The anti-renal amyloidosis potential of rosemary extract: A multidimensional mechanistic study

This study aims to investigate the mechanism of Buyang Huanwu Decoction and Didang Decoction regulating activating transcription factor 6(ATF6)/thioredoxin domain containing 5(TXNDC5) signaling axis to mediate macrophage-myofibroblast transformation(MMT) pathway in chronic kidney disease. Twenty-four SPF SD rats were randomly divided into a blank group(n=9) and a modeling group(n=15). The chronic kidney disease model was established by gavage of adenine for 28 days. After successful modeling, the modeling group was randomly divided into a model group and a Buyang Huanwu Decoction and Didang Decoction group. The blank group and the model group were treated with the same volume of physiological saline, and the Buyang Huanwu Decoction and Didang Decoction group was treated by gavage of Buyang Huanwu Decoction and Didang Decoction(13.8 g·kg~(-1)) daily for 28 days. The 24 h urine was collected on the 28th and 56th days of the experiment, and the 24 h urine protein content was detected. After the last administration, all rats were weighed and anesthetized. The kidney weight of the rats was measured, and the renal index was evaluated. The content of serum creatinine(Scr) and blood urea nitrogen(BUN) was detected by an automatic biochemical analyzer, and the concentrations of serum interleukin-1β(IL-1β), tumor necrosis factor-α(TNF-α), interleukin-10(IL-10), and arginase-1(Arg-1) were detected by enzyme-linked immunosorbent assay. The pathological changes of renal tissue were observed by hematoxylin-eosin staining and Masson staining, and the collagen volume fraction(CVF) was calculated. Transmission electron microscopy(TEM) was used to observe the ultrastructural changes of mitochondria in rat renal tissue cells. Multiplex immunofluorescence was used to detect the co-localization of cluster of differentiation 68(CD68)~+α-smooth muscle actin(α-SMA)~+ and CD68~+α-SMA~+collagen type Ⅰ(COL-Ⅰ)~+ in the kidney. Moreover, real-time quantitative polymerase chain reaction was utilized to quantify the mRNA levels of ATF6, TXNDC5, transforming growth factor-β1(TGF-β1), and α-SMA in renal tissue, and the protein levels of ATF6, TXNDC5, and TGF-β1 in the renal tissue were determined by Western blot. The results showed that compared with the blank group, the model group showed a significantly decreased body weight and significantly increased 24 h urine protein, kidney weight, and renal index. The serum levels of Scr, BUN, IL-1β, and TNF-α were significantly increased, while the serum levels of IL-10 and Arg-1 were significantly decreased. Furthermore, the pathological changes of renal tissue were severe, and CVF was significantly increased. The mitochondrial structure of kidney tissue was disordered. High expression of CD68~+α-SMA~+ cells was found in renal tissue, and CD68~+α-SMA~+ MMT cells significantly co-expressed COL-Ⅰ. Finally, the expression of α-SMA mRNA in renal tissue was significantly increased, and the mRNA and protein expression levels of ATF6, TXNDC5, and TGF-β1 in renal tissue were significantly increased. Buyang Huanwu Decoction and Didang Decoction significantly increased the body weight and the levels of IL-10 and Arg-1 in the serum of chronic kidney disease rats and significantly decreased the content of 24 h urine protein, kidney weight, renal index, and the serum levels of Scr, BUN, IL-1β, and TNF-α. The pathological damage of renal tissue was significantly improved, and the CVF was significantly decreased. The ultrastructural damage of renal tissue was significantly improved, as evidenced by TEM. Moreover, Buyang Huanwu Decoction and Didang Decoction decreased the expression of CD68~+α-SMA~+ and CD68~+α-SMA~+COL-Ⅰ~+ in renal tissue and down-regulated the expression level of α-SMA mRNA and the mRNA and protein levels of ATF6, TXNDC5, and TGF-β1 in renal tissue. In conclusion, Buyang Huanwu Decoction and Didang Decoction can regulate ATF6/TXNDC5 signaling pathway, down-regulate TGF-β1 expression, reduce the occurrence of MMT in chronic kidney disease, reduce renal fibrosis, and improve renal injury, so as to play a protective role in kidney.

