Nx Group Research Articles

Inhibition of α2-adrenoceptor is renoprotective in 5/6 nephrectomy-induced chronic kidney injury rats

In the present study, we investigated the renoprotective effects of long-term treatment with yohimbine, an α2-adrenoceptor inhibitor, in a 5/6 nephrectomy-induced chronic kidney disease (CKD) rat model. Male Sprague–Dawley rats were randomly allocated into the following groups: sham-operated, 5/6-nephrectomized (5/6 Nx), 5/6 Nx + low or high dose of yohimbine (0.3 or 3.0 mg/L in drinking water, respectively), and 5/6 Nx + hydralazine (250 mg/L in drinking water). The 5/6 Nx group presented with renal dysfunction, hypertension, noradrenaline overproduction, and histopathological injuries. Blood pressure decreased in both the yohimbine- and hydralazine-treated groups. Treatment with high dose of yohimbine, but not hydralazine, apparently attenuated urinary protein excretion and noradrenaline concentration of renal venous plasma. Renal fibrosis and upregulated fibrosis-related gene expression were suppressed by high dose of yohimbine. Furthermore, yohimbine, but not hydralazine, treatment ameliorated the urinary concentration ability. These findings suggest that long-term yohimbine treatment can be a useful therapeutic option to prevent the progression of CKD.

Clinical Features in Differentiated Thyroid Carcinoma Stratified By Lymph Node Status

Cervical lymph node (CLN) metastases (mets) often occur in differentiated thyroid cancer (DTC), especially in the central compartment, and are a major predictor of local recurrence. We examined clinical endpoints in three groups of patients based on status of lymph node involvement: those with definite lymph node involvement (N1), negative lymph nodes (N0), and no lymph nodes resected (Nx). We correlated these endpoints with clinical and pathologic features of these patients. Medical records of 261 patients with DTC who underwent thyroidectomy between 2006 and 2018 at our center were reviewed. Lymph node status of patients was categorized based on American Joint Committee on Cancer (AJCC) 8th edition criteria as N1, N0, and Nx. We performed statistical analysis to assess the differences among these groups, using one-way analysis of variance. When significant differences were found, pairwise comparisons were conducted among the three groups. Statistical significance was defined as 2-tailed P<.05 for all tests. There were significant differences among the groups in tumor multicentricity, tumor category/size, AJCC stage, and the presence of thyroglobulin auto-antibodies (TgAbs). There were no difference in age, gender, or histopathology. N1 patients had a higher incidence of multicentricity, larger tumor sizes, and were more likely to have elevated TgAbs. There were no significant differences between the N0 and Nx groups. This study shows that larger and multi-centric tumors are associated with increased likelihood of CLN mets in DTC. We suggest increased vigilance for CLN mets in tumors >2 cm, multicentric tumors, and patients with elevated TgAbs. AJCC = American Joint Committee on Cancer; CLN = cervical lymph node; DTC = differentiated thyroid cancer; FTC = follicular thyroid cancer; mets = metastases; N0 = no cancer in any lymph nodes; N1 = cancer in a lymph node; N1a = cancer in a central compartment lymph node; N1b = cancer in a lateral neck lymph node; Nx = lymph nodes not resected or examined; PTC = papillary thyroid cancer; TgAb = thyroglobulin auto-antibody.

The Association Between Lymph Node Stage and Clinical Prognosis in Thyroid Cancer.

Purpose: To investigate the association between lymph node (N) stage and clinical outcome in thyroid cancer patients with initial distant metastasis.Methods: A total of 3,198 cases (1,435 males and 1,763 females) between 2004 and 2015 with initial distant metastasis were obtained from the surveillance, epidemiology, and end results (SEER) database. Patients with a median follow up time of 13 months and a median age of 66 years were analyzed. A total of 1,407 cases had detailed information regarding the four most common metastatic organs after the year 2010. Kaplan-Meier (KM) analyses, log-rank tests, Cox regression, and logistic regression analyses were used.Results: Among the whole cohort, 33.4% (1,069/3,198), 14.5% (464/3,198), 10.1% (322/3,198), 34.2% (1,094/3,198), and 7.8% (249/3,198) of the patients were at the stage of N0, NX, N1a, N1b, and N1NOS (referring to metastasis to regional lymph nodes but not otherwise specified), respectively. The KM curves demonstrated that the patients at the NX stage had the worst survival. The NX and N1b groups had the highest hazard ratios (HRs) of 1.83 (95%CI 1.46-2.31) and 1.78 (95%CI 1.52-2.10) after adjusting age, race, gender, and tumor size (p < 0.001) compared with N0 group. The lung was the most common metastatic site, with a rate of 51.2% (720/1,407). Compared with the N0 group, N1 patients had higher odds (OR 1.63, 95%CI 1.31-2.01, p < 0.001) for lung metastasis. Similar results were obtained in papillary thyroid cancer (PTC) sub-cohort.Conclusions: Overall, the TC patients at the NX stage had the highest mortality risk, followed by N1b, N1a, and N0 groups. Compared with N0 patients, N1 patients were more likely to have lung metastasis. The poor prognosis for TC patients with the NX stage may make more aggressive treatment reasonable.

