Podocytes are new cellular targets of haemoglobin‐mediated renal damage

Recurrent and massive intravascular haemolysis induces proteinuria, glomerulosclerosis, and progressive impairment of renal function, suggesting podocyte injury. However, the effects of haemoglobin (Hb) on podocytes remain unexplored. Our results show that cultured human podocytes or podocytes isolated from murine glomeruli bound and endocytosed Hb through the megalin–cubilin receptor system, thus resulting in increased intracellular Hb catabolism, oxidative stress, activation of the intrinsic apoptosis pathway, and altered podocyte morphology, with decreased expression of the slit diaphragm proteins nephrin and synaptopodin. Hb uptake activated nuclear factor erythroid‐2‐related factor 2 (Nrf2) and induced expression of the Nrf2‐related antioxidant proteins haem oxygenase‐1 (HO‐1) and ferritin. Nrf2 activation and Hb staining was observed in podocytes of mice with intravascular haemolysis. These mice developed proteinuria and showed podocyte injury, characterized by foot process effacement, decreased synaptopodin and nephrin expression, and podocyte apoptosis. These pathological effects were enhanced in Nrf2‐deficient mice, whereas Nrf2 activation with sulphoraphane protected podocytes against Hb toxicity both in vivo and in vitro. Supporting the translational significance of our findings, we observed podocyte damage and podocytes stained for Hb, HO‐1, ferritin and phosphorylated Nrf2 in renal sections and urinary sediments of patients with massive intravascular haemolysis, such as atypical haemolytic uraemic syndrome and paroxysmal nocturnal haemoglobinuria. In conclusion, podocytes take up Hb both in vitro and during intravascular haemolysis, promoting oxidative stress, podocyte dysfunction, and apoptosis. Nrf2 may be a potential therapeutic target to prevent loss of renal function in patients with intravascular haemolysis. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Evidence for lipid peroxidation by paraquat in the perfused rat lung

In order to evaluate the effect of paraquat on oxidative radical reactions in the lung, we studied MDA production and chemiluminescence (spontaneous and tBuOOH-induced) in the isolated rat lung. After 2 hr of perfusion with 3.0 mM paraquat, MDA content in lung homogenates was 16 +/- 7 nmol/gm dry weight higher than in control lungs (mean +/- S.E., n = 7, p less than 0.05 by paired test); during 30 min of perfusion, malondialdehyde efflux was 33 +/- 15 nmol/gm dry weight higher than in control perfusates (n = 6, p less than 0.05). Spontaneous chemiluminescence was not augmented by 2 hr of perfusion with concentrations of paraquat ranging from 0.75 to 6.0 mM. On the other hand, tBuOOH-induced chemiluminescence was 17% +/- 3 higher immediately after the addition of hydroperoxide and reached a 16% +/- 6 higher plateau for the paraquat-perfused lungs than for control lungs (n = 10, p less than 0.05). Spectral analysis of the light emitted during induced chemiluminescence demonstrated peak intensity between 630 and 730 nm for both control and paraquat-treated lungs. Increased MDA production and increased induced chemiluminescence indicate that perfusion with paraquat enhances lipid peroxidation in the isolated rat lung.     

Organ chemiluminescence: noninvasive assay for oxidative radical reactions.

In situ and perfused rat livers showed a spontaneous chemiluminescence of 7-12 counts/sec . cm2 (corresponding to 7-12 x 10(3) photons/sec . cm2); chemiluminescence was increased up to 30 times by infusion of exogenous hydroperoxides. The chemiluminescence of the perfused liver was oxygen dependent. Ethyl, t-butyl, and cumene hydroperoxides were almost equally effective in inducing light emission in the perfused liver. Glutathione release and chemiluminescence showed a parallel increase upon hydroperoxide supply to the perfused liver. A partial spectral analysis of the chemiluminescence of the perfused liver showed a predominance of red-light-emitting species, presumably arising from the singlet oxygen dimol-emission peaks. Many side reactions derived from the complex free radical sequence of lipid peroxidation could afford the chemistry leading to light emission, which represents only about 10(-14) of the utilization of peroxide.     

Assessment of intra- and interobserver reproducibility of rest and cold pressor test-stimulated myocardial blood flow with <Superscript>13</Superscript>N-ammonia and PET

Purpose We investigated the intraobserver reproducibility of myocardial blood flow (MBF) measurements with PET at rest and during cold pressor test (CPT), and the interobserver agreement. Methods Twenty normal volunteers were studied. Using ¹³N-ammonia, MBF was measured at rest and during CPT and measurement was repeated in a 1-day session (short-term reproducibility; SR). After a follow-up of 2 weeks, MBF was measured again at rest and during CPT and compared with the initial baseline measurement (long-term reproducibility; LR). In addition, adenosine-induced hyperemic MBF increases were assessed. Results Assessment of the SR did not show a significant absolute difference in MBF at rest, MBF during CPT or the endothelium-related change in MBF from rest to CPT (ΔMBF) (0.09 ± 0.10, 0.11 ± 0.09, and 0.08 ± 0.05 ml/g/min; p = NS), and they were linearly correlated (r = 0.72, r = 0.76 and r = 0.84; p < 0.0001). Corresponding values for standard error of the estimate (SEE), as indicative for the range of MBF measurement error, were 0.14, 0.14, and 0.09 ml/g/min. The LR yielded relatively higher but non-significant absolute differences in the MBF at rest, MBF during CPT and ΔMBF (0.10 ± 0.10, 0.14 ± 0.10, and 0.19 ± 0.10 ml/g/min; p = NS), and paired MBFs significantly correlated (r = 0.75, r = 0.71, and r = 0.60; p < 0.001). Corresponding SEEs were 0.13, 0.15, and 0.16 ml/g/min. The interobserver analysis yielded a high correlation for MBF at rest, MBF during CPT, and hyperemic MBF (r = 0.96, SEE=0.04; r = 0.78, SEE=0.11; and r = 0.87, SEE=0.28; p < 0.0001, respectively), and also a good interobserver correlation for ΔMBF (r = 0.62, SEE=0.09; p < 0.003). Conclusion Short- and long-term MBF responses to CPT, as an index for endothelium-related coronary vasomotion, can be measured reproducibly with ¹³N-ammonia PET. In addition, the high interobserver reproducibility for repeat analysis of MBF values suggests the measurements to be largely operator independent. Thomas H. Schindler and Xiao-Li Zhang contributed equally to this paper.     