To observe the effect of acupuncture on serum gastrin content and urinary sodium excretion in spontaneously hypertensive rat (SHR), so as to explore its potential mechanism in the treatment of hypertension. Thirty-two male SHRs were randomly divided into model, hydrochlorothiazide, acupuncture and sham acupuncture groups, with 8 rats in each group, and 8 male Wistar-kyoto rats were taken as the control group. Rats in the hydrochlorothiazide group received gavage of hydrochlorothiazide solution (10 mg·kg-1·d-1), once daily for 4 weeks. Acupuncture was applied to bilateral "Renying" (ST9) and "Zusanli" (ST36) or non-acupoint on both sides for rats in the acupuncture and sham acupuncture groups, with manual stimulation every 10 minutes for a total of 20 minutes, once a day for a total of 4 weeks. The systolic blood pressure of the tail-artery was measured before and 1, 2, 3, and 4 weeks after the intervention. HE staining was used to observe the pathological changes in renal tissue. Serum gastrin contents were detected using enzyme-linked immunosorbent assay (ELISA). Urinary sodium content was measured by atomic absorption spectrometry. The expression levels of cholecystokinin B receptor (CCKBR) and Na+/K+-ATPase proteins in renal tissue were detected by Western blot, and the mRNA expression levels of CCKBR and the α1 subunit of Na+/K+- ATPase (ATP1A1) were detected by fluorescence quantitative PCR. Compared with the control group, the systolic blood pressure of the tail artery in the model group were increased significantly before intervention and at the 1st, 2nd, 3rd and 4th weeks of intervention (P<0.05). Before intervention, the 24 h urine volume of the model, hydrochlorothiazide, acupuncture and sham acupuncture groups were decreased significantly (P<0.05). After intervention, the 24 h urine volume and urinary sodium excretion in the model group were significantly decreased (P<0.05)；the expression levels of CCKBR protein and mRNA in renal tissue were significantly decreased (P<0.05)；the expression levels of Na+/K+-ATPase protein and ATP1A1 mRNA were significantly increased (P<0.05)；the glomerulus was mildly congested with a small amount of lymphocyte infiltration. Compared with the model group, the systolic blood pressure of the tail artery in the hydrochlorothiazide and the acupuncture groups were decreased significantly at the 2nd, 3rd and 4th weeks of intervention (P<0.05)；the 24 h urine volume and urinary sodium excretion in the hydrochlorothiazide and the acupuncture groups were significantly increased (P<0.05)；the serum gastrin content and the expression levels of CCKBR protein and mRNA in renal tissue were significantly increased (P<0.05)；the expression levels of Na+/K+-ATPase protein and ATP1A1 mRNA were significantly decreased (P<0.05)；there were no obvious pathological changes in renal tissue. A small number of lymphocyte focal infiltration around blood vessels was observed in the kidney tissue of the sham acupuncture group. Acupuncture can significantly reduce the tail artery systolic blood pressure of SHR, which may be related to its effect in increasing serum gastrin content and CCKBR expression, inhibiting sodium pump reabsorption, thus promoting urinary sodium excretion.