Inflammation both increases and causes resistance to FGF23 in normal and uremic rats.

Fibroblast growth factor 23 (FGF23) increases phosphorus excretion and decreases calcitriol (1,25(OH)2D) levels. FGF23 increases from early stages of renal failure. We evaluated whether strict control of phosphorus intake in renal failure prevents the increase in FGF23 and to what extent inflammation impairs regulation of FGF23. The study was performed in 5/6 nephrectomized (Nx) Wistar rats fed diets containing 0.2-1.2% phosphorus for 3 or 15 days. FGF23 levels significantly increased in all Nx groups in the short-term (3-day) experiment. However, at 15 days, FGF23 increased in all Nx rats except in those fed 0.2% phosphorus. In a second experiment, Nx rats fed low phosphorus diets (0.2 and 0.4%) for 15 days received daily intraperitoneal lipopolysaccharide (LPS) injections to induce inflammation. In these rats, FGF23 increased despite the low phosphorus diets. Thus, higher FGF23 levels were needed to maintain phosphaturia and normal serum phosphorus values. Renal Klotho expression was preserved in Nx rats on a 0.2% phosphorus diet, reduced on a 0.4% phosphorus diet, and markedly reduced in Nx rats receiving LPS. In ex vivo experiments, high phosphorus and LPS increased nuclear β-catenin and p65-NFκB and decreased Klotho. Inhibition of inflammation and Wnt signaling activation resulted in decreased FGF23 levels and increased renal Klotho. In conclusion, strict control of phosphorus intake prevented the increase in FGF23 in renal failure, whereas inflammation independently increased FGF23 values. Decreased Klotho may explain the renal resistance to FGF23 in inflammation. These effects are likely mediated by the activation of NFkB and Wnt/β-catenin signaling.

Combined Treatment with Omega-3 Fatty Acid and Cholecalciferol Increases 1,25-Dihydroxyvitamin D Levels by Modulating Dysregulation of Vitamin D Metabolism in 5/6 Nephrectomy Rats.

The protein 1α-hydroxylase (CYP27B1) was expressed in liver and omega-3 fatty acid (FA) elevated 1,25-dihydroxyvitamin D [1,25(OH)2D] levels in dialysis patients. The aim of this study was to determine whether omega-3 FA and cholecalciferol have effects on vitamin D metabolism related to CYP27B1 and 24-hydroxylase (CYP24) activities in the kidney and liver of 5/6 nephrectomy (Nx) rats. Male Sprague–Dawley rats were divided into the following groups: sham control, 5/6 Nx, 5/6 Nx treated with cholecalciferol, 5/6 Nx treated with omega-3 FA, and 5/6 Nx treated with cholecalciferol/omega-3 FA. CYP27B1 and CYP24 expression were measured in the liver and kidney. Further, 1,25(OH)2D and 25-hydroxyvitamin D [25(OH)D] levels were measured in serum. Among Nx groups, 1,25(OH)2D and 25(OH)D levels were lowest in the 5/6 Nx group. CYP24 expression was increased in the kidney of the 5/6 Nx rat model, which was found to be reversed by omega-3 FA or cholecalciferol/omega-3 FA supplementation. Decreased CYP27B1 expression was observed in the liver of the 5/6 Nx rats and its expression was recovered by supplementation with cholecalciferol/omega-3 FA. In conclusion, omega-3 FA and cholecalciferol may synergistically increase 1,25(OH)2D levels by inhibiting CYP24 expression in the kidney and liver and activating CYP27B1 expression in the liver of 5/6 Nx rats.

