Chronic kidney disease (CKD) is an increasing and global health problem with a great economic burden for healthcare system. Therefore to slow down the progression of this condition is a main objective in nephrology. It has been extensively reported that microinflammation, immune system deregulation, and oxidative stress contribute to CKD progression. Additionally, dialysis worsens this clinical condition because of the contact of blood with bioincompatible dialytic devices. Numerous studies have shown the close link between immune system impairment and CKD but most have been performed using classical biomolecular strategies. These methodologies are limited in their ability to discover new elements and enable measuring the simultaneous influence of multiple factors. The “omics” techniques could overcome these gaps. For example, transcriptomics has revealed that mitochondria and inflammasome have a role in pathogenesis of CKD and are pivotal elements in the cellular alterations leading to systemic complications. We believe that a larger employment of this technique, together with other “omics” methodologies, could help clinicians to obtain new pathogenetic insights, novel diagnostic biomarkers, and therapeutic targets. Finally, transcriptomics could allow clinicians to personalize therapeutic strategies according to individual genetic background (nutrigenomic and pharmacogenomic). In this review, we analyzed the available transcriptomic studies involving CKD patients.
Chronic kidney disease (CKD) represents an increasing global worldwide health problem particularly in elderly people [
During this condition, patients experience a gradual loss of renal function over time with a progressive decline in the glomerular filtration rate (GFR). An international consensus categorized CKD into 5 stages according to the GFR [
As the kidney is a complex and highly specialized organ, with different functions (e.g., pH, plasma and tissue hydrosaline balance, and vitamin D and erythropoietin production), chronic kidney impairment may also determine significant metabolic and endocrine changes (including acidosis, hyperparathyroidism, and anemia) that may induce relevant clinical complications (e.g., atherosclerosis, pericarditis, osteodystrophy, and uremic encephalopathy) [
Consequently, a healthy life style, with an adequate physical activity and an equilibrate diet, with low salt intake and no smoking habit, can undeniably prevent or slow down the progression of renal damage and lessen complications [
Additionally, the removal of the oxidative stress and microinflammatory insults represents an additional powerful therapeutic target in CKD [
Several factors may be responsible of the chronic immune-inflammatory state and oxidative stress in CKD patients (Figure
Schematic representation of the main factors involved in microinflammation and oxidative stress in chronic kidney disease (CKD). As reported, (1) bioincompatible dialysis devices and plastificants; (2) classical inflammatory cytokines and new emerging biological elements such as pentraxin-3 (PTX3), TNF-like weak inducer of apoptosis (TWEAK), and adipokines; (3) uremia-induced intestinal dysbiosis with an increased translocation of gut bacteria and bacterial components into the circulation; and (4) mitochondrial deregulation may have a central role in the onset of chronic microinflammatory state and oxidative stress and development of malnutrition, inflammation, and atherosclerosis (MIA) syndrome, systemic complications, immune system deregulation, cardiovascular complications, and other systemic comorbidities in CKD patients.
PTX3, a circulating acute phase protein with pattern recognition molecule properties and with antibody-like functions, contributes to innate immunity defence against pathogens and in the regulation of inflammation in CKD [
TWEAK can increase secretion of other cytokines locally in the kidney and its blood levels seem independently associated with coronary artery disease in patients with renal damage [
Adipokines, together with visfatin and leptin, were found higher in CKD and in nondiabetic peritoneal dialysis patients. Interestingly, leptin/adiponectin ratio was able to predict mortality in a group of nondiabetic uremic patients undergoing peritoneal dialysis treatment [
Interestingly, convincing recent evidences suggest that uremia-induced intestinal dysbiosis may have a central role in these processes by increasing the translocation of gut bacteria and bacterial components into the circulation, which can in turn activate systemic inflammation [
A recent cross-sectional study in stage 3-4 CKD demonstrated that indoxyl sulfate and p-cresyl sulfate (nephro- and cardiovascular toxins produced solely by the gut microbiota) were associated with elevated levels of inflammatory biomarkers as well as with increased arterial stiffness [
