Gut microbiota and its metabolites – molecular mechanisms and management strategies in diabetic kidney disease

Diabetic kidney disease (DKD) is one of the major microvascular complications of diabetes mellitus and is also one of the serious risk factors in cardiovascular events, end-stage renal disease, and mortality. DKD is associated with the diversified, compositional, and functional alterations of gut microbiota. The interaction between gut microbiota and host is mainly achieved through metabolites, which are small molecules produced by microbial metabolism from exogenous dietary substrates and endogenous host compounds. The gut microbiota plays a critical role in the pathogenesis of DKD by producing multitudinous metabolites. Nevertheless, detailed mechanisms of gut microbiota and its metabolites involved in the occurrence and development of DKD have not been completely elucidated. This review summarizes the specific classes of gut microbiota-derived metabolites, aims to explore the molecular mechanisms of gut microbiota in DKD pathophysiology and progression, recognizes biomarkers for the screening, diagnosis, and prognosis of DKD, as well as provides novel therapeutic strategies for DKD

Identification of ULK1 as a novel mitophagy-related gene in diabetic nephropathy

BackgroundAccumulating evidence indicates that mitophagy is crucial for the development of diabetic nephropathy (DN). However, little is known about the key genes involved. The present study is to identify the potential mitophagy-related genes (MRGs) in DN.MethodsFive datasets were obtained from the Gene Expression Omnibus (GEO) database and were split into the training and validation set. Then the differentially expressed MRGs were screened and further analyzed for GO and KEGG enrichment. Next, three algorithms (SVM-RFE, LASSO and RF) were used to identify hub genes. The ROC curves were plotted based on the hub genes. We then used the CIBERSORT algorithm to assess the infiltration of 22 types of immune cells and explore the correlation between hub genes and immune cells. Finally, the Nephroseq V5 tool was used to analyze the correlation between hub genes and GFR in DN patients.ResultsCompared with the tubulointerstitium, the expression of MRGs was more noticeably varied in the glomeruli. Twelve DE-MRGs were identified in glomerular samples, of which 11 genes were down-regulated and only MFN1 was up-regulated. GO and KEGG analysis indicated that several enrichment terms were associated with changes in autophagy. Three genes (MFN1, ULK1 and PARK2) were finally determined as potential hub genes by three algorithms. In the training set, the AUROC of MFN1, ULK1 and PARK2 were 0.839, 0.906 and 0.842. However, the results of the validation set demonstrated that MFN1 and PARK2 had no significant difference in distinguishing DN samples from healthy controls, while the AUROC of ULK1 was 0.894. Immune infiltration analysis using CIBERSORT showed that ULK1 was positively related to neutrophils, whereas negatively related to M1 and M2 macrophages. Finally, ULK1 was positively correlated with GFR in Nephroseq database.ConclusionsULK1 is a potential biomarker for DN and may influence the development of diabetic nephropathy by regulating mitophagy

AT2R (Angiotensin II Type 2 Receptor)-Mediated Regulation of NCC (Na-Cl Cotransporter) and Renal K Excretion Depends on the K Channel, Kir4.1

AT2R (AngII [angiotensin II] type 2 receptor) is expressed in the distal nephrons. The aim of the present study is to examine whether AT2R regulates NCC (Na-Cl cotransporter) and Kir4.1 of the distal convoluted tubule. AngII inhibited the basolateral 40 pS K channel (a Kir4.1/5.1 heterotetramer) in the distal convoluted tubule treated with losartan but not with PD123319. AT2R agonist also inhibits the K channel, indicating that AT2R was involved in tonic regulation of Kir4.1. The infusion of PD123319 stimulated the expression of tNCC (total NCC) and pNCC (phosphorylated NCC; Thr(53)) by a time-dependent way with the peak at 4 days. PD123319 treatment (4 days) stimulated the basolateral 40 pS K channel activity, augmented the basolateral K conductance, and increased the negativity of distal convoluted tubule membrane. The stimulation of Kir4.1 was essential for PD123319-induced increase in NCC because inhibiting AT2R increased the expression of tNCC and pNCC only in wild-type but not in the kidney-specific Kir4.1 knockout mice. Renal clearance study showed that thiazide-induced natriuretic effect was larger in PD123319-treated mice for 4 days than untreated mice. However, this effect was absent in kidney-specific Kir4.1 knockout mice which were also Na wasting under basal conditions. Finally, application of AT2R antagonist decreased the renal ability of K excretion and caused hyperkalemia in wild-type but not in kidney-specific Kir4.1 knockout mice. We conclude that AT2R-dependent regulation of NCC requires Kir4.1 in the distal convoluted tubule and that AT2R plays a role in stimulating K excretion by inhibiting Kir4.1 and NCC

Changes in the expression of MMP2, MMP9, and ColIV in stromal cells in oral squamous tongue cell carcinoma: relationships and prognostic implications