Color Change of Different Dual-Cure Resin Cements After Thermocycling

The purpose of this study is to evaluate the effect of thermocycling on the color change of the amine-free dual-cure resin cements. IPS e.max CAD blocs were cut into specimens of 1 mm thickness (N=28) and cemented with one of the 4 different amine-free dual-cure resin cements (NX3 Nexus [NX], Kerr Dental; Variolink Esthetic DC [VE], Ivoclar Vivadent; Panavia V5 [PV], Kuraray Dental; G-CEM Linkforce [GC], GC Corporation) (n=7). A spectrophotometer was used for color measurements. Specimens were subjected to thermocycling (5°C and 55°C; 5000 and 10000 cycles). Normality of data distribution was tested by using the Kolmogorov-Smirnov test. Statistical analysis was performed using a two-way analysis of variance (ANOVA) and Tukey’s multiple comparison tests at a significance level of p&lt;0.05. ∆E values were significantly influenced by the resin cements and the cycle periods (p&lt;0.05). There were no significant differences between NX and VE groups after 5000 thermocycling, however after 10000 thermocycling VE group showed higher ∆E1 values than NX group (p&gt;0.05). There were no statistically significant differences between the ∆E0 and ∆E1 values of the GC group, however the other groups were affected after 10000 thermocycling (p&gt;0.05). Amine-free resin cements used for cementation showed color change after thermocycling except GC group. All resin cements were showed clinically acceptable color change after thermocycling (∆E &lt; 3.5).

Prognostic Value of Lymph Node Status and Lymphadenectomy in Pancreatic Neuroendocrine Tumors

Introduction: Lymph node metastasis (LNM) is common in patients with pancreatic neuroendocrine tumors (pNETs) but the prognostic value and optimal management remains controversial. The objective of the current study was to investigate factors associated with LNM and the prognostic value of LNM and lymphadenectomy in pNETs. Method: A total of 1067 patients who underwent resection for pNETs between 1997 and 2016 were identified from the US Neuroendocrine Tumor Study Group. Clinicopathologic data were reviewed and survival analysis was performed to identify the prognostic factors. Result: There were 549 (51.5%) male, and 518 (48.5%) females, with a median age of 58 years (IQR 47-65 years). There were 525 (49.2%) patients with WHO G1 tumor, 257 (24.1%) with G2 and 32 (3.0%) with G3. Lymphadenectomy was performed in 891 (83.5%) patients, and 256 (24.0%) cases were confirmed to be positive. Multivariable analysis revealed that LNM was associated with head/uncinate location (p=0.021), largest tomur size>2cm (p=0.004), increasing tumor grade (p=0.021), lymph vascular invasion (p<0.001) and distant metastasis (p=0.001). With a median follow up of 34.5 months, 144 (13.5%) patients recurred and 124 (11.6%) patients died of disease progression. The median OS of N1 group was 127.2 moths, which was significantly shorter than N0/Nx group (176.4 months, p<0.001). For patients underwent curative resection (R0/R1, n=1012), the median RFS in N0/Nx group was also superior to N1 group (not reached vs. 91.1 months, p<0.001). In addition, number of positive lymph node>5 was associated with worse OS (p=0.013). Lymphadenectomy was mostly performed in distal pancreatectomy (59.9%), tumor size >2cm (89.7%), and grade G3 (93.8%). The median OS in Nx group (190.9 months) was similar to N0 group (176.4 months, p=0.265). Conclusion: LNM are valuable predictor of poor outcomes for pNETs. Surgery without LA is safe in selected patients with small G1 lesions, normal sized and soft lymph nodes.

Raman Spectroscopic Analysis to Detect Reduced Bone Quality after Sciatic Neurectomy in Mice.

Bone mineral density (BMD) is a commonly used diagnostic indicator for bone fracture risk in osteoporosis. Along with low BMD, bone fragility accounts for reduced bone quality in addition to low BMD, but there is no diagnostic method to directly assess the bone quality. In this study, we investigated changes in bone quality using the Raman spectroscopic technique. Sciatic neurectomy (NX) was performed in male C57/BL6J mice (NX group) as a model of disuse osteoporosis, and sham surgery was used as an experimental control (Sham group). Eight months after surgery, we acquired Raman spectral data from the anterior cortical surface of the proximal tibia. We also performed a BMD measurement and micro-CT measurement to investigate the pathogenesis of osteoporosis. Quantitative analysis based on the Raman peak intensities showed that the carbonate/phosphate ratio and the mineral/matrix ratio were significantly higher in the NX group than in the Sham group. There was direct evidence of alterations in the mineral content associated with mechanical properties of bone. To fully understand the spectral changes, we performed principal component analysis of the spectral dataset, focusing on the matrix content. In conclusion, Raman spectroscopy provides reliable information on chemical changes in both mineral and matrix contents, and it also identifies possible mechanisms of disuse osteoporosis.

