Retinal inflammation in murine models of type 1 and type 2 diabetes with diabetic retinopathy.

AIMS/HYPOTHESIS: The loss of pericytes surrounding the retinal vasculature in early diabetic retinopathy underlies changes to the neurovascular unit that lead to more destructive forms of the disease. However, it is unclear which changes lead to loss of retinal pericytes. This study investigated the hypothesis that chronic increases in one or more inflammatory factors mitigate the signalling pathways needed for pericyte survival. METHODS: Loss of pericytes and levels of inflammatory markers at the mRNA and protein levels were investigated in two genetic models of diabetes, Ins2Akita/+ (a model of type 1 diabetes) and Leprdb/db (a model of type 2 diabetes), at early stages of diabetic retinopathy. In addition, changes that accompany gliosis and the retinal vasculature were determined. Finally, changes in retinal pericytes chronically incubated with vehicle or increasing amounts of IFNγ were investigated to determine the effects on pericyte survival. The numbers of pericytes, microglia, astrocytes and endothelial cells in retinal flatmounts were determined by immunofluorescence. Protein and mRNA levels of inflammatory factors were determined using multiplex ELISAs and quantitative reverse transcription PCR (qRT-PCR). The effects of IFNγ on the murine retinal pericyte survival-related platelet-derived growth factor receptor ß (PDGFRß) signalling pathway were investigated by western blot analysis. Finally, the levels of cell death-associated protein kinase C isoform delta (PKCδ) and cleaved caspase 3 (CC3) in pericytes were determined by western blot analysis and immunocytochemistry. RESULTS: The essential findings of this study were that both type 1 and 2 diabetes were accompanied by a similar progression of retinal pericyte loss, as well as gliosis. However, inflammatory factor expression was dissimilar in the two models of diabetes, with peak expression occurring at different ages for each model. Retinal vascular changes were more severe in the type 2 diabetes model. Chronic incubation of murine retinal pericytes with IFNγ decreased PDGFRß signalling and increased the levels of active PKCδ and CC3. CONCLUSIONS/INTERPRETATION: We conclude that retinal inflammation is involved in and sustains pericyte loss as diabetic retinopathy progresses. Moreover, IFNγ plays a critical role in reducing pericyte survival in the retina by reducing activation of the PDGFRß signalling pathway and increasing PKCδ levels and pericyte apoptosis.

In Vivo Mechanism of Action of Sodium Caprate for Improving the Intestinal Absorption of a GLP1/GIP Coagonist Peptide.

Sodium caprate (C10) has been widely evaluated as an intestinal permeation enhancer for the oral delivery of macromolecules. However, the effect of C10 on the intestinal absorption of peptides with different physicochemical properties and its permeation-enhancing effect in vivo remains to be understood. Here, we evaluated the effects of C10 on intestinal absorption in rats with a glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GIP-GLP1) dual agonist peptide (LY) and semaglutide with different enzymatic stabilities and self-association behaviors as well as the oral exposure of the LY peptide in minipigs. Furthermore, we investigated the mechanism of action (MoA) of C10 for improving the intestinal absorption of the LY peptide in vivo via live imaging of the rat intestinal epithelium and tissue distribution of the LY peptide in minipigs. The LY peptide showed higher proteolytic stability in pancreatin and was a monomer in solution compared to that in semaglutide. C10 increased in vitro permeability in the minipig intestinal organoid monolayer to a greater extent for the LY peptide than for semaglutide. In the rat jejunal closed-loop model, C10 increased the absorption of LY peptide better than that of semaglutide, which might be attributed to higher in vitro proteolytic stability and permeability of the LY peptide. Using confocal live imaging, we observed that C10 enabled the rapid oral absorption of a model macromolecule (FD4) in the rat intestine. In the duodenum tissues of minipigs, C10 was found to qualitatively reduce the tight junction protein level and allow peptide uptake to the intestinal cells. C10 decreased the transition temperature of the artificial lipid membrane, indicating an increase in membrane fluidity, which is consistent with the above in vivo imaging results. These data indicated that the LY's favorable physicochemical properties combined with the effects of C10 on the intestinal mucosa resulted in an ∼2% relative bioavailability in minipigs.

Major Fatty Acid Profiles and Bioactivity of Seed Oils from Ten†Tree Peony Cultivars as a Potential Raw Material Source for the Cosmetics and Healthy Products.