Abstract Background Type IV collagen (ColIV) is the most important scaffold for the basement membrane (BM) proteins, and plays an important role in regulating and limiting tumour invasion and metastasis. Methods Here, we observed the changes in morphology and distribution of type IV collagen (ColIV) in the basement membrane (BM) surrounding nests of carcinoma in 48 patients with oral tongue squamous cell (OTSCC). We examined the correlation between the expressions of ColIV, MMP-2 and MMP-9 and the prognosis of OTSCC patients. The intensity and patterns of expression were assessed immunohistochemically using anti-human mouse monoclonal MMP-2, MMP-9 and Col IV antibodies. Statistical analyses were performed to determine the prognostic correlations of ColIV, MMP-2, and MMP-9 levels. Results MMP-2 and MMP-9 expressions in OTSCC were higher than those in normal oral mucosa and dysplastic oral mucosa group(MMP-2 iOD: 66.40 ± 24.20, 134.69 ± 37.08, and 357.79 ± 116.78; MMP-9 iOD: 88.05 ± 23.85, 307.13 ± 93.22, and 791.31 ± 260.52; in normal, dysplastic oral mucosa, and tumour tissues, respectively, P Conclusions Degradation of ColIV was closely related to increased MMP-2 and MMP-9 expression; MMP-9 have more important function than MMP-2 during the cancer development. Monitoring changes in the expression of ColIV, MMP-2, and MMP-9 may be a useful technique for assessing prognoses in OTSCC patients.</p

Kir4.1/Kir5.1 Activity Is Essential for Dietary Sodium Intake-Induced Modulation of Na-Cl Cotransporter

BACKGROUND: Dietary sodium intake regulates the thiazide-sensitive Na-Cl cotransporter (NCC) in the distal convoluted tubule (DCT). Whether the basolateral, inwardly rectifying potassium channel Kir4.1/Kir5.1 (a heterotetramer of Kir4.1/Kir5.1) in the DCT is essential for mediating the effect of dietary sodium intake on NCC activity is unknown. METHODS: We used electrophysiology, renal clearance techniques, and immunoblotting to examine effects of Kir4.1/Kir5.1 in the DCT and NCC in wild-type and kidney-specific Kir4.1 knockout mice. RESULTS: Low sodium intake stimulated basolateral Kir4.1/Kir5.1 activity, increased basolateral K(+) conductance, and hyperpolarized the membrane. Conversely, high sodium intake inhibited the potassium channel, decreased basolateral K(+) currents, and depolarized the membrane. Low sodium intake increased total and phosphorylated NCC expression and augmented hydrochlorothiazide-induced natriuresis; high sodium intake had opposite effects. Thus, elevated NCC activity induced by low sodium intake was associated with upregulation of Kir4.1/Kir5.1 activity in the DCT, whereas inhibition of NCC activity by high sodium intake was associated with diminished Kir4.1/Kir5.1 activity. In contrast, dietary sodium intake did not affect NCC activity in knockout mice. Further, Kir4.1 deletion not only abolished basolateral K(+) conductance and depolarized the DCT membrane, but also abrogated the stimulating effects induced by low sodium intake on basolateral K(+) conductance and hyperpolarization. Finally, dietary sodium intake did not alter urinary potassium excretion rate in hypokalemic knockout and wild-type mice. CONCLUSIONS: Stimulation of Kir4.1/Kir5.1 by low intake of dietary sodium is essential for NCC upregulation, and inhibition of Kir4.1/Kir5.1 induced by high sodium intake is a key step for downregulation of NCC

Deletion of Kir5.1 Impairs Renal Ability to Excrete Potassium During Increased Dietary Potassium Intake

BACKGROUND: The basolateral potassium channel in the distal convoluted tubule (DCT), comprising the inwardly rectifying potassium channel Kir4.1/Kir5.1 heterotetramer, plays a key role in mediating the effect of dietary potassium intake on the thiazide-sensitive NaCl cotransporter (NCC). The role of Kir5.1 (encoded by Kcnj16) in mediating effects of dietary potassium intake on the NCC and renal potassium excretion is unknown. METHODS: We used electrophysiology, renal clearance, and immunoblotting to study Kir4.1 in the DCT and NCC in Kir5.1 knockout (Kcnj16(-/-) ) and wild-type (Kcnj16(+/+) ) mice fed with normal, high, or low potassium diets. RESULTS: We detected a 40-pS and 20-pS potassium channel in the basolateral membrane of the DCT in wild-type and knockout mice, respectively. Compared with wild-type, Kcnj16(-/-) mice fed a normal potassium diet had higher basolateral potassium conductance, a more negative DCT membrane potential, higher expression of phosphorylated NCC (pNCC) and total NCC (tNCC), and augmented thiazide-induced natriuresis. Neither high- nor low-potassium diets affected the basolateral DCT\u27s potassium conductance and membrane potential in Kcnj16(-/-) mice. Although high potassium reduced and low potassium increased the expression of pNCC and tNCC in wild-type mice, these effects were absent in Kcnj16(-/-) mice. High potassium intake inhibited and low intake augmented thiazide-induced natriuresis in wild-type but not in Kcnj16(-/-) mice. Compared with wild-type, Kcnj16(-/-) mice with normal potassium intake had slightly lower plasma potassium but were more hyperkalemic with prolonged high potassium intake and more hypokalemic during potassium restriction. CONCLUSIONS: Kir5.1 is essential for dietary potassium\u27s effect on NCC and for maintaining potassium homeostasis