Decreased expression of transforming growth factor-β1 and α-smooth muscle actin contributes to the protection of lotensin against chronic renal failure in rats

Background: Lotensin has been shown to have a protective function in the early stage of chronic renal failure. However, its role in the intermediate and late stages of chronic renal failure remains largely unknown. The present study aimed to investigate the role and underlying mechanism of lotensin in advanced chronic kidney disease.Methods: Female Wistar rats were randomly divided into three groups (n = 10): sham group, 5/6 nephrectomy (5/6 Nx) group, and lotensin group (oral administration of lotensin for 9 weeks following 5/6 Nx). Rats were sacrificed and pathological parameters were measured. Western blot assay and immunohistochemical staining were performed to detect the expression of transforming growth factor-β1 (TGF-β1) and α-smooth muscle actin (α-SMA) in kidney tissues.Results: Compared to the 5/6 Nx group, lotensin administration significantly decreased 5/6 Nx-induced elevation in blood urea nitrogen, serum creatinine and 24-h urinary protein excretion (UPE) rates, but markedly increased red blood cell count, plasma albumin and hemoglobin levels, along with improved renal morphology. Mechanistically, lotensin dramatically downregulated the renal expression of TGF-β1 and α-SMA induced by 5/6 Nx.Conclusions: Lotensin protects against advanced chronic kidney disease in rats with 5/6 Nx through the downregulation of TGF-β1 and α-SMA.

Sodium butyrate ameliorates insulin resistance and renal failure in CKD rats by modulating intestinal permeability and mucin expression.

BackgroundThe associated increase in the lipopolysaccharide (LPS) levels and uremic toxins in chronic kidney disease (CKD) has shifted the way we focus on intestinal microbiota. This study shows that a disruption of the intestinal barrier in CKD promotes leakage of LPS from the gut, subsequently decreasing insulin sensitivity. Butyrate treatment improved the intestinal barrier function by increasing colonic mucin and tight junction (TJ) proteins. This modulation further ameliorated metabolic functions such as insulin intolerance and improved renal function.MethodsRenal failure was induced by 5/6th nephrectomy (Nx) in rats. A group of Nx and control rats received sodium butyrate in drinking water. The Nx groups were compared with sham-operated controls.ResultsThe Nx rats had significant increases in serum creatinine, urea and proteinuria. These animals had impaired glucose and insulin tolerance and increased gluconeogenesis, which corresponded with decreased glucagon-like peptide-1 (GLP-1) secretion. The Nx animals suffered significant loss of intestinal TJ proteins, colonic mucin and mucin 2 protein. This was associated with a significant increase in circulating LPS, suggesting a leaky gut phenomenon. 5′adenosine monophosphate-activated protein kinase (AMPK) phosphorylation, known to modulate epithelial TJs and glucose metabolism, was significantly reduced in the intestine of the Nx group. Anti-inflammatory cytokine, interleukin 10, anti-bacterial peptide and cathelicidin-related antimicrobial peptide were also lowered in the Nx cohort. Butyrate treatment increased AMPK phosphorylation, improved renal function and controlled hyperglycemia.ConclusionsButyrate improves AMPK phosphorylation, increases GLP-1 secretion and promotes colonic mucin and TJ proteins, which strengthen the gut wall. This decreases LPS leakage and inflammation. Taken together, butyrate improves metabolic parameters such as insulin resistance and markers of renal failure in CKD animals.

Dietary Changes Involving Bifidobacterium longum and Other Nutrients Delays Chronic Kidney Disease Progression