As a new woody oleaginous crop, tree peony is now being widely developed and utilized, which attributed to the outstanding oil-use features regarding the fatty acid profiles and bioactivity. The major fatty acid profiles and bioactivity of seed oils in ten tree peony varieties collected from a common garden were investigated in the present study. The results showed that the oil yields, fatty acid profiles, bioactivity of seed oils existed significant variations among ten tree peony varieties (P<0.05). Considered the application value, 'Fengdan' (FD) and 'Ziban' (ZB) was the optimal resources as the high oil yields, rich unsaturated fatty acids especially high α-linolenic acid of 40.46 %, great antioxidant activity with low IC50 values, high ABTS and FRAP values, and strong antimicrobial activity with high DIZ and low MIC/MBC values. The study also confirmed seed oil of tree peony as the potential raw materials sources in functional food, pharmaceuticals and cosmetics for human health.

Comparative Analysis of the Chemical Consistency Between the Traditional and Mixed Decoction of Maimendong Decoction by Ultra-Performance Liquid Chromatography Coupled to Quadrupole with Time-of-Flight Mass Spectrometry (UPLC-QTOF-MS)-Based Chemical Profiling Approach.

Take Maimendong Decoction (MMDD), one of the Chinese classic herbal formulas, as an object to evaluate the chemical consistency between traditional decoction and mixed decoction. The ultra-performance liquid chromatography coupled to quadrupole with time-of-flight mass spectrometry-based chemical profiling approach has been utilized. A total of 48 major peaks are detected from these two decoctions under the present chromatographic and mass spectrometry conditions. The results of negative ion mode show nine significant inconsistencies. Liquiritin, ginsenoside Ro and ginsenoside Rg5/Rk1 are detected with higher intensity in traditional preparation sample than the mixed decoction, while licoisoflavone A is higher in mixed decoction samples than the traditional one. The mechanisms involved in the chemical changes were assumed to be anti-inflammation, anti-oxidative stress and so on, suggesting these two different preparation approaches of MMDD may lead to a possibility of discrepant clinical outcomes.

Synopsis of Sweet! Mouse Models of Diabetic Kidney Disease.

Diabetes mellitus (types 1 and 2) is the leading cause of glomerular disease and end-stage renal disease in most developed countries, with estimates that one-third of people living with diabetes will develop diabetic kidney disease (DKD). The current standard of care medications slow but do not arrest progression of kidney disease, and therefore, therapy for DKD is a highly unmet medical need for patients. To discover and test novel and durable new therapies, it is necessary to develop animal models of human DKD, which authentically recapitulate the human disease state and provide translatable efficacy to human patients. Here, we review selected mouse models of human DKD, which demonstrate many of the features of type 2 human DKD.

Progressive Renal Disease Established by Renin-Coding Adeno-Associated Virus-Driven Hypertension in Diverse Diabetic Models.

Progress in research and developing therapeutics to prevent diabetic kidney disease (DKD) is limited by a lack of animal models exhibiting progressive kidney disease. Chronic hypertension, a driving factor of disease progression in human patients, is lacking in most available models of diabetes. We hypothesized that superimposition of hypertension on diabetic mouse models would accelerate DKD. To test this possibility, we induced persistent hypertension in three mouse models of type 1 diabetes and two models of type 2 diabetes by adeno-associated virus delivery of renin (ReninAAV). Compared with LacZAAV-treated counterparts, ReninAAV-treated type 1 diabetic Akita/129 mice exhibited a substantial increase in albumin-to-creatinine ratio (ACR) and serum creatinine level and more severe renal lesions. In type 2 models of diabetes (C57BKLS db/db and BTBR ob/ob mice), compared with LacZAAV, ReninAAV induced significant elevations in ACR and increased the incidence and severity of histopathologic findings, with increased serum creatinine detected only in the ReninAAV-treated db/db mice. The uninephrectomized ReninAAV db/db model was the most progressive model examined and further characterized. In this model, separate treatment of hyperglycemia with rosiglitazone or hypertension with lisinopril partially reduced ACR, consistent with independent contributions of these disorders to renal disease. Microarray analysis and comparison with human DKD showed common pathways affected in human disease and this model. These results identify novel models of progressive DKD that provide researchers with a facile and reliable method to study disease pathogenesis and support the development of therapeutics.