Role of WNK4 and kidney-specific WNK1 in mediating the effect of high dietary K+ intake on ROMK channel in the distal convoluted tubule

With-no-lysine kinase 4 (WNK4) and kidney-Specific (KS)-WNK1 regulate ROMK (Kir1.1) channels in a variety of cell models. We now explore the role of WNK4 and KS-WNK1 in regulating ROMK in the native distal convoluted tubule (DCT)/connecting tubule (CNT) by measuring TPNQ (ROMK inhibitor)-sensitive K+ currents with whole-cell recording. TPNQ-sensitive K+ currents in DCT2/CNT of KS-WNK1-/- and WNK4-/- mice were significantly smaller than that of WT mice. In contrast, the basolateral K+ channels (a Kir4.1/5.1 heterotetramer) in the DCT were not inhibited. Moreover, WNK4-/- mice were hypokalemic while KS-WNK1-/- mice had normal plasma K+ level. High K+ (HK) intake significantly increased TPNQ-sensitive K+ currents in DCT2/CNT of WT and WNK4-/- mice but not in KS-WNK1-/- mice. However, TPNQ-sensitive K+ currents in the cortical collecting duct (CCD) were normal not only under control conditions but also significantly increased in response to HK in KS-WNK1-/- mice. This suggests that the deletion of KS-WNK1-induced inhibition of ROMK occurs only in the DCT2/CNT. Renal clearance study further demonstrated that the deletion of KS-WNK1 did not affect the renal ability of K+ excretion under control conditions and during increasing K+ intake. Also, HK intake did not cause hyperkalemia in KS-WNK1-/- mice. We conclude that KS-WNK1 but not WNK4 is required for HK-intake-induced stimulation of ROMK activity in DCT2/CNT. However, KS-WNK1 is not essential for HK-induced stimulation of ROMK in the CCD and the lack of KS-WNK1 does not affect net renal K+ excretion

Deletion of Renal Nedd4-2 Abolishes the Effect of High Sodium Intake (HS) on Kir4.1, ENaC, and NCC and Causes Hypokalemia During High HS

Neural precursor cell expressed developmentally downregulated protein 4-2 (Nedd4-2) regulates the expression of Kir4.1, thiazide-sensitive NaCl cotransporter (NCC), and epithelial Na+ channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN), and Nedd4-2 deletion causes salt-sensitive hypertension. We now examined whether Nedd4-2 deletion compromises the effect of high-salt (HS) diet on Kir4.1, NCC, ENaC, and renal K+ excretion. Immunoblot analysis showed that HS diet decreased the expression of Kir4.1, Ca2+-activated large-conductance K+ channel subunit-α (BKα), ENaCβ, ENaCγ, total NCC, and phospho-NCC (at Thr53) in floxed neural precursor cell expressed developmentally downregulated gene 4-like (Nedd4lfl/fl) mice, whereas these effects were absent in kidney-specific Nedd4-2 knockout (Ks-Nedd4-2 KO) mice. Renal clearance experiments also demonstrated that Nedd4-2 deletion abolished the inhibitory effect of HS diet on hydrochlorothiazide-induced natriuresis. Patch-clamp experiments showed that neither HS diet nor low-salt diet had an effect on Kir4.1/Kir5.1 currents of the distal convoluted tubule in Nedd4-2-deficient mice, whereas we confirmed that HS diet inhibited and low-salt diet increased Kir4.1/Kir5.1 activity in Nedd4lflox/flox mice. Nedd4-2 deletion increased ENaC currents in the ASDN, and this increase was more robust in the cortical collecting duct than in the distal convoluted tubule. Also, HS-induced inhibition of ENaC currents in the ASDN was absent in Nedd4-2-deficient mice. Renal clearance experiments showed that HS intake for 2 wk increased the basal level of renal K+ excretion and caused hypokalemia in Ks-Nedd4-2-KO mice but not in Nedd4lflox/flox mice. In contrast, plasma Na+ concentrations were similar in Nedd4lflox/flox and Ks-Nedd4-2 KO mice on HS diet. We conclude that Nedd4-2 plays an important role in mediating the inhibitory effect of HS diet on Kir4.1, ENaC, and NCC and is essential for maintaining normal renal K+ excretion and plasma K+ ranges during long-term HS diet.NEW & NOTEWORTHY The present study suggests that Nedd4-2 is involved in mediating the inhibitory effect of high salt (HS) diet on Kir4.1/kir5.1 in the distal convoluted tubule, NaCl cotransporter function, and epithelial Na+ channel activity and that Nedd4-2 plays an essential role in maintaining K+ homeostasis in response to a long-term HS diet. This suggests the possibility that HS intake could lead to hypokalemia in subjects lacking proper Nedd4-2 E3 ubiquitin ligase activity in aldosterone-sensitive distal nephron