To investigate the effects of astragaloside IV (AS-IV) on podocyte injury of diabetic nephropathy (DN) and reveal its potential mechanism. In in vitro experiment, podocytes were divided into 4 groups, normal, high glucose (HG), inositol-requiring enzyme 1 (IRE-1) α activator (HG+thapsigargin 1 µmol/L), and IRE-1α inhibitor (HG+STF-083010, 20 µmol/L) groups. Additionally, podocytes were divided into 4 groups, including normal, HG, AS-IV (HG+AS-IV 20 µmol/L), and IRE-1α inhibitor (HG+STF-083010, 20 µmol/L) groups, respectively. After 24 h treatment, the morphology of podocytes and endoplasmic reticulum (ER) was observed by electron microscopy. The expressions of glucose-regulated protein 78 (GRP78) and IRE-1α were detected by cellular immunofluorescence. In in vivo experiment, DN rat model was established via a consecutive 3-day intraperitoneal streptozotocin (STZ) injections. A total of 40 rats were assigned into the normal, DN, AS-IV [AS-IV 40 mg/(kg·d)], and IRE-1α inhibitor [STF-083010, 10 mg/(kg·d)] groups (n=10), respectively. The general condition, 24-h urine volume, random blood glucose, urinary protein excretion rate (UAER), urea nitrogen (BUN), and serum creatinine (SCr) levels of rats were measured after 8 weeks of intervention. Pathological changes in the renal tissue were observed by hematoxylin and eosin (HE) staining. Quantitative reverse transcription-polymerase chain reaction (RT-PCR) and Western blot were used to detect the expressions of GRP78, IRE-1α, nuclear factor kappa Bp65 (NF-κBp65), interleukin (IL)-1β, NLR family pyrin domain containing 3 (NLRP3), caspase-1, gasdermin D-N (GSDMD-N), and nephrin at the mRNA and protein levels in vivo and in vitro, respectively. Cytoplasmic vacuolation and ER swelling were observed in the HG and IRE-1α activator groups. Podocyte morphology and ER expansion were improved in AS-IV and IRE-1α inhibitor groups compared with HG group. Cellular immunofluorescence showed that compared with the normal group, the fluorescence intensity of GRP78 and IRE-1α in the HG and IRE-1α activator groups were significantly increased whereas decreased in AS-IV and IRE-1α inhibitor groups (P<0.05). Compared with the normal group, the mRNA and protein expressions of GRP78, IRE-1α, NF-κ Bp65, IL-1β, NLRP3, caspase-1 and GSDMD-N in the HG group was increased (P<0.05). Compared with HG group, the expression of above indices was decreased in the AS-IV and IRE-1α inhibitor groups, and the expression in the IRE-1α activator group was increased (P<0.05). The expression of nephrin was decreased in the HG group, and increased in AS-IV and IRE-1α inhibitor groups (P<0.05). The in vivo experiment results revealed that compared to the normal group, the levels of blood glucose, triglyceride, total cholesterol, BUN, blood creatinine and urinary protein in the DN group were higher (P<0.05). Compared with DN group, the above indices in AS-IV and IRE-1α inhibitor groups were decreased (P<0.05). HE staining revealed glomerular hypertrophy, mesangial widening and mesangial cell proliferation in the renal tissue of the DN group. Compared with the DN group, the above pathological changes in renal tissue of AS-IV and IRE-1α inhibitor groups were alleviated. Quantitative RT-PCR and Western blot results of GRP78, IRE-1α, NF-κ Bp65, IL-1β, NLRP3, caspase-1 and GSDMD-N were consistent with immunofluorescence analysis. AS-IV could reduce ERS and inflammation, improve podocyte pyroptosis, thus exerting a podocyte-protective effect in DN, through regulating IRE-1α/NF-κ B/NLRP3 signaling pathway.

Selection of modeling time for type 2 diabetes mellitus mouse

To observe the expression of cellular Fas-associated death domain-like interleukin-1β converting enzyme inhibit protein (c-FLIP) in sepsis mice with acute kidney injury (SAKI) and explore its significance. Thirty male ICR mice were divided into the normal control group (Normal group), sham operation group (Sham group) and SAKI group by random number table method, with 10 mice in each group. The SAKI model of mice was established by cecal ligation and puncture (CLP); the Sham group was not ligated and the cecum was not punctured, and other surgical procedures were the same as the SAKI group; the Normal group did not experience any treatment. The serum and renal tissues of mice in each group were harvested 24 hours after CLP model establishment. The levels of serum creatinine (SCr) and blood urea nitrogen (BUN) were detected by enzyme linked immunosorbent assay (ELISA). The renal tissues were stained with hematoxylin-eosin (HE), and the pathological changes of renal tissues were observed under light microscope and the severity of injury was determined. The expression of c-FLIP in renal tissues was detected by immunohistochemistry. The expression of c-FLIP, Bax and caspase-3 protein in renal tissue was detected by Western Blot. The correlation between c-FLIP expression and Bax, caspase-3 protein expressions in renal tissues were analyzed by Pearson test. In the Normal group and the Sham group, the renal tubular epithelial cells were regular and intact, and no interstitial inflammatory cell infiltration was observed; the renal injury score was both 1.30±0.48; immunohistochemistry showed a large amount of c-FLIP positive expression in renal tubular epithelial cells (IA: 120.20±3.87, 116.70±3.46); Western Blot showed high expression of c-FLIP in renal tissues (c-FLIP/GAPDH: 0.99±0.01, 0.98±0.02), and low expressions of Bax and caspase-3 (Bax/GAPDH: 0.16±0.04, 0.19±0.03, caspase-3/GAPDH: 0.24±0.04, 0.23±0.05). Compared with the Sham group, in the SAKI group, renal tubular epithelial cells were degenerated and necrosis, and a large number of interstitial inflammatory cells infiltrated, the renal injury score was significantly increased (4.60±0.52 vs. 1.30±0.48, P < 0.01); the levels of SCr and BUN were significantly increased [SCr (μmol/L): 193.90±13.54 vs. 24.50±3.78, BUN (mmol/L): 81.60±7.26 vs. 5.20±0.92, both P < 0.01]; the c-FLIP positive cells in renal tissues was significantly reduced (IA: 17.11±0.82 vs. 116.70±3.46, P < 0.01); the expression of c-FLIP protein in renal tissues was significantly decreased (c-FLIP/GAPDH: 0.29±0.03 vs. 0.98±0.02, P < 0.01), while the expressions of Bax and caspase-3 protein were significantly increased (Bax/GAPDH: 0.87±0.06 vs. 0.19±0.03, caspase-3/GAPDH: 0.88±0.07 vs. 0.23±0.05, both P < 0.01]. The correlation analysis showed that the c-FLIP protein was significantly negatively correlated with Bax (r = -0.468, P = 0.029) and caspase-3 protein expressions (r = -0.663, P = 0.004). The expression level of c-FLIP protein was significantly down-regulated in renal tissue of SAKI, and its down-regulation mechanism was associated with increased apoptosis of renal tubular epithelial cells, which could be an effective target for the treatment of SAKI.

