Effects of high amylose resistant starch on gut microbiota and uremic toxin levels in patients with stage G3a-G4 chronic kidney disease: a randomized trial.

OBJECTIVE: This study was designed to determine the effect of 16 weeks of supplementation with Hi-maize 260 resistant starch on the gut microbiota, uremic toxins (indoxyl sulfate and p-cresyl sulfate), markers of inflammation and oxidative stress along with vascular function in patients with stage G3a-G4 chronic kidney disease (CKD). DESIGN: & Methods: This was a double-blind, placebo-controlled, parallel-arm, randomized controlled trial. Sixty-eight patients with stage G3a-G4 CKD were randomized to either resistant starch with usual care or placebo and usual care. Patients attended four testing sessions: two baseline visits, and follow-up visits at 8 and 16 weeks. Fasting blood samples, resting brachial and central blood pressures, along with arterial stiffness, were collected at visits (1 or 2), and weeks 8 and 16. A stool sample was collected for analysis of microbial composition at baseline and week 16. Patients were randomized after the baseline visits. RESULTS: Patients receiving the resistant starch had a reduction in p-cresyl sulfate at week 16. This reduction was associated with a decrease in microbial α-diversity between baseline and week 16 (Chao1 p=0.014, Shannon p=0.017, PD p= 0.046, and Simpson p=0.017) as well as increases in Subdoligranulum (p=0.03) and Oscillospiraceae UCG 002 (p=0.02) and decreases in Bacteroides (p=0.009).There were no changes in microbial beta diversity and other biomarkers or markers of vascular function following the 16-week period Conclusion: Sixteen weeks of supplementation of resistant starch in patients with stage G3a-G4 CKD led to changes in microbial composition that were associated with a significant reduction in p-cresyl sulfate.

Targeting Lactobacillus johnsonii to reverse chronic kidney disease.

Accumulated evidence suggested that gut microbial dysbiosis interplayed with progressive chronic kidney disease (CKD). However, no available therapy is effective in suppressing progressive CKD. Here, using microbiomics in 480 participants including healthy controls and patients with stage 1-5 CKD, we identified an elongation taxonomic chain Bacilli-Lactobacillales-Lactobacillaceae-Lactobacillus-Lactobacillus johnsonii correlated with patients with CKD progression, whose abundance strongly correlated with clinical kidney markers. L. johnsonii abundance reduced with progressive CKD in rats with adenine-induced CKD. L. johnsonii supplementation ameliorated kidney lesion. Serum indole-3-aldehyde (IAld), whose level strongly negatively correlated with creatinine level in CKD rats, decreased in serum of rats induced using unilateral ureteral obstruction (UUO) and 5/6 nephrectomy (NX) as well as late CKD patients. Treatment with IAld dampened kidney lesion through suppressing aryl hydrocarbon receptor (AHR) signal in rats with CKD or UUO, and in cultured 1-hydroxypyrene-induced HK-2 cells. Renoprotective effect of IAld was partially diminished in AHR deficiency mice and HK-2 cells. Our further data showed that treatment with L. johnsonii attenuated kidney lesion by suppressing AHR signal via increasing serum IAld level. Taken together, targeting L. johnsonii might reverse patients with CKD. This study provides a deeper understanding of how microbial-produced tryptophan metabolism affects host disease and discovers potential pathways for prophylactic and therapeutic treatments for CKD patients.

Restless Legs Syndrome in Chronic Kidney Disease- a Systematic Review.

Objectives: The objective of this review is to provide updated information on the epidemiology, correlating factors and treatment of chronic kidney disease associated restless legs syndrome (CKD-A-RLS) in both adult and pediatric population. Materials and Methods: We have reviewed the Medline search and Google Scholar search up to May 2022, using key words restless legs syndrome, chronic kidney disease and hemodialysis and kidney transplant. The reviewed articles were studied for epidemiology, correlating factors, as well as pharmacologic and non-pharmacologic treatment options. Results: Our search revealed 175 articles, 111 were clinical trials or cross- sectional studies and 64 were review articles. All 111 articles were retrieved and studied in detail. Of these, 105 focused on adults and 6 on children. A majority of studies on dialysis patients reported a prevalence between 15-30%, which is notably higher than prevalence of RLS in general population (5-10%). The correlation between presence of CKD-A-RLS with age, gender, abnormalities of hemogram, iron, ferritin, serum lipids, electrolytes and parathyroid hormones were also reviewed. The results were inconsistent and controversial. Limited studies have reported on the treatment of CKD-A-RLS. Non-pharmacological treatment focused on the effect(s) of exercise, acupuncture, massage with different oils and infra-red light whereas, pharmacologic treatment options include the effects of dopaminergic drugs, Alpha2-Delta ligands (gabapentin and pregabalin), vitamins E and C, and intravenous iron infusion. Conclusion: This updated review showed that RLS is two to three times more common in patients with CKD compared to the general population. More patients with CKD-A-RLS demonstrated increased mortality, increased incidence of cardiovascular accident, depression, insomnia and impaired quality of life than those with CKD without RLS. Dopaminergic drugs such as levodopa, ropinirole, pramipexole and rotigotine as well as calcium channel blockers (gabapentin and pregabalin) are helpful for treatment of RLS. High quality studies with these agents are currently underway and hopefully confirm the efficacy and practicality of using these drugs in CKD-A-RLS. Some studies have shown that aerobic exercise and massage with lavender oil can improve symptoms of CKD-A- RLS suggesting that these measures can be useful as adjunct therapy.