Minimum absolute lymphocyte counts during radiation are associated with a worse prognosis in patients with unresectable hepatocellular carcinoma.

BACKGROUND: Peripheral blood lymphocytes play an important role in antitumour immunity. We examined the relationship between the minimum absolute lymphocyte counts (Min ALCs) during radiotherapy (RT) and clinical outcomes in patients with hepatocellular carcinoma (HCC). METHODS: Data from a total of 69 HCC patients who had received RT were retrospectively analysed. Peripheral blood lymphocytes were measured before RT, weekly during RT and after RT. Regression and mixed-effect models were used to assess the relationships with and potential predictors of overall survival (OS). Receiver-operating characteristic (ROC) curve analysis was used to define optimal cut-off points of continuous variables for outcomes. RESULTS: The median follow up was 30 months (range, 4-68 months). The median survival time (MST), 1-year OS rate and 2-year OS rate of the whole group were 25 months, 51% and 39%, respectively. The average circulating lymphocyte counts declined during RT (1493.19 versus 503.48 cells/µl, p < 0.001). A lower Min ALC was associated with worse OS (p = 0.001), with a cut-off value of 450 cells/µl (sensitivity and specificity, 50% and 70.6%, respectively). The MSTs, 1-year OS rates and 2-year OS rates were 15 months versus 47 months, 27% versus 78% and 4% versus 71% for patients with relatively lower (⩽450 cells/µl) and higher Min ALCs (>450 cells/µl), respectively (p < 0.001). After adjusting for potential confounders, multivariate Cox regression analysis demonstrated that Min ALC independently predicted patients' OS (HR, 0.32; 95% CI, 0.15-0.69). CONCLUSIONS: Lower Min ALCs during RT may act as a worse prognostic factor for HCC after RT.

Comparison of intra-arterial chemoembolization with and without radiotherapy for advanced hepatocellular carcinoma with portal vein tumor thrombosis: a meta-analysis.

PURPOSE: Numerous studies have tried to combine transarterial chemoembolization (TACE) or hepatic arterial infusion chemotherapy (HAIC) with radiotherapy (RT) for the treatment of hepatocellular carcinoma (HCC) patients with portal vein tumor thrombus (PVTT). However, the efficacy of TACE or HAIC combined with RT versus TACE or HAIC alone remains controversial. Thus, we performed a meta-analysis to compare the efficacy and safety of intra-arterial chemoembolization combined with RT versus intra-arterial chemoembolization alone for the treatment of HCC patients with PVTT. METHODS: PubMed, Embase, and Cochrane Library databases were systematically searched for eligible studies. Two authors independently reviewed the abstracts, extracted relevant data and rated the quality of studies. The major end points were objective response rate (ORR), overall survival (OS), and adverse events. RESULTS: Eight studies with a total of 1,760 patients were included in this meta-analysis. The pooled results showed that intra-arterial chemoembolization combined with RT significantly improved ORR of PVTT (OR, 4.22; 95% CI, 3.07-5.80; P<0.001) and OS (HR, 0.69; 95% CI, 0.57-0.83; P=0.001), but did not affect ORR of primary liver tumor (OR, 1.37; 95% CI, 0.67-2.79; P=0.390). The incidence of grade 3 or 4 leukopenia (OR, 5.80; 95% CI, 2.478-13.56; P<0.001) and thrombocytopenia (OR, 3.77; 95% CI, 1.06-13.43; P=0.041) was higher in the intra-arterial chemoembolization plus RT group than in the intra-arterial chemoembolization group. CONCLUSION: Combination therapy of intra-arterial chemoembolization and RT for HCC patients with PVTT could bring higher ORR of PVTT and better survival benefits. This combination therapy was also associated with a significantly increased risk of adverse events. However, they were mostly mild to moderate and successfully treated with conservative treatment.

Superiority of helical tomotherapy on liver sparing and dose escalation in hepatocellular carcinoma: a comparison study of three-dimensional conformal radiotherapy and intensity-modulated radiotherapy.