Additionally, it has been largely reported that CKD patients develop a complex immune dysfunction with an interaction between the innate and adaptive systems, in which immune activation (hypercytokinemia and acute phase response) and immune suppression (impairment of response to infections and poor development of adaptive immunity) coexist [
CKD progression and dialysis procedure contribute also to protein-energy malnutrition, probably mediated by proinflammatory cytokines that can affect appetite and increased protein hydrolysis and muscle protein breakdown [
To minimize these dialysis-related conditions [
Oxidative stress and mitochondrial deregulation play a major role in CKD and, already from the early stage of CKD, several markers of oxidative stress (e.g., malondialdehyde, F2 isoprostanes, and advanced oxidation protein products) are plentiful with a concomitant decrease of antioxidants (e.g., superoxide dismutase, glutathione peroxidase, and vitamins E and C) [
The accumulation of uremic toxins, through the direct augmentation of NADPH oxidase and xanthine oxidoreductase activities [
Likewise, during PD conventional dialysis solutions, containing high concentrations of glucose and glucose degradation products, may increase ROS production in human peritoneal mesothelial cells with consequent loss of ultrafiltration capacity, increased vascular density, and development of fibrosis [
Oxidative stress is also accountable for the onset and development of severe clinical complications (including cardiovascular disease, atherosclerosis, hypertension, anemia, and malnutrition) with a consequent low quality of life, high risk of hospitalization, and short survival of CKD patients in both conservative and dialysis treatment [
Notably, recent studies have suggested that mitochondria could be implicated in this CKD-associated prooxidative machinery [
Structurally, they present an outer and inner membrane, the latter of which would be impermeable to all molecules in the absence of specific carriers and contains the OXPHOS complexes. Electrons derived from metabolic reducing equivalents (NADH and FADH2) enter into the electron transport chain through either complex I or complex II and via respiratory chain to molecular oxygen which is finally reduced to water. This exergonic process is used to pump protons from the mitochondrial matrix into the intermembrane space, creating an electrochemical gradient used by complex V for ATP synthesis [
During this process a small percentage (0.4–4%) of electrons may “leak” from the respiratory chain (in particular at complexes I and III) and partially reduce oxygen, forming superoxide anion (
Additionally, during CKD, patients’ cells undergo reduction in mitochondrial DNA (mtDNA) copy number, loss of mitochondrial membrane potential
Mitochondria are also involved in apoptosis and epithelial to mesenchymal transition of renal tubular epithelial cells contributing to the fibrogenic process [
In the last two decades, researchers have tried to identify key regulators of the intricate inflammatory pathway activated by uremia “per se” and by dialysis, but, most of the time, this research strategy based on single factor analysis is limited and biased. Hence, a simultaneous multifactorial analysis in CKD appears more powerful and effective.
The recent development and extension of high-throughput technologies have allowed reaching the above-mentioned objective. In particular, microarray technology, which allows the study of the entire transcriptome thanks to the hybridization of nucleic acid (RNA) with dozens of thousands of DNA probes attached to a solid support (such as glass, plastic or silicon), has been revealed to be promising. Briefly, transcripts extracted from samples are labeled with fluorescent dyes and hybridize to their complementary targets. Light intensity is then an indirect measurement of gene expression. Transcriptome is the sum of RNA transcripts that comprehend messenger RNAs, ribosomal and transfer RNA, and regulatory noncoding RNAs [
This technology produces a large amount of raw data that require specific statistical and bioinformatics tools in order to avoid or minimize false positive and to obtain “more conservative” results. In this context, a well conducted validation process by using standardized classical biomolecular methodologies can reduce these biases.
As the other high-throughput (omics) sciences, no prior hypothesis is made. Relationships among top selected genes are, then, translated into biological pathway by using specific software for functional analysis (e.g., Ingenuity Pathway Analysis) [
In the last ten years numerous studies have used this approach in nephrology and the results have been very useful in discovering new insights in the pathogenesis of CKD as well as in the comorbidities associated with renal failure (Table
Relevant studies using transcriptomics in nephrology.