Tetrahydrocurcumin Add-On therapy to losartan in a rat model of diabetic nephropathy decreases blood pressure and markers of kidney injury.

Tetrahydrocurcumin (THC), a principal metabolite of curcumin, was tested in a rat model of type 2 diabetes mellitus. THC was administered via daily oral gavage with the lipid carrier polyenylphosphatidylcholine (PPC) as add-on therapy to losartan (angiotensin receptor blocker) to examine effects on kidney oxidative stress and fibrosis. A combination of unilateral nephrectomy, high-fat diet and low-dose streptozotocin was used to induce diabetic nephropathy in male Sprague-Dawley rats. Animals with fasting blood glucose >200 mg/dL were randomized to PPC, losartan, THC + PPC or THC + PPC + losartan. Untreated chronic kidney disease (CKD) animals had proteinuria, decreased creatinine clearance, and evidence of kidney fibrosis on histology. THC + PPC + losartan treatment significantly lowered blood pressure concurrent with increased messenger RNA levels of antioxidant copper-zinc-superoxide dismutase and decreased protein kinase C-α, kidney injury molecule-1 and type I collagen in the kidneys; there was decreased albuminuria and a trend for increased creatinine clearance compared to untreated CKD rats. There was decreased fibrosis on kidney histology in PPC-only and THC-treated CKD rats. Plasma levels of kidney injury molecule-1 were decreased in THC + PPC + losartan animals. In summary, add-on THC to losartan therapy improved antioxidant levels and decreased fibrosis in the kidneys, and lowered blood pressure in diabetic CKD rats.

Intrarenal 1-methoxypyrene, an aryl hydrocarbon receptor agonist, mediates progressive tubulointerstitial fibrosis in mice.

Recent studies have shown that endogenous metabolites act via aryl hydrocarbon receptor (AhR) signalling pathway in tubulointerstitial fibrosis (TIF) pathogenesis. However, the mechanisms underlying endogenous metabolite-mediated AhR activation are poorly characterised. In this study, we conducted untargeted metabolomics analysis to identify the significantly altered intrarenal metabolites in a mouse model of unilateral ureteral obstruction (UUO). We found that the levels of the metabolite 1-methoxypyrene (MP) and the mRNA expression of AhR and its target genes CYP1A1, CYP1A2, CYP1B1 and COX-2 were progressively increased in the obstructed kidney at Weeks 1, 2 and 3. Furthermore, these changes were positively correlated with progressive TIF in UUO mice. In NRK-52E, RAW 264.7 and NRK-49F cells, MP dose-dependently upregulated the mRNA expression of AhR and its four target genes and the protein expression of nuclear AhR, accompanied by the upregulated protein expression of collagen I, α-SMA and fibronectin, as well as downregulated E-cadherin expression. Consistently, oral administration of MP in mice progressively enhanced AhR activity and upregulated profibrotic protein expression in the kidneys; these effects were partially inhibited by AhR knockdown in MP-treated mice and cell lines. In addition, we screened and identified erythro-guaiacylglycerol-ß-ferulic acid ether (GFA), which was isolated from Semen plantaginis, as a new AhR antagonist. GFA significantly attenuated TIF in MP-treated NRK-52E cells and mice by partially antagonising AhR activity. Our results suggest that MP activates AhR signalling, thus mediating TIF through epithelial-mesenchymal transition and macrophage-myofibroblast transition. MP is a crucial metabolite that contributes to TIF via AhR signalling pathway.

Dihydroartemisinin suppresses renal fibrosis in mice by inhibiting DNA-methyltransferase 1 and increasing Klotho.