Objective To observe the expression of cellular Fas-associated death domain-like interleukin-1β converting enzyme inhibit protein (c-FLIP) in sepsis mice with acute kidney injury (SAKI) and explore its significance. Methods Thirty male ICR mice were divided into the normal control group (Normal group), sham operation group (Sham group) and SAKI group by random number table method, with 10 mice in each group. The SAKI model of mice was established by cecal ligation and puncture (CLP); the Sham group was not ligated and the cecum was not punctured, and other surgical procedures were the same as the SAKI group; the Normal group did not experience any treatment. The serum and renal tissues of mice in each group were harvested 24 hours after CLP model establishment. The levels of serum creatinine (SCr) and blood urea nitrogen (BUN) were detected by enzyme linked immunosorbent assay (ELISA). The renal tissues were stained with hematoxylin-eosin (HE), and the pathological changes of renal tissues were observed under light microscope and the severity of injury was determined. The expression of c-FLIP in renal tissues was detected by immunohistochemistry. The expression of c-FLIP, Bax and caspase-3 protein in renal tissue was detected by Western Blot. The correlation between c-FLIP expression and Bax, caspase-3 protein expressions in renal tissues were analyzed by Pearson test. Results In the Normal group and the Sham group, the renal tubular epithelial cells were regular and intact, and no interstitial inflammatory cell infiltration was observed; the renal injury score was both 1. 30±0. 48; immunohistochemistry showed a large amount of c-FLIP positive expression in renal tubular epithelial cells (IA: 120. 20±3. 87, 116. 70±3. 46); Western Blot showed high expression of c-FLIP in renal tissues (c-FLIP/GAPDH: 0. 99±0. 01, 0. 98±0. 02), and low expressions of Bax and caspase-3 (Bax/GAPDH: 0. 16±0. 04, 0. 19±0. 03, caspase-3/GAPDH: 0. 24±0. 04, 0. 23±0. 05). Compared with the Sham group, in the SAKI group, renal tubular epithelial cells were degenerated and necrosis, and a large number of interstitial inflammatory cells infiltrated, the renal injury score was significantly increased (4. 60±0. 52 vs. 1. 30±0. 48, P < 0. 01); the levels of SCr and BUN were significantly increased [SCr (μmol/L): 193. 90±13. 54 vs. 24. 50±3. 78, BUN (mmol/L): 81. 60±7. 26 vs. 5. 20±0. 92, both P < 0. 01]; the c-FLIP positive cells in renal tissues was significantly reduced (IA: 17. 11±0. 82 vs. 116. 70±3. 46, P < 0. 01); the expression of c-FLIP protein in renal tissues was significantly decreased (c-FLIP/GAPDH: 0. 29±0. 03 vs. 0. 98±0. 02, P < 0. 01), while the expressions of Bax and caspase-3 protein were significantly increased (Bax/GAPDH: 0. 87±0. 06 vs. 0. 19±0. 03, caspase-3/GAPDH: 0. 88±0. 07 vs. 0. 23±0. 05, both P < 0. 01]. The correlation analysis showed that the c-FLIP protein was significantly negatively correlated with Bax (r = -0. 468, P = 0. 029) and caspase-3 protein expressions (r = -0. 663, P = 0. 004). Conclusions The expression level of c-FLIP protein was significantly down-regulated in renal tissue of SAKI, and its down-regulation mechanism was associated with increased apoptosis of renal tubular epithelial cells, which could be an effective target for the treatment of SAKI. Key words: c-FLIP; Sepsis; Acute kidney injury; Apoptosis