BACKGROUND AND PURPOSE: To compare the difference of liver sparing and dose escalation between three-dimensional conformal radiotherapy (3DCRT), intensity-modulated radiotherapy (IMRT), and helical tomotherapy (HT) for hepatocellular carcinoma. PATIENTS AND METHODS: Sixteen unresectable HCC patients were enrolled in this study. First, some evaluation factors of 3DCRT, IMRT, and HT plans were calculated with prescription dose at 50 Gy/25 fractions. Then, the doses were increased using HT or IMRT independently until either the plans reached 70 Gy or any normal tissue reached the dose limit according to quantitative analysis of normal tissue effects in the clinic criteria. RESULTS: The conformal index of 3DCRT was lower than that of IMRT (P<0.001) or HT (P<0.001), and the homogeneity index of 3DCRT was higher than that of IMRT (P<0.001) or HT (P<0.001). HT took the longest treatment time (P<0.001). For V 50% (fraction of normal liver treated to at least 50% of the isocenter dose) of the normal liver, there was a significant difference: 3DCRT > IMRT > HT (P<0.001). HT had a lower D mean (mean dose) and V 20 (V n, the percentage of organ volume receiving ≥n Gy) of liver compared with 3DCRT (P=0.005 and P=0.005, respectively) or IMRT (P=0.508 and P=0.007, respectively). D mean of nontarget normal liver and V 30 of liver were higher for 3DCRT than IMRT (P=0.005 and P=0.005, respectively) or HT (P=0.005 and P=0.005, respectively). Seven patients in IMRT (43.75%) and nine patients in HT (56.25%) reached the isodose 70 Gy, meeting the dose limit of the organs at risk. CONCLUSION: HT may provide significantly better liver sparing and allow more patients to achieve higher prescription dose in HCC radiotherapy.

Astaxanthin reduces type 2 diabetic­associated cognitive decline in rats via activation of PI3K/Akt and attenuation of oxidative stress.

Astaxanthin (AST) is an oxygenated derivative of carotenoid, which possesses a strong antioxidant activity. AST can effectively remove active oxygen from the body, and is thus considered to have an important role in disease prevention and treatment. The present study aimed to determine the effects of AST on type 2 diabetic­associated cognitive decline (DACD) in rats. Rats were intraperitoneally injected with streptozotocin (STZ), in order to establish a model of diabetes mellitus (DM). A total of 40 rats were randomly divided into five groups: The control group, the DM group, the AST (50 mg/kg) group, the AST (100 mg/kg) group, and the AST+LY294002 group (AST, 50 mg/kg and LY, 0.25 µg/100 g). Following a 14­day treatment with AST, the body weight, blood glucose levels and cognitive function were determined. In addition, the protein expression levels of phosphatidylinositol 3­kinase (PI3K)/Akt, glutathione peroxidase and superoxide dismutase activity, glutathione and malondialdehyde content, and inducible nitric oxide synthase (iNOS), caspase­3 and caspase­9 activity were detected in the rats with DM. AST clearly augmented body weight and reduced blood glucose levels in rats with DM. Furthermore, treatment with AST significantly improved the cognitive function of rats with DM. Treatment with AST activated the PI3K/Akt pathway, and suppressed oxidative stress in the DM rats. In the cerebral cortex and hippocampus of the rats with DM, the activities of iNOS, caspase­3 and caspase­9 were markedly reduced. Furthermore, treatment with the Akt inhibitor LY294002 reduced the effectiveness of AST on DACD in rats. In conclusion, AST may reduce type 2 DACD in rats via activation of PI3K/Akt and attenuation of oxidative stress.

Naringin ameliorates cognitive deficits via oxidative stress, proinflammatory factors and the PPARγ signaling pathway in a type 2 diabetic rat model.

