Caspase-11/GSDMD contributes to the progression of hyperuricemic nephropathy by promoting NETs formation.

Hyperuricemia is an independent risk factor for chronic kidney disease (CKD) and promotes renal fibrosis, but the underlying mechanism remains largely unknown. Unresolved inflammation is strongly associated with renal fibrosis and is a well-known significant contributor to the progression of CKD, including hyperuricemia nephropathy. In the current study, we elucidated the impact of Caspase-11/Gasdermin D (GSDMD)-dependent neutrophil extracellular traps (NETs) on progressive hyperuricemic nephropathy. We found that the Caspase-11/GSDMD signaling were markedly activated in the kidneys of hyperuricemic nephropathy. Deletion of Gsdmd or Caspase-11 protects against the progression of hyperuricemic nephropathy by reducing kidney inflammation, proinflammatory and profibrogenic factors expression, NETs generation, α-smooth muscle actin expression, and fibrosis. Furthermore, specific deletion of Gsdmd or Caspase-11 in hematopoietic cells showed a protective effect on renal fibrosis in hyperuricemic nephropathy. Additionally, in vitro studies unveiled the capability of uric acid in inducing Caspase-11/GSDMD-dependent NETs formation, consequently enhancing α-smooth muscle actin production in macrophages. In summary, this study demonstrated the contributory role of Caspase-11/GSDMD in the progression of hyperuricemic nephropathy by promoting NETs formation, which may shed new light on the therapeutic approach to treating and reversing hyperuricemic nephropathy.

GSDMD and GSDME synergy in the transition of acute kidney injury to chronic kidney disease.

BACKGROUND AND HYPOTHESIS: Acute kidney injury (AKI) could progress to chronic kidney disease (CKD) and the AKI-CKD transition has major clinical significance. A growing body of evidence has unveiled the role of pyroptosis in kidney injury. We postulate that GSDMD and GSDME exert cumulative effects on the AKI-CKD transition by modulating different cellular responses. METHODS: We established an AKI-CKD transition model induced by folic acid in wildtype (WT), Gsdmd-/-, Gsdme-/-, and Gsdmd-/-Gsdme-/- mice. Tubular injury, renal fibrosis and inflammatory responses were evaluated. In vitro studies were conducted to investigate the interplay among tubular cells, neutrophils, and macrophages. RESULTS: Double deletion of Gsdmd and Gsdme conferred heightened protection against AKI, mitigating inflammatory responses, including the formation of neutrophil extracellular traps (NETs), macrophage polarization and differentiation, and ultimately renal fibrosis, compared with wildtype mice and mice with single deletion of either Gsdmd or Gsdme. Gsdme, but not Gsdmd deficiency, shielded tubular cells from pyroptosis. GSDME-dependent tubular cell death stimulated NETs formation and prompted macrophage polarization towards a pro-inflammatory phenotype. Gsdmd deficiency suppressed NETs formation and subsequently hindered NETs-induced macrophage-to-myofibroblast transition (MMT). CONCLUSION: GSDMD and GSDME collaborate to contribute to AKI and subsequent renal fibrosis induced by folic acid. Synchronous inhibition of GSDMD and GSDME could be an innovative therapeutic strategy for mitigating the AKI-CKD transition.

Influence of doctors' perception on the diagnostic status of chronic kidney disease: results from 976 409 individuals with electronic health records in China.