Background: Recent studies suggest that prebiotic and/or probiotic treatments ameliorate kidney function in humans and animals by improving the gut environment. However, the gut microbiota and kidney disease interactions remain to be determined. This study investigated whether synbiotics modulate the gut microbiota and ameliorate kidney function using a rat model of chronic kidney disease (CKD). As uremic toxins are associated with CKD-related mineral and bone disorder, the secondary aim was to evaluate the relationship between synbiotics and secondary hyperparathyroidism (SHPT). Methods: 5/6 nephrectomy (Nx) rats were developed as the CKD model. Sham-operated (sham) rats were used as the control. To investigate the effectiveness of prebiotics (glutamine, dietary fiber, and oligosaccharide) and probiotics (Bifidobacterium longum strain; GFOB diet), rats were randomly assigned to 4 groups: Nx group fed the GFOB diet (n = 10); Nx group fed the control (CON) diet (n = 10); sham group fed the GFOB diet (n = 5); and sham group fed the control diet (n = 5). Blood, feces, and kidney samples were collected and analyzed. Results: Serum creatinine (Cre) and blood urea nitrogen in the Nx GFOB group were significantly lower than those in the Nx CON group. Serum indoxyl sulfate in the Nx GFOB group was lower than that in the Nx CON group, and significantly correlated with serum Cre. Inorganic phosphorus and intact parathyroid hormone in the Nx GFOB group were significantly lower than those in the Nx CON group. Conclusion: Improving the gut environment using synbiotics ameliorated kidney function and might be a pharmacological treatment for SHPT without any serious adverse events.

Men are from Mars, Women are from Venus. Gender specific changes in pulmonary function in 1 year old rats after neonatal hyperoxia exposure

RationalePreterm birth affects about 11% of pregnancies worldwide. The Institute of Medicine estimated the cost associated with premature birth to be $36 billion in 2016. Lungs in preterm infants are in the critical saccular stage of lung development and susceptible to the oxygen‐induced chronic lung disease known as bronchopulmonary dysplasia (BPD). BPD is characterized by functional abnormalities including decreased saccular/alveolar septation, decreased microvascular cross‐sectional area with or without pulmonary hypertension, airway injury with or without increased airway reactivity, and abnormalities of control of breathing in various combinations. While the initial inflammation improves, chronic inflammation ensues for a long time. We tested how neonatal hyperoxia exposure affects lung function one year after birth. We postulated an increase in lung collagen levels reflected by reduced pulmonary function.Methods6 newborn male and female rats in each group were exposed to Hyperoxia (Hx; 85% O2) or Normoxia (Nx; 21% O2) for 14 days, then aged to 1 year in Nx. At one year, lung function was measured using a FlexiVent (Scireq, Montreal, Canada). Using a single frequency forced oscillation manoeuvre and a low‐frequency forced oscillation technique the lung was tested for compliance and resistance. Total lung collagen was determined by a colorimetric assay using Ehrlich's reagent.ResultsAnalysis of one year old rats showed no difference in weight and body length between the Nx and the Hx group within each gender. The inspiratory capacity (IC) was significantly higher in the Hx exposed male rats compared to the Nx exposed male rats (12.6±0.6 ml vs 14.0±0.9 ml; p=0.02). The difference in IC in female rats was significantly lower in the Hx group compared to the Nx group (11.3±0.4 vs 10.1±0.6 ml; p=0.002). Dynamic compliance was significantly increased in the male Hx group compared to the male Nx group (1.43±0.05 vs 1.61±0.11 ml/cm H2O; p=0.04). In contrast, the female Hx group showed significantly reduced compliance compared to the female Nx group (1.32±0.05 vs 1.15±0.17 ml/cm H2O). In addition, the PV loop area was higher in the male Hx group compared to the male Nx group and no difference was found between the two female groups. Total collagen in lung tissue was not different between the male Hx and Nx groups but was higher in the female Hx compared to the Nx group.ConclusionOne year after neonatal Hx exposure, the male lung shows increased compliance and inspiratory capacity compared to the Nx exposed rats suggesting a trend to emphysema. In contrast, female rats show signs of lung fibrosis reflected by reduced compliance and increased total collagen content. This data suggests that neonatal oxygen exposure initiates alterations in inflammatory and repair pathways that are different between male and female rats.Support or Funding InformationNHLBI: R01 HL115061‐01 (Eldridge)This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Long-term exercise restores hydrogen sulfide in the kidney and contributes to exercise benefits in 5/6 nephrectomized rats