Naringenin is a flavonoid polyphenolic compound, which facilitates the removal of free radicals, oxidative stress and inflammation. The present study aimed to obtain a better understanding of the effects of curcumin on the regulation of diabetes­associated cognitive decline, and its underlying mechanisms. An experimental diabetes mellitus (DM) rat model was induced by streptozoticin (50 mg/kg). Following treatment with naringin (100 and 200 mg/kg) for 16 weeks, the body weight and blood glucose levels of the DM rats were measured. A morris water maze test was used to analyze the effects of naringin on the cognitive deficit of the DM rats. The levels of oxidative stress, proinflammatory factors, caspase­3 and caspase­9, and the protein expression of peroxisome proliferator­activated receptor γ (PPARγ) were quantified in the DM rats using a commercially­available kit and western blot assay, respectively. In addition, a GW9662 PPARγ inhibitor (0.3 mg/kg) was administered to the DM rats to determine whether PPARγ affected the effects of naringin on the cognitive deficit of the DM rats. The results demonstrated that naringin increased the body weight, blood glucose levels, and cognitive deficits of the DM rats. The levels of oxidative stress and proinflammatory factors in the naringin­treated rats were significantly lower, compared with those of the DM rats. In addition, naringin activated the protein expression of PPARγ, and administration of the PPARγ inhibitor decreased the protein expression of PPARγ, and attenuated the effects of naringin on cognitive deficit. The results also demonstrated that naringin decreased the expression levels of caspase­3 and caspase­9 in the DM rats. These results suggested that naringin ameliorated cognitive deficits via oxidative stress, proinflammatory factors and the PPARγ signaling pathway in the type 2 diabetic rat model. Furthermore, oxidative stress, proinflammatory factors and PPARγ signaling may be involved in mediating these effects.

Promoting effect of vanadium on catalytic activity of Pt/Ce-Zr-O diesel oxidation catalysts.

A series of Pt-V/Ce-Zr-O diesel oxidation catalysts was prepared using the impregnation method. The catalytic activity and sulfur resistance of Pt-V/Ce-Zr-O were investigated in the presence of simulated diesel exhaust. The effect of vanadium on the structure and redox properties of the catalysts was also investigated using the Brunauer-Emmett-Teller method, X-ray diffraction, H2 temperature-programmed reduction, CO temperature-programmed desorption, X-ray photoelectron spectroscopy, and Energy Dispersive Spectroscopy. Results showed that the Pt particles were well dispersed on the Ce-Zr-O carrier through the vanadium isolation effect, which significantly improved the oxidation activity toward CO and hydrocarbons. An electron-withdrawing phenomenon occurred from V to Pt, resulting in an increase in the metallic nature of platinum, which was beneficial to hydrocarbon molecular activation.

Paeoniflorin inhibits proliferation and induces apoptosis of human glioma cells via microRNA-16 upregulation and matrix metalloproteinase-9 downregulation.

Paeoniflorin is one of the active ingredients of the commonly used herbal medicine derived from Paeonia, which exhibits anticancer properties. MicroRNA-16 (miR-16) is upregulated in CD133(-) cells, but downregulated in CD133(+) cells from glioma tissue. Matrix metalloproteinase-9 (MMP-9) expression in glioma tissue samples is significantly higher than that in healthy brain tissue samples. Therefore, miR-16 and MMP-9 expression may be associated with glioma pathogenesis. In the present study, the effects of paeoniflorin on glioma were analyzed. U87 cells were treated with paeoniflorin at 0, 5, 10 and 20 µΜ concentrations. The results suggested that paeoniflorin inhibited U87 cell proliferation and accelerated cell apoptosis. In the present study paeoniflorin treatment increased miR-16 expression and reduced MMP-9 protein expression in U87 cells. Additionally, the results of the present study suggested that miR-16 may regulate MMP-9 expression in miR-16-transfected U87 cells. Furthermore, anti-miR-16 antibodies were used in order to investigate the apoptotic effects of paeoniflorin on U87 cells. The results demonstrated that paeoniflorin inhibits proliferation and induces apoptosis of human glial cells, via miR-16 upregulation and MMP-9 downregulation.

Baicalein alters PI3K/Akt/GSK3ß signaling pathway in rats with diabetes-associated cognitive deficits.

Our present investigation focused on assessing the neuroprotective potential of baicalein (BAC) against diabetes-associated cognitive deficit (DACD) using a diabetic model and further figure out the potential molecular mechanisms. Diabetic rat model was established by streptozotocin (STZ). Vehicle or BAC by the doses of 2 and 4 mg/kg was intraperitoneally injected once a day for seven consecutive weeks. Memory function was evaluated by Morris water maze test and avoidance passive test. The activities of acetylcholinesterase (AChE), choline acetylase (ChAT), caspase-9 and caspase-3 in STZ-induced diabetic rats' hippocampus were detected via responsive commercial kits. Western blot assay were used to determine the protein levels of phospho-phosphatidylinositol 3-kinase (p-PI3K), phospho-Akt (p-Akt), and phospho-glycogen synthase kinase-3ß (p-GSK3ß). Our results showed that BAC remarkably increased body weight and ChAT activity, decreased blood glucose level and AChE activity as well as improved cognitive deficits in diabetic rats. Additionally, it was also found that treatment with BAC to diabetes obviously stimulated the p-PI3K and p-Akt and inhibited the level of p-GSK3ß. Furthermore, the neuronal apoptosis was also prevented after BAC treatment by decreasing caspase-9 and caspase-3 activities in diabetic rats' hippocampus. It is concluded that BAC exerted beneficial effects against DACD in rats and its neuroprotection might be linked with activating PI3K and Akt phosphorylation accompanied with suppressing the phosphorylated level of GSK3ß. These results hint that BAC is likely to be served as an adjuvant therapy to conventional anti-hyperglycemic regimens as well as DACD.