Effects of Yiniao Recipe on serum antidiuretic hormone level and plasma ratio of cAMP to cGMP in rats with kidney-yang deficiency

The aim of the present study was to examine the protective effect of curcumin (CUR) and curcumin nanoparticles (CURNPs) as antioxidants on the nephrotoxicity induced by lead (Pb). CUR (60 mg/kg b.w) and CURNPs (30 mg/kg b.w) were evaluated using Pb (30 mg/kg b.w.) induced oxidative renal damage in rats. Administration of Pb significantly (p<0.001) increased the levels of renal function test such as creatinine (CRN) and blood urea nitrogen (BUN) and uric acid (UA). Furthermore, Pb significantly decreased glutathione (GSH) levels, superoxide dismutase (SOD), glutathione reductase (GR), catalase (CAT) and glutathione peroxidase (GPx) activities (p<0.001). CUR and CURNPs displayed a renal protective effect as evident by significant decrease in CRN, BUN, UA levels and elevated the antioxidant status of renal tissues. The administration of Pb also induced histopathological changes in renal tissue; however, in groups treated by CUR and CURNPs, there was amelioration in pathological changes in renal tissue. These findings suggest that Nano CUR could be a potential compound in combating the oxidative damage of renal tissue as it increases curcumin bioavailability.

Quercetin-targeted AKT1 regulates the Raf/MEK/ERK signaling pathway to protect against doxorubicin-induced nephropathy in mice

To investigate the effects of liriodendrin on the intestinal flora and the ferroptosis signaling pathway in renal tissue of rats with sepsis-induced acute kidney injury (AKI). Thirty male Sprague-Dawley (SD) rats were randomly divided into sham operation group (Sham group), sepsis model induced by cecal ligation and puncture group (CLP group), and liriodendrin intervention group (CLP+LIR group), with 10 rats in each group. The CLP+LIR group was given 0.2 mL of 100 mg/kg liriodendrin by gavage 2 hours before modeling; Sham group and CLP group were given the same volume of normal saline by gavage. The samples were collected after anesthesia 24 hours after modeling. The pathological changes of renal tissue were observed by hematoxylin-eosin (HE) staining. The levels of inflammatory factors such as tumor necrosis factor-α (TNF-α), interleukins (IL-1β, IL-6) were detected by enzyme linked immunosorbent assay (ELISA). The levels of renal function indicators such as creatinine (Cr), and urea nitrogen (UREA) in peripheral blood, and the content of malondialdehyde (MDA) and Fe2+ in renal tissue were detected. Western blotting was used to detect the expressions of nuclear factor E2-related factor 2 (Nrf2), glutathione peroxidase 4 (GPX4) and heme oxygenase-1 (HO-1) in renal tissues. The changes of intestinal flora were detected by 16S rDNA high-throughput sequencing. Compared with the Sham group, the CLP group showed significantly enlarged glomeruli, noticeable renal interstitial edema, disorganized kidney tissue, and significantly increased pathological scores. The contents of TNF-α, IL-1β, IL-6, Cr, and UREA in peripheral blood and the levels of MDA and Fe2+ in renal tissue were significantly increased. The protein expressions of Nrf2, GPX4, and HO-1 in renal tissue were significantly down-regulated. The species richness of intestinal flora decreased significantly, and the relative abundances of pathogenic bacteria such as Morganella, Citrobacter, Proteus, Klebsiella, Shigella, Aggregatibacter, and Enterococcus increased significantly, while the relative abundances of beneficial bacteria such as Butyricimonas, Veillonella, Prevotella, Lactobacillus, Bifidobacterium, and Ruminococcus decreased significantly. Compared with the CLP group, CLP+LIR group could significantly reduce the pathological damage of renal tissue, the pathological score significantly decreased (1.80±0.84 vs. 4.20±1.30, P < 0.05), and improve the composition of intestinal flora, reduce the relative abundances of pathogenic bacteria such as Proteus, Klebsiella, Shigella, Aggregatibacter, and Enterococcus, and significantly increase the relative abundances of Lactobacillus, Bifidobacterium, and Ruminococcus, significantly reduce the contents of TNF-α, IL-1β, IL-6, Cr, and UREA in peripheral blood and the levels of MDA and Fe2+ in renal tissue [blood TNF-α (ng/L): 191.31±7.23 vs. 254.90±47.89, blood IL-1β (ng/L): 11.15±4.04 vs. 23.06±1.67, blood IL-6 (ng/L): 163.20±17.83 vs. 267.69±20.92, blood Cr (μmol/L): 24.14±4.25 vs. 41.17±5.43, blood UREA (mmol/L): 4.59±0.90 vs. 8.01±1.07, renal MDA (μmol/g): 9.67±0.46 vs. 16.05±0.88, renal Fe2+ (mg/g): 0.71±0.07 vs. 0.93±0.04, all P < 0.05], and increase the protein expressions of Nrf2, GPX4, and HO-1 (Nrf2/GAPDH: 1.21±0.01 vs. 0.39±0.01, GPX4/GAPDH: 0.74±0.04 vs. 0.48±0.04, HO-1/GAPDH: 0.91±0.01 vs. 0.41±0.02, all P < 0.05). Liriodendrin has an obvious protective effect on sepsis-induced AKI. The mechanism may involve regulating the intestinal flora, increasing the activation of the Nrf2/HO-1/GPX4 signaling pathway in renal tissue, and reducing ferroptosis.