Reference | Comparison | Tissue/cells | Selected genes |
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[ |
HD versus PD versus CKD | PBMC |
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ATOX1, RELA, CSDE1, MIF, LTB4R, GSS, NFRKB | |||
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HRH1, OLR1, CHST4, S100A8, CXCL12, GPX7 | |||
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IL8RB, HDAC5, BCL6 P | |||
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[ |
PRE-HD versus POST-HD | Blood |
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TNF-A, IL-8, IL-18, IL-1RN, IL-4R, IL-10R, IFN- | |||
HD high CRP versus HD low CRP |
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IL-1RN, IL-4R, IL-10R, IFN- | |||
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[ |
HD versus HS | Muscle |
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SP3, MEF2A, MAF, TCF8, SMARCA1, DICER1, SFRS11, HMGN3, UPF3A, EPM2A, SOS2, DEK, CLK1, CDC10, LAF4, BMI1, DDX17, MAPK6, ANAPC13, MYBPC1, C6orf111, KIAA0740, ART3, BIRC2, RABGGTB, OA48-18, CSE1L, SH3GLB1, MAP2K4, GLRX, PIP5K3, SLC35A1, VPS26, PXMP1, SRP54, SCP-2, SUCLA2, DMD, PRDX3, NDUFA5, NRIP1, XPO1, PSMC6, SEPP1, AXOT, LANCL1, SHOC2, FAM8A1, UBE1C, UBL3, PJA2, YME1L1, ELF2, OGT, IRS1, GATM, DLD, BZRP, PICALM, CAST, ANGPT1, ANK3, AKAP9, Rif1, CBX3, CBX1, ZNF146, MYH8, Tl132, MORC3, ZC3H11A, PURA, FLJ13110, GBAS, KTN1, SLC30A9, Tre | |||
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PTK9L, IGFBP4, TRAP1, TAX1BP3, LGALS3BP, GNAI2, HBA1, HBB | |||
PRE-HD versus POST-HD |
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FST, GADD45A, GADD45B, IGFBP4, SAT, C-FOS, JUN-B, THBD, HES1, CCL2, CEBPD, BTG2, FOSL2, MYC, THBD, ZFP36, JE, NFIL3, SERPINB1, SCL39A14, NNMT, ARID5B | |||
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TOB1 | |||
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[ |
semisynthetic versus full-synthetic dialysis membrane | PBMC |
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[ |
PMN stimulated with shredded hollow fibres of CU or PS versus unstimulated PMN | PMN |
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AXUD1, FTH1, LIF, PTGS2, MGC12815, IL-1b, CCL3, CXCL1, SOCS3, PPIF, SPAG9, ACPP, DCT, GLA, GNS, PFKFB3, PLAU, USP36, SFRS3, DDX48, FLJ23231, PTD004, GNA13, HBEGF, DPYSL3, ARL8, GPR4, RASL11, DUSP2, EDN1, EDN3, EDNRB, JUN, FOS, EGR1, EGR2, DDIT3, EGR3, ELL2, NR4A3, TFAP2A, STAR, SEC31L1, ATP13A3, PHACTR1, TncRNA | |||
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FADD, FLI1, SOLH, YPEL3 | |||
PMN stimulated with |
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GADD45B, BIRC3, IER3, IER5, SGK, CDH24, ICAM1, CSF1, VEGF, NBS1, CCL18, CCL20, CCL3, CD48, CXCL1, CXCL2, CXCL3, IL1A, IL1B, IL1RN, LIF, MGC12815, NFIL3, PTGS2, SOCS3, TNF, TNFAIP6, EGR1, EGR2, ETS2, HIVEP1, ISL1, JUN, MAFF, MAFG, NFKB1, NFKBIA, NFKBIE, NFKBIZ, NR4A3, TFAP2A, TNFAIP3, XBP1, ZFHX1B, B4GALT5, DCT, FPGS, GCH1, GLA, GNPDA1, LOC285533, OAZIN, PLAU, PPIF, PPP1R15B, DDX48, FLJ23231, NMES1, SFMBT2, SNAPC3, TIFA, PHACTR1, ARL8, CALCA, CDC42EP3, DPYSL3, DUSP2, EDN1, EDN3, EHD1, GAB2, GPR4, MAPK6, NSMAF, SLC35B2, RHCG, SPAG9, VANGL1, VPS18, KCNJ2, AQP9 | |||
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FADD, LAMB1, MEF2C, HNRPUL1, NDP52, YPEL3, DUSP6 | |||
PMN stimulated with |
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CCL20, CXCL3, CCL3, IL1A, TNF, NFKBIA, NFKBIE, NFKBIZ, NFKB1, TNFAIP3, PLAU, IER5, ICAM3 | |||
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[ |
HD versus CKD III-IV | PBMC |
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[ |
HD versus HS | Blood |
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[ |
Mice with intraperitoneal injection of chlorhexidine gluconate versus control | Parietal peritoneum |
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[ |
PD/HD versus CKD III-IV/HS | PBMC |
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[ |
HD versus CKD II-III versus HS | PBMC |
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[ |
HD versus HS | PBMC |
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Our group, in the 2008, has published one of the first studies using this technology in dialysis [
CXCL12 and its receptor, CXCR4, are important modulators of inflammation and immune response. IL8RB binds IL-8, a chemokine with proinflammatory and chemotactic activity [
These data were in line with those obtained by Friedrich et al. [
Similar results were obtained by Shah et al. [
Transcriptomic profile was also influenced by different dialysis procedures. Wilflingseder et al. clearly demonstrated with microarray analysis conducted in PBMCs of four stable HD patients that a large group of genes (
Surprisingly, these results were different from those published by Hochegger et al. [