ABSTRACTPhysical exercise is shown to have protective effects on chronic kidney disease (CKD). CKD itself is associated with a reduction in renal hydrogen sulfide (H2S) concentration. This study was designed to investigate whether protective effects of exercise in 5/6 nephrectomized (5/6 NX) rats is associated with H2S levels in the kidney? Twenty four male Wistar rats weighing 250–300 g were assigned into 4 groups: 1- Sham 2- Sham exercise 3–5/6 NX 4–5/6 NX+exercise. To induce CKD, 4 days after removing upper and lower one-third parts of the left kidney, total right nephrectomy was performed. In the Sham groups, anesthesia and surgery were performed like the other groups without removal of the kidney mass. Exercise was performed by treadmill at a speed of 18 m/min for 8 weeks. At the end of the twelfth week, blood and kidney samples were collected to measure renal function (levels of plasma urea and creatinine), oxidative stress markers (renal MDA level and SOD activity), and histological indices. Eight weeks exercise significantly improved serum creatinine, BUN, renal MDA level, SOD activity, renal sympathetic nerve activity (RSNA), hypertension, and renal histology in addition to renal H2S level compared to the 5/6 NX group. The results suggest that regular exercise improves renal oxidative status and ameliorates renal damage, hypertension, and RSNA in 5/6 nephrectomized rats. These improvements by exercise might be associated with the increase in renal H2S level.

Prognostic significance of nephrectomy in metastatic renal cell carcinoma treated with systemic cytokine or targeted therapy: A 16-year retrospective analysis

We compared progression-free survival (PFS) and overall survival (OS) among 292 metastatic renal cell carcinoma (mRCC) patients either undergoing nephrectomy (Nx, 61.6%) or not (non-Nx, 38.4%), stratified according to the MSKCC and Heng risk models, treated with either immunotherapy (IT, 45.2%) or targeted therapy (TT, 54.8%) between 2000 and 2015. During the follow-up duration of 16.6 months, PFS/OS of the Nx (6.0/30 months) and non-Nx (3.0/6.0 months) groups were significantly different despite differences among baseline parameters (p < 0.05). The intermediate- and poor-risk patients defined using either model showed significantly longer PFS and OS in the Nx group than in the non-Nx group (p < 0.05). After stratifying groups by systemic therapy and risk models, both the Nx and non-Nx groups showed no significant differences in intermediate and poor-risk models (p > 0.05). In both synchronous and metachronous mRCC patients, both PFS and OS showed similar survivals; the Nx group had significantly longer PFS and OS than the non-Nx group, even after considering each systemic therapy and prognostic model. Nx showed a significant positive benefit in PFS and OS compared to no Nx upon patient stratification according to the MSKCC and Heng risk models. The metastatic type did not significantly affect survival between the two groups.

High calcium diet alleviates 5/6 nephrectomy-induced bone deteriorations of lumbar vertebrae in mice.

Dietary calcium (Ca) supplementation has beneficial effects on bone health. However, it is not clear whether a high calcium diet (HCD) following 5/6 nephrectomy (5/6 Nx) is beneficial to bone health. The aim of the present study was to examine the effects of an HCD on bone metabolism using a chronic kidney disease (CKD) mouse model. Male C57BL/6J mice were divided into three groups: Sham group, 5/6 Nx group and 5/6 Nx + HCD group. Mice were sacrificed 12 weeks post-surgery. Calcium (Ca) and creatinine (Cr) were measured using standard colorimetric methods and picric acid methods, respectively. Bone metabolism-associated markers, FGF-23, PTH, ALP-b and TRAP-5b were measured using ELISA kits. Lumbar vertebrae histomorphological analysis was performed using hematoxylin and eosin staining. The expression of osteoprotegerin (OPG) and receptor activator of nuclear factor κB ligand (RANKL) mRNA was detected using reverse transcription-quantitative polymerase chain reaction. Impaired renal function and histopathological damage was indicated in 5/6 Nx mice. However, HCD had no significant effects on these changes in 5/6 Nx mice. Notably, mineral metabolism disorder and histopathological damage to lumbar vertebrae were markedly improved in HCD-treated 5/6 Nx mice. Compared with 5/6 Nx mice, HCD supplementation significantly elevated the ratio of OPG/RANKL and inhibited RANKL mRNA expression in lumbar vertebrae. To conclude, the present findings indicated that increased Ca intake is effective in increasing bone mineral content of the lumbar vertebrae in 5/6 Nx mice. These results may provide a basis for the clinical use of dietary Ca supplementation as a therapeutic approach to treat CKD-induced disturbance of mineral metabolism and bone loss.

Triiodothyronine attenuates the progression of renal injury in a rat model of chronic kidney disease.