Objective To evaluate the role of thioredoxin-interacting protein(TXNIP)/oligomerization domain-like receptor family pyrin domain-containing 3(NLRP3)signaling pathway in renal ischemia-reperfusion(I/R)injury in diabetic rats. Methods Pathogen-free healthy male Sprague-Dawley rats, aged 8-12 weeks, weighing 200-220 g, were used in the study.Diabetes mellitus was induced by intraperitoneal injection of 1% streptozotocin 65 mg/kg and confirmed by blood glucose ≥16.7 mmol/L 3 days later.Twenty-four diabetic rats were divided into 3 groups(n=8 each)using a random number table: sham operation group(group S), renal I/R group(group I/R)and resveratrol(TXNIP inhibitor)group(group R). Resveratrol 10 mg/kg was intraperitoneally injected every day for 7 consecutive days starting from 3rd week after successful establishment of the model in group R. At 4th week after successful establishment of the model, renal I/R was produced by occlusion of bilateral renal pedicles for 25 min followed by reperfusion in anesthetized rats in group R. The animals were sacrificed at 48 h of reperfusion, and renal specimens were obtained for microscopic examination of pathologic changes and for measurement of malondialdehyde(MDA)content, superoxide dismutase(SOD)activity and superoxide anion scavenging capability(using colorimetric method), interleukin-1beta(IL-1β)and IL-18 contents(by enzyme-linked immunosorbent assay), cell apoptosis(using TUNEL)and expression of TXNIP, NLRP3 and caspase-1 in renal tissues(using Western blot). Blood samples were obtained from the left ventricle for determination of serum urea nitrogen(BUN)and creatinine(Cr)concentrations. Results Compared with group S, the serum Cr concentration and apoptosis index were significantly increased, superoxide anion scavenging capability in renal tissues was decreased, and the expression of TXNIP, NLRP3 and caspase-1 was up-regulated in I/R and R groups, and the serum BUN concentration and contents of MDA, IL-1β and IL-18 in renal tissues were increased, the SOD activity was decreased(P<0.05), and the pathological changes of renal tissues were aggravated in group I/R.Compared with group I/R, the serum BUN and Cr concentrations were significantly decreased, the contents of MDA, IL-1β and IL-18 and apoptosis index were decreased, the SOD activity and superoxide anion scavenging capability were increased, the expression of TXNIP, NLRP3 and caspase-1 was down-regulated(P<0.05), and the pathological changes of renal tissues were significantly attenuated in group R. Conclusion The pathophysiological mechanism of renal I/R injury is associated with the activation of TXNIP/NLRP3 signaling pathway in diabetic rats. Key words: Thioredoxins; NLR family, pyrin domain-containing 3 protein; Diabetes mellitus; Kindey; Reperfusion injury