Subsequently, our research group performed a research project aimed at understanding the influence of dialysis on the PBMCs’ immunotranscriptome [
Among the genes downregulated in HD patients, we identified those encoding the human leukocyte antigen- (HLA-) G, a nonclassical major histocompatibility complex class I molecule that differs from other HLA class I molecules with regard to its low polymorphism, restricted tissue distribution, slow turnover, immunosuppressive properties, and limited peptide diversity [
Under physiological conditions, the production of HLA-G protein is restricted to trophoblast [
HLA-G possesses the capability to bind inhibitory receptors such as the immunoglobulin-like transcripts 2 and 4 (ILT2, ILT4) and the killer immunoglobulin-like receptor (KIR)2DL4/CD158d with inhibitory effects [
HLA-G may also have a direct immune-inhibitory function through blocking effector cells and indirect immune-inhibitory activity by regulatory cell generation. Via the direct inhibitory functions, HLA-G is able to inhibit the cytolytic activity and proliferation of NK [
Therefore, it is plausible that the lower HLA-G expression in HD patients may determine a hyperactivation of T cells and NK that could explain the different immune response of dialyzed patients to viral infections and tumors.
Recently, Scherer et al. [
Moreover, Yokoi et al. [
This growth factor was found not only in fibroblasts and mesothelial cells within the underlying submesothelial compact zones of mice, but also in human peritoneal biopsy samples and peritoneal dialysate effluent. In wild-type mice, CG treatment increased peritoneal permeability, increased mRNA level of TGF-
Also genes involved in proteoglycans biosynthesis/metabolism appear differentially expressed in dialyzed patients [
These results demonstrated that PBMCs of uremic patients undergoing both peritoneal and hemodialysis exhibit a chronic activation of the biosynthetic proteoglycans transcriptomic pattern with heparanase being a central biological element. This enzyme could be considered important for rolling and leukocytes mobility in response of pathological dialysis stimuli. In future, a pharmacological modulation of HPSE could definitely mitigate these effects and reduce the frequent CKD-associated vascular comorbidities.
Transcriptomic analysis demonstrated different expression of several genes involved in oxidative phosphorylation system (OXPHOS) and mitochondrial function in ESRD/HD patients compared to healthy subjects [
Mitochondria are major source of cellular ATP molecules, but if damaged, they may generate high levels of ROS with massive clinical systemic consequences. Therefore, modulating their function and biogenesis could turn to be a valuable therapeutic option.
Finally, a combined research strategy between classical biomolecular strategies and high-throughput techniques showed the Nod-like receptor protein 3 (NLRP3) inflammasome activation in dialyzed CKD patients [
NLRP3 inflammasome can be activated by a lot of exogenous and endogenous stimuli: pathogen-associated molecular patterns (PAMPs), such as bacterial and viral RNA [
The activation of NLRP3 inflammasome requires 2 specific signals. The first, or priming signal, converges on NF-
ROS being able to activate the proinflammatory transcription factors [
The central role of mitochondrial ROS and NLRP3 activation has been reported also in the pathogenesis of albumin-induced renal tubular injury [
A correct analysis of transcriptomic/microarray results may be useful to identify valuable biomarkers and to uncover new therapeutic targets for CKD. This strategy could also facilitate the employment of new available molecules/drugs in nephrology (Figure
Site of action of most common endogenous and food derived antioxidants, phytochemicals, and conventional drugs with favorable antioxidant side effects and new available more selective anti-inflammatory medications. Some food derived antioxidants and drug (captopril) have both direct antioxidant effect acting as a scavenger of free radicals or inhibiting lipid peroxidation and indirect effect by modulating the activity of transcription factors NF-
Mainly, endogenous and food derived antioxidants, phytochemicals, conventional drugs with favorable antioxidant side effects, and mitochondria-targeted molecules seem promising tools [
Among endogenous and food derived antioxidants L-carnitine, coenzyme Q10 (CoQ10), alpha-lipoic acid (ALA), omega 3 polyunsaturated fatty acids (Omega-3 PUFAs), and vitamins E and vitamin C have demonstrated direct and indirect antioxidant actions in clinical studies conducted in CKD and HD patients [
Several phytochemicals such as thymoquinone (from
Also conventional and routinely used drugs may have favorable antioxidant side effects. For example, captopril has a thiol group in its structure that scavenges ROS and increases antioxidants enzyme levels.