This study was designed to investigate whether and how triiodothyronine (T3) affects renal function in an experimental model of chronic kidney disease. Twenty-four female rats were divided into the following groups: sham-operated control group (n = 8), 5/6 nephrectomized group (Nx, n = 8), and 5/6 nephrectomized group treated with T3 for 2 weeks (T3-Nx, n = 8). T3 administration significantly decreased serum levels of urea, creatinine, tumour necrosis factorα, and interleukin-6 compared with serum levels in the Nx group. The levels of malondialdehyde, transforming growth factor β, fibronectin, and collagen IV, as well as the expression of inducible nitric oxide synthase, nuclear factor κB, poly(ADP-ribose) polymerase, caspase-3, and Bax were all significantly decreased, though not normalized, in the remnant kidney of rats in the T3-Nx group compared with Nx rats. Glutathione, heme oxygenase-1 levels, as well as endothelial nitric oxide synthase expression were increased in the remnant kidney of the T3-Nx group. Histological studies revealed focal necrosis of renal tubules associated with inflammatory cell infiltration and fibrosis in the Nx group. These changes were alleviated in T3-Nx rats. This study showed that T3 administration attenuated the clinical and histological signs of renal injury in 5/6 nephrectomized rats by mitigating renal oxidative stress, inflammation, apoptosis, and fibrosis.

Jian‐Pi‐Yi‐Shen Formula Regulates Inflammatory Cytokines Production in 5/6 Nephrectomized Rats via Suppression of NF‐κB Activation

Jian-Pi-Yi-Shen formula (JPYSF) is a Chinese herbal decoction used for treating chronic kidney disease (CKD) for over 20 years with good efficiency. However, the mechanism lacks solid evidence. In the present study, we tested the hypothesis that JPYSF may retard CKD progression via inhibition of inflammation in 5/6 nephrectomy (5/6 Nx) rat model. The 5/6 Nx rats were randomly divided into 2 groups: 5/6 Nx group and JPYSF group. Sham-operated rats served as control. JPYSF (2.06 g/kg/d) were administrated by gavage to 5/6 Nx rats daily for 6 weeks. Results showed that JPYSF treatment significantly improved kidney function and pathological injury in 5/6 Nx rats. Multiplex analysis of cytokines revealed that JPYSF reduced proinflammatory cytokines and increased anti-inflammatory cytokine production. Furthermore, JPYSF inhibited the activation of nuclear factor-kappa B (NF-κB) signaling pathway. In conclusion, our data demonstrated that JPYSF remarkably retards development and progression of CKD in a 5/6 Nx rat model, which may be associated with inhibition of inflammation via NF-κB signaling pathway.

Disorder of gut amino acids metabolism during CKD progression is related with gut microbiota dysbiosis and metagenome change

Chronic kidney disease (CKD) is a worldwide public health problem. Uremic retention solutes such as indoxyl sulfate (IS) and p-cresyl sulfate (PCS) are accumulated in CKD patients and are associated with the incidence of CKD progression. Amino acids are the major precursors of uremic retention solutes in gut. The dynamic change of amino acid metabolism in the gut during CKD progression has not been reported previously. In this paper, we studied the dynamic change of gut IS/PCS precursor and amino acid metabolism profile during CKD progression in 5/6 nephrectomized (5/6Nx) rats model. The related gut microbiota and metagenome profile was also studied. Rat plasma, urine and feces were collected at different time points after nephrectomization. Plasma IS and PCS, fecal indole (the precursor of IS), p-cresol (the precursor of PCS) and 19 kinds of amino acids were analyzed by LC–MS. During CKD progression, 5/6 Nx rats showed increased plasma IS, PCS concentration and increased fecal indole, p-cresol concentration. 5/6 Nx rats also showed disordered gut amino acids metabolism profile which became more significant along with the progession of CKD. The abundance of some specific gut bacteria also changed significantly in 5/6 Nx rats. The 16S rDNA sequencing data of gut microbiota was further analyzed by an online tool PICRUSt, a large-scale computational method for metagenomes prediction with 16S rDNA sequencing data. The content of each gene was compared between the two groups by Mann-Whitney U test, and then the significantly regulated genes in 5/6 Nx group were subjected to KEGG website. The amino acid metabolism related genes were picked out. Most of these genes are more abundant in 5/6 Nx groups. Our study showed that gut amino acids metabolism profile was disordered with CKD progression, which was highly related to the gut microbiota dysbiosis and metagenome change. And that regulation of gut amino acids metabolism pathway may be a possible way to intervene the progression of CKD.

Effect of Intranasally Delivered rh-VEGF165 on Angiogenesis Following Cerebral Hypoxia-Ischemia in the Cerebral Cortex of Newborn Piglets.