This study investigated the effect of multi-glycosides of Tripterygium wilfordii(GTW) on renal injury in diabetic kidney disease(DKD) rats through Nod-like receptor protein 3(NLRP3)/cysteine-aspartic acid protease-1(caspase-1)/gsdermin D(GSDMD) pyroptosis pathway and the mechanism. To be specific, a total of 40 male SD rats were randomized into the normal group(n=8) and modeling group(n=34). In the modeling group, a high-sugar and high-fat diet and one-time intraperitoneal injection of streptozotocin(STZ) were used to induce DKD in rats. After successful modeling, they were randomly classified into model group, valsartan(Diovan) group, and GTW group. Normal group and model group were given normal saline, and the valsartan group and GTW group received(ig) valsartan and GTW, respectively, for 6 weeks. Blood urea nitrogen(BUN), serum creatinine(Scr), alanine ami-notransferase(ALT), albumin(ALB), and 24 hours urinary total protein(24 h-UTP) were determined by biochemical tests. The pathological changes of renal tissue were observed based on hematoxylin and eosin(HE) staining. Serum levels of interleukin-1β(IL-1β) and interleukin-18(IL-18) were detected by enzyme-linked immunosorbent assay(ELISA). Western blot was used to detect the expression of pyroptosis pathway-related proteins in renal tissue, and RT-PCR to determine the expression of pyroptosis pathway-related genes in renal tissue. Compared with the normal group, the model group showed high levels of BUN, Scr, ALT, and 24 h-UTP and serum levels of IL-1β and IL-18(P&lt;0.01), low level of ALB(P&lt;0.01), severe pathological damage to kidney, and high protein and mRNA levels of NLRP3, caspase-1, and GSDMD in renal tissue(P&lt;0.01). Compared with the model group, valsartan group and GTW group had low levels of BUN, Scr, ALT, and 24 h-UTP and serum levels of IL-1β and IL-18(P&lt;0.01), high level of ALB(P&lt;0.01), alleviation of the pathological damage to the kidney, and low protein and mRNA levels of NLRP3, caspase-1, and GSDMD in renal tissue(P&lt;0.01 or P&lt;0.05). GTW may inhibit pyroptosis by decreasing the expression of NLRP3/caspase-1/GSDMD in renal tissue, thereby relieving the inflammatory response of DKD rats and the pathological injury of kidney.

Long noncoding (Lnc)RNA np_5318 has been proved to be involved in renal injury, while its functionality in renal ischemia-reperfusion (I/R) injury is unknown. Therefore, the present study aimed to investigate the role of lncRNA np_5318 in the development of renal I/R injury. Renal I/R injury model and I/R cell model were established in vitro. The expression of np_5318 in I/R cell was inhibited by small interfering (si)-np_5318 and increased by pc-np_5318. Renal function was detected and evaluated by automatic biochemical tests. Immunohistochemical staining was performed to detect the expression cluster of differentiation (CD)31, transforming growth factor (TGF)-β1 and (mothers against decapentaplegic homolog 3) Smad3 in renal tissue. The interaction between np_5318 and Smad3 was verified by chromatin immunoprecipitation (ChIP). Western blotting was performed to detect the expression levels of TGF-β1, Smad3 and phosphorylated (p)-Smad3 in renal tissue and renal cells. Expression of np_5318 in renal tissue and renal cells was detected by reverse transcription-quantitative PCR. Relative cell viability was confirmed by MTT assay. Renal function was impaired and pathological changes in renal tissue were observed in the renal I/R injury group, indicating the renal I/R injury model was successfully established. Compared with the sham group, the expression level of np_5318 significantly increased in the renal I/R injury group. ChIP data confirmed the interaction between np_5318 and Smad3. The expression of TGF-β1, Smad3 and p-Smad3 in renal tissue was also significantly increased in the renal I/R injury group. Furthermore, the I/R cell model in vitro was successfully constructed and np_5318 in I/R group was significantly increased compared with the control group. Cell growth was significantly suppressed in the I/R group compared with the control group. Additionally, transfection with pc-np_5318 significantly inhibited cell growth of I/R cells at 48 and 72 h. While inhibition of np_5318 by si-np_5318 significantly increased the cell growth of I/R cells at 48 and 72 h. Moreover, the level of TGF-β1, p-Smad3 and Smad3 was significantly increased in the I/R group compared with the control group, and transfection with pc-np_5318 significantly increased the level of TGF-β1, p-Smad3 and Smad3. While inhibition of np_5318 by si-np_5318 significantly suppressed the level of TGF-β1, p-Smad3 and Smad3.LncRNA np_5318 may participate in the development of renal I/R injury through TGF-β/Smad signaling pathway.