Chrysin ameliorates diabetes-associated cognitive deficits in Wistar rats.

Chrysin (CH) is an important natural plant flavonoid and possesses diverse pharmacological activities. Our present investigations aimed to assess the neuroprotection of CH against diabetes-associated cognitive decline (DACD) in a rat model of diabetes and exploring its potential mechanism. Diabetic model was induced by intraperitoneal injection of streptozotocin. Then, they were treated with vehicle or CH by doses of 30 and 100 mg/kg for 26 days. Learning and memory function was evaluated by Morris water maze test. The oxidative indicators [malondialdehyde (MDA), catalase (CAT), superoxide dismutase (SOD) and glutathione (GSH)], NF-κB p65 unit, TNF-α, IL-1ß, IL-6 and caspase-3 were measured in cerebral cortex and hippocampus using corresponding commercial kits. The diabetic rats showed marked reductions in body weight, percentage of time spent in target quadrant and number of times of crossing platform, coupled with increases in plasma glucose levels, escape latency, mean path length and oxidative stress (increased MDA level and decreased CAT and SOD as well as reduced GSH), NF-κB p65 unit, TNF-α, IL-1ß, IL-6 and caspase-3 in cerebral cortex and hippocampus. Moreover, CH supplement dramatically reversed the corresponding behavioral, biochemical and molecular alterations in diabetes. The alterations of swimming speed among different groups were not observed after CH adminstration. In conclusion, our current work discloses that CH remarkably alleviates DACD and suggests that oxidative stress, inflammation and apoptotic cascades are linked with diabetes-associated cognitive deficits. These findings point toward the therapeutic potential of CH in DACD.

SOD1, but not SOD3, deficiency accelerates diabetic renal injury in C57BL/6-Ins2(Akita) diabetic mice.

Superoxide dismutase (SOD) is a major defender against excessive superoxide generated under hyperglycemia. We have recently reported that renal SOD1 (cytosolic CuZn-SOD) and SOD3 (extracellular CuZn-SOD) isoenzymes are remarkably down-regulated in KK/Ta-Ins2(Akita) diabetic mice, which exhibit progressive diabetic nephropathy (DN), but not in DN-resistant C57BL/6- Ins2(Akita) (C57BL/6-Akita) diabetic mice. To determine the role of SOD1 and SOD3 in DN, we generated C57BL/6-Akita diabetic mice with deficiency of SOD1 and/or SOD3 and investigated their renal phenotype at the age of 20 weeks. Increased glomerular superoxide levels were observed in SOD1(-/-)SOD3(+/+) and SOD1(-/-)SOD3(-/-) C57BL/6-Akita mice but not in SOD1(+/+)SOD3(-/-) C57BL/6-Akita mice. The SOD1(-/-)SOD3(+/+) and SOD1(-/-)SOD3(-/-) C57BL/6-Akita mice exhibited higher glomerular filtration rate, increased urinary albumin levels, and advanced mesangial expansion as compared with SOD1(+/+)SOD3(+/+) C57BL/6-Akita mice, yet the severity of DN did not differ between the SOD1(-/-)SOD3(+/+) and SOD1(-/-)SOD3(-/-) C57BL/6-Akita groups. Increased renal mRNA expression of transforming growth factor-ß1 (TGF-ß1) and connective tissue growth factor (CTGF), reduced glomerular nitric oxide (NO), and increased renal prostaglandin E2 (PGE2) production were noted in the SOD1(-/-)SOD3(+/+) and SOD1(-/-)SOD3(-/-) C57BL/6-Akita mice. This finding indicates that such renal changes in fibrogenic cytokines, NO, and PGE2, possibly caused by superoxide excess, would contribute to the development of overt albuminuria by promoting mesangial expansion, endothelial dysfunction, and glomerular hyperfiltration. The present results demonstrate that deficiency of SOD1, but not SOD3, increases renal superoxide in the setting of diabetes and causes overt renal injury in nephropathy-resistant diabetic mice, and that SOD3 deficiency does not provide additive effects on the severity of DN in SOD1-deficient C57BL/6-Akita mice.