Background: Vascular endothelial growth factor (VEGF) stimulates vascular genesis and angiogenesis. Cerebral Hypoxia-Ischemia (HI) leads to the reduction of vasculature in the cerebral cortex of newborn piglets. Objective: The present study tests the hypothesis that post-hypoxia intranasal administration of recombinant human VEGF165 (rh-VEGF165) for 3 days increases the vascular density in the cerebral cortex of newborn piglets without promoting neovascularization. Design/Methods: Ventilated newborn piglets were divided into three groups (n = 5/group): normoxic (Nx), hypoxic-ischemic (HI), and HI treated with intranasal rh-VEGF165rh-VEGF165 (HI-VEGF). HI piglets were exposed to HI (0.05 FiO2) for 30 min. Recombinant h-VEGF165 (100 ng/kg) was administered 15 min after HI and then once daily for 3 days. The animals were perfused transcardially and coronal brains sections were processed for Isolectin, Hoechst, and ki-67 cell proliferation marker staining. To assess the vascular density, 30–35 fields per animal section were manually counted using image J software. Results: The vascular density (vessels/mm2) was 42.0 ± 8.0 in the Nx group, 26.4 ± 4.8 (p < 0.05 vs. Nx) in the HI group, and 46.0 ± 11.9 (p < 0.05 vs. HI) in the HI-VEGF group. When stained for newly formed vessels, via Ki-67 staining, the vascular density was 5.4 ± 3.6 in the Nx group (p < 0.05 vs. HI), 10.2 ± 2.1 in the HI group, and 10.9 ± 2.9 in the HI-VEGF group (p = 0.72 vs. HI). HI resulted in a decrease in vascular density. Intranasal rh-VEGF165rh-VEGF165 resulted in the attenuation of the HI-induced decrease in vascular density. However, rh-VEGF165 did not result in the formation of new vascularity, as evident by ki-67 staining. Conclusions: Intranasal rh-VEGF165 may prevent the HI-induced decrease in the vascular density of the brain and could serve as a promising adjuvant therapy for hypoxic-ischemic encephalopathy (HIE).

Effects of Tomatine on Aldosterone Release from Zona Glomerulosa Cells in 5/6 Nephrectomized Male Rats

Chronic kidney disease (CKD) is a common risk. Tomatine (Tt) is a glycoalkaloid extracted from tomato plants. To study the CKD, the 5/6 nephrectomy (5/6 NX) in rats was performed. The purpose of this study was to clarify the actions of Tt on the function of zona glomerulosa (ZG) cells. Rat ZG cells were prepared and treated with Tt in the presence or absence of angiotensin II (Ang II). The release of aldosterone was dose-dependently decreased by the administration of Tt. Then, male rats were divided into six groups including [1] sham+ oil, [2] sham+Tt (0.4 mg/kg), [3] sham+Tt (2 mg/kg), [4] 5/6 NX+oil, [5] 5/6 NX +Tt (0.4 mg/kg), and [6] 5/6 NX +Tt (2 mg/kg). Six weeks after 5/6 NX and sham-operation, Tt and oil were injected subcutaneously once per day for 7 days. In nephropathy rats, both blood creatinine and urine nitrogen increased as the markers of renal dysfunction. With metabolic cage housing, the water consumption and urine output declined after treatment of Tt in rats. After injection of Tt, the mean arterial pressure was slightly declined. The circulating levels of Na^+, K^+, and Cl^- were not altered but the osmolality was significantly raised in 5/6 NX rats. The plasma and medium levels of aldosterone were measured by radioimmunoassay. The levels of plasma aldosterone in intact groups were not altered, but markedly raised in 5/6 NX groups after Tt treatment. Immunoblotting was performed to analyze the protein expressions of angiotensin II Type 1 receptor (AT1R), steroidogenic acute regulatory protein (StAR), and cholesterol side-chain cleavage (P450scc) enzyme in rat adrenocortex tissue. These results indicated that the expression of AT1R was up-regulated in 5/6 NX rats but decreased by the administration of Tt. These results suggested that Tt may act directly on ZG cells to reduce the release of aldosterone in response to angiotensin II via attenuating the cyclic adenosine monophosphate (cAMP) pathway. The reduction of Tt on aldosterone release in CKD rats induced by 5/6 NX were at least in part due to the inhibition of Tt on the elevated protein expression of AT1R in CKD rats.