This study examined the effect of sulodexide on podocyte injury in rats with adriamycin nephropathy (AN). A total of 36 healthy male SD rats were randomly assigned to three groups: control group, AN group and sulodexide treatment group. Rat models of AN were established by a single tail intravenous injection of adriamycin (6.5 mg/kg) in both AN group and sulodexide treatment group. Sulodexide (10 mg/kg) was administered the rats in the treatment group once daily by garage from the first day of model establishment until the 14th day or the 28th day. Samples of 24-h urine and renal cortex tissues were harvested at day 14, 28 after the model establishment. Excretion of 24-h urinary protein was measured by Coomassie brilliant blue method. The pathological changes in renal tissues were observed by light microscopy and electron microscopy respectively. Heparanase mRNA was detected by RT-PCR. Expressions of desmin, CD2AP and heparanase were determined by immunohistological staining. The results showed that the expressions of heparanase mRNA and protein were increased in the glomeruli of AN rats at day 14 and 28 after the model establishment, which was accompanied by the increased expression of desmin and CD2AP. The mRNA and protein expression of heparanase was decreased in the sulodexide-treated rats as compared with AN rats at day 14 and 28. And, the protein expression of desmin and CD2AP was reduced as with heparanase in the sulodexide- treated rats. Proteinuria and podocyte foot process effacement were alleviated in the AN rats after sulodexide treatment. There was a positive correlation between the expression of heparanase and the expression of desmin and CD2AP (as well as 24-h urinary protein excretion). It was concluded that increased heparanase is involved in podocyte injury. Sulodexide can maintain and restore podocyte morphology by inhibiting the expression of heparanase in AN.

PurposeThis study aimed to investigate whether ursolic acid (UA) mitigates renal inflammation, oxidative stress and fibrosis by regulating the angiotensin II type 1 receptor-associated protein (ARAP1)/angiotensin II type 1 receptor (AT1R) signaling pathway and subsequently alleviating renal damage.Methodsdb/db mice were divided randomly into a diabetic nephropathy (DN) group and a UA treatment group. Light microscopy and electron microscopy were used to observe pathological changes in renal tissues. Immunohistochemistry (IHC) was employed to examine changes in the expression of ARAP1, AT1R, 8-hydroxydeoxyguanosine (8-OHdG), NADPH oxidase 2 (NOX2), the extracellular matrix protein fibronectin (FN), IL-1β and IL-18 in renal tissues. Western blotting and RT-qPCR were used to detect the respective changes in the protein and mRNA levels of ARAP1, AT1R, NOX4, NOX2, transforming growth factor-β1 (TGF-β1), FN, collagen IV, IL-1β and IL-18 in renal tissues and mesangial cells. In addition, immunofluorescence staining was employed to examine changes in FN and NOX2 expression in mesangial cells.ResultsUA treatment effectively reduced the body weights and blood glucose levels of db/db mice (p<0.05) as well as the urinary albumin/creatinine ratio (p<0.05). In addition, the renal tissue lesions and glomerulosclerosis index of the db/db mice were significantly improved after treatment (p<0.01). Histochemical analysis results showed significantly lower expression levels of ARAP1, AT1R, FN, NOX2, 8-OHdG, IL-1β and IL-18 in renal tissues in the UA treatment group than in the DN group. Western blotting and RT-qPCR data also revealed UA-induced decreases in the renal levels of the ARAP1, AT1, NOX4, NOX2, TGF-β1, FN, collagen IV, IL-1β and IL-18 proteins in vivo and/or in vitro (p<0.01). ARAP1 knockdown effectively reduced the expression of NOX2 and FN in vitro.ConclusionUA alleviated renal damage in type 2 diabetic db/db mice by downregulating proteins in the ARAP1/AT1R signaling pathway to inhibit extracellular matrix accumulation, renal inflammation, fibrosis and oxidative stress.