BACKGROUND: The diagnostic status of chronic kidney disease (CKD) and its underlying reasons provide evidence that can improve CKD management. However, the situation in developing countries remains under-investigated. METHODS: Adults with electronic health records (EHRs; 2008-19) in Yinzhou, China were included. The gold standard for CKD was defined as having persistently reduced estimated glomerular filtration rate (eGFR), albuminuria/proteinuria, haematuria or a history of CKD. CKD stages (G1-G5) were defined by eGFR. Clinical diagnosis of CKD in the real world setting was evaluated using International Classification of Diseases (ICD)-10 codes related to primary cause or stages of CKD. The specialty of doctors who administered the serum creatinine (SCr) tests and who made the primary-cause/CKD-staging diagnoses was analysed. The accuracy of CKD-staging codes was assessed. RESULTS: Altogether, 85 519 CKD patients were identified from 976 409 individuals with EHRs. Of them, 10 287 (12.0%) having persistent urinary abnormalities or labelled with CKD-related ICD codes did not receive SCr tests within 12 months before or after the urine tests. Among 75 147 patients who received SCr tests, 46 150 (61.4%) missed any CKD-related codes, 6857 (35.7%) were merely labelled with primary-cause codes, and only 2140 (2.9%) were labelled with CKD-staging codes. The majority of CKD patients (51.6-91.1%) received SCr tests from non-nephrologists, whereas CKD-staging diagnoses were mainly from nephrologists (52.3-64.8%). Only 3 of 42 general hospitals had nephrologists. The CKD-staging codes had high specificity (>99.0%) but low sensitivity (G3-G4: <10.0%). CONCLUSIONS: Under-perception of CKD among doctors, rather than unsatisfactory health-seeking behaviour or low detection rates, was the main cause of under-diagnosis of CKD in China. Intensification of CKD education among doctors with different specialties might bring about immediate effective improvement in the diagnosis and awareness of CKD.

The physiological modulation by intracellular kinases of hippocampal γ-oscillation in vitro.

Hippocampal network oscillations at gamma frequency band (γ-oscillation, 20-80 Hz) are synchronized synaptic activities generated by the interactions between the excitatory and inhibitory interneurons and are associated with higher brain function such as learning and memory. Despite extensive studies about the modulation of intracellular kinases on synaptic transmission and plasticity, little is known about the effects of these kinases on γ-oscillations. In this study, we examined the effects of several critical intracellular kinases such as cyclic AMP-dependent protein kinase (PKA), protein kinase B (PKB)/Akt, protein kinase C (PKC), extracellular-regulated protein kinases (ERK) and AMP-activated protein kinase (AMPK), known to regulate synaptic transmission, on hippocampal γ-oscillations in vitro. We found that AMPK inhibitor but not PKA, PKC, or ERK inhibitor, strongly enhanced the power of γ-oscillation (γ-power) and that Akt inhibitor weakly increased γ-power. Western blot analysis confirmed that AMPK inhibitor reduced the expression of p-AMPK but not total AMPK. By using the slice whole cell voltage-clamp technique, we found that AMPK inhibitor increased the frequency but not amplitude of spontaneous inhibitory postsynaptic currents (sIPSC) and had no effect on either frequency or amplitude of spontaneous excitatory postsynaptic currents (sEPSC). Therefore, AMPK activation negatively modulates hippocampal γ-oscillation via modulation of the inhibitory neurons.

Effect of SGLT2 inhibitor on renal function in patients with type 2 diabetes mellitus: a systematic review and meta-analysis of randomized controlled trials.

OBJECTIVE: This study summarizes the evidence from randomized controlled trials (RCTs) to assess the effects of SGLT2 inhibitors on renal function and albuminuria in patients with type 2 diabetes. MATERIALS/METHODS: We searched PubMed, Web of Science, Cochrane Library and EMBASE for reports published up to March 2018 and included RCTs reporting estimated glomerular filtration rate (eGFR) and/or urine albumin/creatinine ratio (UACR) changes. Data extraction and assessment of research quality based on Cochrane risk biasing tools. Data were calculated to represent the standardized mean difference (SMD) for each study, and the SMDs with 95% confidence intervals (CIs) were pooled using a random effects model. RESULTS: Fifty-one studies were included that evaluated eGFR levels, and 17 studies were included that evaluated UACR levels. A meta-analysis showed that SGLT2 inhibitors had no significant effect on eGFR levels (SMD - 0.02, 95% CI - 0.06, 0.03, p = 0.45), and eGFR reduction was observed in the subsets of the duration of the trial 12 < duration ≤ 26 weeks (SMD - 0.08, 95% CI - 0.13, - 0.02, p = 0.005) and mean baseline eGFR < 60 ml/min per 1.73 square meters (SMD - 0.22, 95% CI - 0.37, - 0.07, p = 0.004). We found that SGLT2 inhibitors reduced UACR levels in patients with type 2 diabetes (SMD - 0.11, 95% CI - 0.17, - 0.05, p = 0.0001). Compared with monotherapy, the combination with other hypoglycemic agents can reduce albuminuria levels (SMD - 0.13, 95% CI - 0.19, - 0.06, p < 0.0001). CONCLUSIONS: The effect of SGLT2 inhibitor on eGFR in patients with T2DM was not statistically significant, but it was effective in reducing albuminuria levels.