Renal fibrosis is an unavoidable end result of all forms of progressive chronic kidney diseases (CKD). Discovery of efficacious drugs against renal fibrosis is in crucial need. In a preliminary study we found that a derivative of artemisinin, dihydroartemisinin (DHA), exerted strong renoprotection, and reversed renal fibrosis in adenine-induced CKD mouse model. In this study we investigated the anti-fibrotic mechanisms of DHA, particularly its specific target in renal cells. Renal fibrosis was induced in mice by unilateral ureteral obstruction (UUO) or oral administration of adenine (80 mg · kg-1), the mice received DHA (30 mg · kg-1 · d-1, i.g.) for 14 or 21 days, respectively. We showed that DHA administration markedly attenuated the inflammation and fibrotic responses in the kidneys and significantly improved the renal function in both the renal fibrosis mouse models. In adenine-treated mice, DHA was more effective than 5-azacytidine against renal fibrosis. The anti-fibrotic effects of DHA were also observed in TGF-ß1-treated HK-2 cells. In order to determine the target protein of DHA, we conducted pull-down technology coupled with shotgun proteomics using a small-molecule probe based on the structure of DHA (biotin-DHA). As a results, DNA methyltransferase 1 (DNMT1) was identified as the anti-fibrotic target of DHA in 3 different types of renal cell lines (HK-2, HEK293 and 3T3). We demonstrated that DHA directly bound to Asn 1529 and Thr 1528 of DNMT1 with a Kd value of 8.18 µM. In primary mouse renal tubular cells, we showed that DHA (10 µM) promoted DNMT1 degradation via the ubiquitin-proteasome pathway. DHA-reduced DNMT1 expression effectively reversed Klotho promoter hypermethylation, which led to the reversal of Klotho protein loss in the kidney of UUO mice. This subsequently resulted in inhibition of the Wnt/ß-catenin and TGF-ß/Smad signaling pathways and consequently conferred renoprotection in the animals. Knockdown of Klotho abolished the renoprotective effect of DHA in UUO mice. Our study reveals a novel pharmacological activity for DHA, i.e., renoprotection. DHA exhibits this effect by targeting DNMT1 to reverse Klotho repression. This study provides an evidence for the possible clinical application of DHA in the treatment of renal fibrosis.

1-Hydroxypyrene mediates renal fibrosis through aryl hydrocarbon receptor signalling pathway.

BACKGROUND AND PURPOSE: In chronic kidney disease (CKD), patients inevitably reach end-stage renal disease and require renal transplant. Evidence suggests that CKD is associated with metabolite disorders. However, the molecular pathways targeted by metabolites remain enigmatic. Here, we describe roles of 1-hydroxypyrene in mediating renal fibrosis. EXPERIMENTAL APPROACH: We analysed 5406 urine and serum samples from patients with Stage 1-5 CKD using metabolomics, and 1-hydroxypyrene was identified and validated using longitudinal and drug intervention cohorts as well as 5/6 nephrectomised and adenine-induced rats. KEY RESULTS: We identified correlations between the urine and serum levels of 1-hydroxypyrene and the estimated GFR in patients with CKD onset and progression. Moreover, increased 1-hydroxypyrene levels in serum and kidney tissues correlated with decreased renal function in two rat models. Up-regulated mRNA expression of aryl hydrocarbon receptor and its target genes, including CYP1A1, CYP1A2 and CYP1B1, were observed in patients and rats with progressive CKD. Further we showed up-regulated mRNA expression of aryl hydrocarbon receptor and its three target genes, plus up-regulated nuclear aryl hydrocarbon receptor protein levels in mice and HK-2 cells treated with 1-hydroxypyrene, which caused accumulation of extracellular matrix components. Treatment with aryl hydrocarbon receptor short hairpin RNA or flavonoids inhibited mRNA expression of aryl hydrocarbon receptor and its target genes in 1-hydroxypyrene-induced HK-2 cells and mice. CONCLUSION AND IMPLICATIONS: Metabolite 1-hydroxypyrene was demonstrated to mediate renal fibrosis through activation of the aryl hydrocarbon receptor signalling pathway. Targeting aryl hydrocarbon receptor may be an alternative therapeutic strategy for CKD progression.

Diabetes and the Gut Microbiome.