Better nephrology for mice--and man.

The use of creatinine to estimate glomerular filtration rate in patients is prone to well-described artifacts that impact its interpretation. Eisner et al. now show that the impact of creatinine secretion on creatinine clearance is even larger in mice than in humans, raising questions regarding the utility of creatinine for measuring glomerular filtration rate in mice.

Reduction of renal superoxide dismutase in progressive diabetic nephropathy.

Superoxide excess plays a central role in tissue damage that results from diabetes, but the mechanisms of superoxide overproduction in diabetic nephropathy (DN) are incompletely understood. In the present study, we investigated the enzyme superoxide dismutase (SOD), a major defender against superoxide, in the kidneys during the development of murine DN. We assessed SOD activity and the expression of SOD isoforms in the kidneys of two diabetic mouse models (C57BL/6-Akita and KK/Ta-Akita) that exhibit comparable levels of hyperglycemia but different susceptibility to DN. We observed down-regulation of cytosolic CuZn-SOD (SOD1) and extracellular CuZn-SOD (SOD3), but not mitochondrial Mn-SOD (SOD2), in the kidney of KK/Ta-Akita mice which exhibit progressive DN. In contrast, we did not detect a change in renal SOD expression in DN-resistant C57BL/6-Akita mice. Consistent with these findings, there was a significant reduction in total SOD activity in the kidney of KK/Ta-Akita mice compared with C57BL/6-Akita mice. Finally, treatment of KK/Ta-Akita mice with a SOD mimetic, tempol, ameliorated the nephropathic changes in KK/Ta-Akita mice without altering the level of hyperglycemia. Collectively, these results indicate that down-regulation of renal SOD1 and SOD3 may play a key role in the pathogenesis of DN.

Measurement of glomerular filtration rate in conscious mice.

Glomerular filtration rate (GFR) is an important index of renal function and routinely used in patient care and basic research to evaluate progression of renal diseases or test the efficacy of novel therapeutic strategies. Determination of GFR in mouse models has been mostly practiced in anesthetized animals, which is not suitable for serial monitoring of GFR in the individual mouse. In this chapter, we outline two approaches for determining GFR in conscious mice including 1) determination of urinary excretion of fluorescein-labelled inulin (FITC-inulin), and 2) determination of plasma FITC-inulin decay following a single bolus injection. The GFR values determined using these two methods are comparable. The sensitivity of the methods in reflecting renal function was validated in nephrectomized mice and early stage diabetic mice. The effects of inbred mouse genetic background on GFR values are also discussed in this chapter.

Markers of glycemic control in the mouse: comparisons of 6-h- and overnight-fasted blood glucoses to Hb A1c.

The present studies examined the relationship between fasting blood glucose and Hb A(1c) in C57BL/6J, DBA/2J, and KK/HlJ mice with and without diabetes mellitus. Daily averaged blood glucose levels based on continuous glucose monitoring and effects of 6-h vs. overnight fasting on blood glucose were determined. Daily averaged blood glucose levels were highly correlated with Hb A(1c), as determined with a hand-held automated device using an immunodetection method. R(2) values were 0.90, 0.95, and 0.99 in KK/HIJ, C57BL/6J, and DBA/2J, respectively. Six-hour fasting blood glucose correlated more closely with the level of daily averaged blood glucose and with Hb A(1c) than did blood glucose following an overnight fast. To validate the immunoassay-determined Hb A(1c), we also measured total glycosylated hemoglobin using boronate HPLC. Hb A(1c) values correlated well with total glycosylated hemoglobin in all three strains but were relatively lower than total glycosylated hemoglobin in diabetic DBA/2J mice. These results show that 6-h fasting glucose provides a superior index of glycemic control and correlates more closely with Hb A(1c) than overnight-fasted blood glucose in these strains of mice.