Radix Puerariae and Fructus Crataegi mixture inhibits renal injury in type 2 diabetes via decreasing of AKT/PI3K.

BACKGROUND: Radix puerariae (RP) is a herbal medicines for diabetes, mainly because of anti-oxidative, insulin resistance and hypoglycemic effect. Fructus crataegi (FC) also possesses strong antioxidant activity in vitro. This study focused on the effects of herbal mixture of RP and FC (RPFC) on renal protection through a diabetic rat model. METHODS: Type 2 Diabetic model was established with high fat diet followed by injecting rats a low dose of STZ (25 mg/kg body weight). Rats were randomly divided into five groups: normal, high fat diet, diabetes mellitus, high fat diet plus RPFC prevention, and RPFC prevention before diabetes mellitus. RPFC was given to rats daily by intragastric gavage. The blood bio-chemical index and renal pathological changes were examined. The later includes hematoxylin and eosin staining, periodic acid schiff staining, and Masson trichrome staining. Protein levels of were determined by Western blot and immunohistochemical staining. mRNA levels were detected by RT-PCR. RESULTS: Rats prevented with RPFC resulted in decreasing blood glucose with corresponding vehicle treated rats. Glomerulus mesangial matrix expansion, renal capsule constriction, and renal tubular epithelial cell edema were less severe following RPFC prevention. Moreover, RPFC prevention reduced protein levels of PI3K, AKT, α-SMA and collagen IV in the kidney of diabetic rats. CONCLUSION: Combined prevention with RPFC may inhibit the PI3K/AKT pathway in the kidney, thereby prevent renal injury in diabetic rats.

[High glucose dialysate enhances peritoneal fibrosis through upregulating glucose transporters GLUT1 and SGLT1].

Objective: To investigate the role of glucose transporter 1 (GLUT1) and sodium-glucose cotransporter 1 (SGLT1) in high glucose dialysate-induced peritoneal fibrosis. Methods: Thirty six male SD rats were randomly divided into 6 groups (6 in each):normal control group, sham operation group, peritoneal dialysis group (PD group), PD+phloretin group (PD+T group), PD+phlorizin group (PD+Z group), PD+phloretin+phlorizin group (PD+T+Z group). Rat model of uraemia was established using 5/6 nephrotomy, and 2.5% dextrose peritoneal dialysis solution was used in peritoneal dialysis. Peritoneal equilibration test was performed 24 h after dialysis to evaluate transport function of peritoneum in rats; HE staining was used to observe the morphology of peritoneal tissue; and immunohistochemistry was used to detect the expression of GLUT1, SGLT1, TGF-ß1 and connective tissue growth factor (CTGF) in peritoneum. Human peritoneal microvascular endothelial cells (HPECs) were divided into 5 groups:normal control group, peritoneal dialysis group (PD group), PD+phloretin group (PD+T group), PD+phlorezin group (PD+Z group), and PD+phloretin+phlorezin group (PD+T+Z group). Real time PCR and Western blotting were used to detect mRNA and protein expressions of GLUT1, SGLT1, TGF-ß1, CTGF in peritoneal membrane and HPECs. Results:In vivo, compared with sham operation group, rats in PD group had thickened peritoneum, higher ultrafiltration volume, and the mRNA and protein expressions of GLUT1, SGLT1, CTGF, TGF-ß1 were significantly increased (all P<0.05); compared with PD group, thickened peritoneum was attenuated, and the mRNA and protein expressions of GLUT1, SGLT1, CTGF, TGF-ß1 were significantly decreased in PD+T, PD+Z and PD+T+Z groups (all P<0.05). Pearson's correlation analysis showed that the expressions of GLUT1, SGLT1 in peritoneum were positively correlated with the expressions of TGF-ß1 and CTGF (all P<0.05). In vitro, the mRNA and protein expressions of GLUT1, SGLT1, TGF-ß1, CTGF were significantly increased in HPECs of peritoneal dialysis group (all P<0.05), and those in PD+T, PD+Z, and PD+T+Z groups were decreased (all P<0.05). Pearson's correlation analysis showed that the expressions of GLUT1, SGLT1 in HPECs were positively correlated with the expressions of TGF-ß1 and CTGF (all P<0.05). Conclusion: High glucose peritoneal dialysis fluid may promote peritoneal fibrosis by upregulating the expressions of GLUT1 and SGLT1.