Unfortunately, most of these molecules are unable to reach mitochondria at therapeutic dosage. For this reason “shuttle” molecules to better deliver antioxidants into the mitochondria are a growing field of interest.
First synthesized mitochondria target molecule was MitoE that is vitamin E conjugated with a lipophilic molecule, triphenylphosphonium (TPP). Then, with the same technique, MitoQ (a quinone linked to TPP), MitoSOD (a mimetic of MnSOD linked to TPP), and Mito-TEMPO (a nitroxide linked to TPP) were also produced. Phase I and II clinical trials are ongoing for MitoQ [
More recently even more efficacious peptides have been introduced: Szeto-Schiller peptides (SS) and mitochondrial cell-penetrating peptides (mt-CPPs) [
Moreover, pharmaceutical companies are initiating new research programs to discover and develop potent, selective anti-inflammatory medications. Among them, bardoxolone methyl (activators of Nrf2) has been the first to reach full clinical development. It induces the antioxidant and cytoprotective transcription factor Nrf2, reduces the proinflammatory activity of the IKK-
Another interesting compound is the primary amine of ethanolamine [
PEA was first discovered in the late 1950s by studying the antiallergic and anti-inflammatory activity exerted by dietary supplementation with egg yolk, peanut oil, or soybean lecithin [
Anti-inflammatory and protective activities of PEA were confirmed in several models of inflammation, that is, carrageenan-induced paw edema, adjuvant-induced arthritis, tuberculin hypersensitivity, and ischemia reperfusion injury [
Finally, also peroxisome proliferator-activated receptors (PPARs), nuclear hormone receptors that stimulate transcription of genes by binding to specific DNA sequences, have demonstrated beneficial effects on vascular function [
Moreover PPAR-
Moreover, the renoprotective effect of exogenous PPAR-
Therefore, to obtain the best results with these drugs it could be important to personalize administration and to properly identify the “right patient for the right medication.” Transcriptomic strategy could help to reach this objective.
Transcriptomic analysis, although still not largely employed in nephrology, has demonstrated great speculative potentialities. Several key regulators of the immune-inflammatory and oxidative stress pathways in CKD have been identified by using this innovative technology.
Additionally, it has demonstrated a unique capability to help clinicians to personalize patients’ treatment according to their multigenetic expression fingerprint. A “customized” drug and diet administration (nutrigenomic and pharmacogenomic), by permitting the introduction of innovative therapeutic protocols including new antioxidant and anti-inflammatory compounds (e.g., endogenous and food derived antioxidants, phytochemicals, and mitochondria-targeted molecules), could definitely have a remarkable clinical and therapeutic impact.
Recently, M. R. Shahidi Bonjar and L. Shahidi Bonjar [
However, to enforce the routine use of this methodology, researchers should work more to make data analysis and interpretation easily accessible to medical doctor not expert in statistics and bioinformatics and to reduce cost-consuming of microarray experiments.
For these reasons, at the moment, we are still far from large employment of this methodology in the nephrology research and in daily clinical practice. To achieve this objective a multidisciplinary network (including medical doctors, biologists, and statisticians) should be developed.
Chronic kidney disease
Glomerular filtration rate
End stage renal disease
Renal replacement therapies
Hemodialysis
Peritoneal dialysis
Angiotensin-converting enzyme
Angiotensin receptor blockers
Peripheral blood mononuclear cells
Tumor necrosis factor-
Pentraxin-3
TNF-like weak inducer of apoptosis
Reactive oxygen species
Oxidative phosphorylation system
Nicotinamide adenine dinucleotide
Flavin adenine dinucleotide
Mitochondrial DNA
Superoxide anion
C-reactive protein
High-sensitive C reactive protein
Polymorphonuclear neutrophils
Human leukocyte antigen
Natural killer
Chlorhexidine gluconate
Healthy subjects
Heparanase
Pathogen-associated molecular patterns
Damage-associated molecular patterns
Alpha-lipoic acid
Coenzyme Q10
Omega 3 polyunsaturated fatty acids
Triphenylphosphonium
Szeto-Schiller
Mitochondrial cell-penetrating peptides
Nuclear factor erythroid 2-related factor 2
Primary amine ethanolamine
N-Acylethanolamine
Peroxisome proliferator-activated receptors
Bovine serum albumin
Quantitative microarray detector
Homeostasis-oriented microarray column.
The authors declare that there is no conflict of interests regarding the publication of this paper.