Gut dysbiosis in diabetes mellitus is associated with decreased short-chain fatty acids and epithelial barrier disruption. Microbial-derived toxins move across the "leaky gut" and incur systemic inflammation and insulin resistance. In children, gut dysbiosis has been associated with risk of developing type 1 diabetes mellitus. In animal models, the obesity phenotype is transferable via microbiota transplantation. Plant-based low protein diets and certain anti-diabetic drugs have been associated with positive microbiome effects. Clinical trials with prebiotics and probiotics have yielded mixed results. Further investigations are needed to evaluate the gut microbiome as a potential therapeutic target for diabetes prevention and management.

Polyporus Umbellatus Protects Against Renal Fibrosis by Regulating Intrarenal Fatty Acyl Metabolites.

Background: Chronic renal failure (CRF) results in significant dyslipidemia and profound changes in lipid metabolism. Polyporus umbellatus (PPU) has been shown to prevent kidney injury and subsequent kidney fibrosis. Methods: Lipidomic analysis was performed to explore the intrarenal profile of lipid metabolites and further investigate the effect of PPU and its main bioactive component, ergone, on disorders of lipid metabolism in rats induced by adenine. Univariate and multivariate statistical analyses were performed for choosing intrarenal differential lipid species in CRF rats and the intervening effect of n-hexane extract of PPU and ergone on CRF rats. Results: Compared with control group, decreased creatinine clearance rate indicated declining kidney function in CRF group. Based on the lipidomics, we identified 65 lipid species that showed significant differences between CRF and control groups. The levels of 12 lipid species, especially fatty acyl lipids including docosahexaenoic acid, docosapentaenoic acid (22n-3), 10,11-Dihydro-12R-hydroxy-leukotriene C4, 3-hydroxydodecanoyl carnitine, eicosapentaenoic acid, hypogeic acid and 3-hydroxypentadecanoic acid had a strong linear correlation with creatinine clearance rate, which indicated these lipid species were associated with impaired renal function. In addition, receiver operating characteristics analysis showed that 12 lipid species had high area under the curve values with high sensitivity and specificity for differentiating CRF group from control group. These changes are related to the perturbation of fatty acyl metabolism. Treatment with PPU and ergone improved the impaired kidney function and mitigated renal fibrosis. Both chemometrics and cluster analyses showed that rats treated by PPU and ergone could be separated from CRF rats by using 12 lipid species. Intriguingly, PPU treatment could restore the levels of 12 lipid species, while treatment with ergone could only reverse the changes of six fatty acids in CRF rats. Conclusion: Altered intrarenal fatty acyl metabolites were implicated in pathogenesis of renal fibrosis. PPU and ergone administration alleviated renal fibrosis and partially improved fatty acyl metabolism. These findings suggest that PPU exerted its renoprotective effect by regulating fatty acyl metabolism as a potential biochemical mechanism. Therefore, these findings indicated that fatty acyl metabolism played an important role in renal fibrosis and could be considered as an effective therapeutic avenue against renal fibrosis.

Impact of Circulating N-Acylethanolamine Levels with Clinical and Laboratory End Points in Hemodialysis Patients.

BACKGROUND: Patients with ESRD on maintenance hemodialysis (MHD) are particularly susceptible to dysregulation of energy metabolism, which may manifest as protein energy wasting and cachexia. In recent years, the endocannabinoid system has been shown to play an important role in energy metabolism with potential relevance in ESRD. N-acylethanolamines are a class of fatty acid amides which include the major endocannabinoid ligand, anandamide, and the endogenous peroxisome proliferator-activated receptor-α agonists, oleoylethanolamide (OEA) and palmitoylethanolamide (PEA). METHODS: Serum concentrations of OEA and PEA were measured in MHD patients and their correlations with various clinical/laboratory indices were examined. Secondarily, we evaluated the association of circulating PEA and OEA levels with 12-month all-cause mortality. RESULTS: Both serum OEA and PEA levels positively correlated with high-density lipoprotein-cholesterol levels and negatively correlated with body fat and body anthropometric measures. Serum OEA levels correlated positively with serum interleukin-6 (IL-6) (rho = 0.19; p = 0.004). Serum PEA and IL-6 showed a similar but nonsignificant trend (rho = 0.12; p = 0.07). Restricted cubic spline analyses showed that increasing serum OEA and PEA both trended toward higher mortality risk, and these associations were statistically significant for PEA (PEA ≥4.7 pmol/mL; reference: PEA <4.7 pmol/mL) after adjustments in a Cox model (hazard ratio 2.99; 95% confidence interval 1.04, 8.64). CONCLUSIONS: In MHD patients, OEA and PEA are significantly correlated with variables related to lipid metabolism and body mass. Additionally, higher serum levels of PEA are associated with mortality risk. Future studies are needed to examine the potential mechanisms responsible for these findings and their clinical implications.