Cytotoxic ent-kaurane diterpenoids from Isodon macrophyllus.

Two new ent-kaurane diterpenoids, dayecrystals D-E (1-2), together with nine known compounds, isojaponin A (3), rabdosin A (4), lushanrubescensin J (5), wikstroemioidin B (6), maoyecrystal C (7), rabdosin B (8), isodonal (9), shikokianin (10), and effusanin A (11), were isolated from the leaves of Isodon macrophyllus. The structures of the new compounds were elucidated using 1D and 2D NMR spectroscopy. The (13)C-NMR spectral data of compound 4 are reported for the first time. All of the compounds were tested for their cytotoxicities against DU145 and LoVo human tumor cells. Compounds 4, 10, and 11 showed inhibitory effects on DU145 cells with IC(50) values 5.90, 4.24, and 3.16 microM, and LoVo cells with IC(50) values 14.20, 17.55, and 3.02 microM, respectively.

[Chemical constituents of Isodon macrophyllus (II)].

OBJECTIVE: To study the chemical constituents of Isodon macrophyllus. METHODS: Compounds were spearated and purified by silica gel column chromatography and their structures were elucidated on the basis of spectral data and chemical evidences. RESULTS: Six compounds were obtained from EtOAc extract of leaves of Isodon macroplhllus and identified as rabdophyllin H(I), lushan-ruhescensins F(II), maovecrystal J(III), Taibaijap onicain A(IV), rabdosinate(V), dancostero(VI). CONCLUSION: Co mpounds II-IV were isolated from this plant for the first time.

[Effect of calpain on the degradation of tau protein in rat brain cortex extracts].

Calpain is a calcium-activated protease and has two ubiquitously distributed mammalian isoforms, namely calpain 1 (calpain I, mu-calpain and CAPN1) and calpain 2 (calpain II, m-calpain and CAPN2). Calpains regulate the function of many proteins by limited proteolysis. To determine the nature of different subtypes of calpain on degradation of microtubule-associated protein tau, the rat cortex extracts were incubated with 0.2 mmol/L, 1 mmol/L, 3 mmol/L and 5 mmol/L of CaCl(2 )for 15 min at 37 degrees C, respectively, and it was found that Ca(2+) treatment at concentrations 1-5 mmol/L led to significant proteolysis of the tau protein and this degradation was blocked by calpain inhibitor, calpeptin. In addition, when the extracts containing 1 mmol/L CaCl(2 )were treated with mu-calpain inhibitor (0.05 micromol/L of calpastatin) or m-calpain inhibitor (100 micromol/L calpain inhibitor IV) or both, the Ca(2+)-induced degradation of tau protein was blocked to about 8.6% 92.5% and 97.8% compared with the group with 1 mmol/L CaCl(2), respectively. These data suggest that both mu-calpain and m-calpain in brain cortex extracts are activated by Ca(2+) and both of them degraded tau protein, although, m-calpain plays a more important role in proteolysis of the tau protein.