagen type III Forward: 5 ′ -5 ′ -GGTCACTTTCACT GGTTGACGA-3 ′ and Pro-collagen type III Reverse: 5 ′ -TTGAATATCAAACACGCAAGGC-3 ′ ; SOD Forward: 5 ′ -GACAAACCTGAGCCCTAACGG-3 ′ and SOD Reverse: 5 ′ -CTTCTTGCAAACTATG-3 ′ ; TGF-β1 Forward: 5 ′ -ATACGCCTGAGTGGCTGTCT-3 ′ and TGF-β1 Reverse: 5 ′ -TGGGACTGATCCCATTGATT-3 ′ ; TNF-α Forward: 5 ′ -CACGCTTTCTGTCTACTGA-3 ′ and TNF-α Reverse 5 ′ -GGACTCCGTGATGTCTAAGT-3 ′ ; TSP-1 Forward: 5 ′ -CTTTGCTGGTGCCAAGTGTA-3 ′ and TSP-1 Reverse: 5 ′ -CGACGTCTTTGCACTGGATA-3 ′ ; TRB3 Forward: 5 ′ -TGATGCTGTCTGGATGACAA-3 ′ and TRB3 Reverse: 5 ′ -GTGAATGGGGACTTTGGTCT-3 ′ ; VCAM-1 Forward: 5 ′ -GAAGCCGGTCATGGTCAAGT-3 ′ and VCAM-1 Reverse: 5 ′ -GACGGTCACCCTTGAACAG TTC-3 ′ ; VEGF Forward: 5 ′ -GAGAATTCGGCCCCAA CCATGAACTTTCTGCT-3 ′ and VEGF Reverse: 5 ′ -G AGCATGCCCTCCTGCCCGGCTCACCGC-3 ′ ; beta-actin Forward: 5 ′ -TGTGCTATGTTGCCCTAGACTT C-3 ′ and beta-actin Reverse: 5 ′ -CGGACTCATCGTA CTCCTGCT-3 ′ .…”

Effect of SDF-1/CXCR4 axis on the migration of transplanted bone mesenchymal stem cells mobilized by erythropoietin toward lesion sites following spinal cord injury

“…Erythropoietin (EPO) is a glycoprotein that mediates the differentiation and apoptosis of a number of non-hematopoietic cells via the erythropoietin receptor (EPOR) ( 20 ). There is increasing evidence for the different roles of EPO in various types of diseases, such as calvarial defects ( 21 ), chronic renal failure ( 22 ) myocardial infarction ( 23 ), osteonecrosis of the femoral head ( 24 ), spinal cord injury ( 25 , 26 ), diabetes mellitus ( 27 ) and acute lung injury ( 28 , 29 ). The study by Zwezdaryk et al ( 30 ) confirmed that EPOR is also expressed on the surface of BMSCs.…”

Iron-Hepcidin Dysmetabolism, Anemia and Renal Hypoxia, Inflammation and Fibrosis in the Remnant Kidney Rat Model

“…Possible contaminating remnants of genomic DNA were eliminated by treating these preparations with deoxyribonuclease I (amplification grade) prior to RT-qPCR amplification. Reverse transcription and relative quantification of gene expression were performed as previously described [ 24 ]. Real-time qPCR reactions were performed for the following genes: EPO , EPOR , Transferrin receptor 2 ( TfR2 ), Hepcidin ( Hamp ), Ferroportin ( SLC40A1 ), Hemojuvelin ( HJV ), Transferrin ( TF ), Hemochromatosis ( Hfe ), +IRE-Divalent Metal Transporter 1 ( DMT1 ), Transferrin recptor 1 ( TfR1 ), Matriptase-2 ( TMPRSS6 ), Interleukin-6 ( IL-6 ) and Bone Morphogenic Protein 6 ( BMP6 ); vascular endothelial growth factor (VEGF) , interleukin-1 beta (IL-1β) , nuclear transcription factor kappa B ( NF-kB) , connective tissue growth factor ( CTGF) and tumor necrosis factor alpha (TNF-α) which were normalized in relation to the expression of beta-actin ( Actb ), and 18S ribosomal subunit ( 18S ).…”