A threshold concentration of FSH is needed during IVM of ex vivo collected human oocytes

Journal of Assisted Reproduction and Genetics - To investigate the effect of different FSH concentrations on human oocyte maturation in vitro and its impact on gene expression of key factors in the...     

Oxidative DNA damage estimated by oxo8dG in the liver of guinea-pigs supplemented with graded dietary doses of ascorbic acid and alpha- tocopherol

Dietary antioxidants may influence cancer risk and aging by modifying oxidative damage. The effect of graded dietary doses of the antioxidant vitamins C and E on oxidative DNA damage was studied in the liver of guinea-pigs under normal conditions. Like human beings, guinea-pigs cannot synthesize ascorbate and alpha-tocopherol. In one experiment, three groups of 6-8 guinea-pigs were fed diets containing 15 mg of vitamin E/kg chow and three different amounts of vitamin C (33,660 or 13,200 mg/kg) for 5 weeks. In a second experiment, three groups of seven guinea-pigs were fed diets containing 660 mg of vitamin C/kg and three different amounts of vitamin E (15, 150 or 1500 mg/kg) for 5 weeks. The three graded levels of each vitamin respectively represent marginal deficiency, an optimum supplementation and a megadose. Oxidative damage to liver DNA was estimated by measuring 8-oxo-7,8- dihydro-2'-deoxyguanosine (oxo8dG) referred to deoxyguanosine (dG) by means of high-performance liquid chromatography with simultaneous electrochemical-coulometric and ultraviolet detection. The level of ascorbate in the liver was 0.034 +/- 0.051, 1.63 +/- 1.06 and 1.99 +/- 0.44 micromol/g in the low, medium and high dose ascorbate groups (59- fold variation). The liver concentration of alpha-tocopherol was 28 +/- 11, 63 +/- 18 and 187 +/- 34 nmol/g in the low, medium and high dose alpha-tocopherol groups (7-fold variation). The level of oxo8dG in the liver DNA was 1.89 +/- 0.32, 1.94 +/- 0.78 and 1.93 +/- 0.65 per 10(5) dG in the low, medium and high dose ascorbate groups (no effect: P > 0.05). In the low, medium and high dose alpha-tocopherol groups oxo8dG level in the liver DNA was 2.85 +/- 0.70, 2.74 +/- 0.66 and 2.61 +/- 0.92 per 10(5) dG (no effect: P > 0.05). It is concluded that even very large variations in the content of the antioxidant vitamins C and E in the diet and liver have no influence on the steady-state level of oxidative damage to guanine in the liver DNA of normal unstressed guinea-pigs.      

Skeletal muscle protein anabolic response to increased energy and insulin is preserved in poorly controlled type 2 diabetes

Type 2 diabetes (T2DM) subjects failing diet treatment are characterized by hyperinsulinemia and insulin resistance leading to fasting and postprandial hyperglycemia and hyperlipidemia. Energy is essential for allowing the process of protein synthesis to proceed. Additionally, insulin can stimulate protein synthesis in human muscle. The aims of this study were to determine if poorly controlled T2DM affects postabsorptive muscle protein anabolism, and if the muscle anabolic response to hyperinsulinemia with high energy availability is maintained. Control (n = 6) and T2DM subjects (n = 6) were studied in the postabsorptive state and during an isoenergetic high nutritional energy clamp (relative to postabsorptive state). Muscle protein synthesis and breakdown (nmol . min(-1) . 100 g leg muscle(-1)) were assessed using stable isotope methodology, femoral arterio-venous sampling, muscle biopsies, and a three-pool model to calculate protein turnover. Postabsorptive phenylalanine net balance and whole body rate of appearance (Ra) were not different between groups; however, basal muscle protein breakdown was higher in T2DM (94 +/- 9) than in controls (58 +/- 12) (P < 0.05) and muscle protein synthesis tended (P = 0.07) to be elevated in T2DM (66 +/- 14) compared with controls (39 +/- 6). During the clamp, net balance increased, whole body Ra and muscle protein breakdown decreased (P < 0.05), and muscle protein synthesis tended to decrease (P = 0.08) to a similar extent in both groups. We conclude that postabsorptive muscle protein turnover is elevated in poorly controlled T2DM, however, there is no excessive loss of muscle protein because net balance is not different from controls. Moreover, the anabolic response to increased insulin and energy availability is maintained in T2DM.     

Redox interactions of nitric oxide with dopamine and its derivatives

Nitric oxide (*NO) is a ubiquitous diffusible messenger in the central nervous system. *NO and derived nitrogen species may interact with catecholamines, thus, modifying not only its regulatory actions but also producing oxidants and free radicals that are likely to trigger toxic pathways in the nervous system. Oxidative pathways and chain oxidation reactions triggered by catecholamines may be broken by ascorbate and glutathione, of which there is ample supply in the brain. At the subcellular level, mitochondria and cytosolic dopamine storage vesicles are likely to provide site-specific settings for *NO and catecholamines interactions. Thus, a complex picture emerges in which the steady- state levels of the individual reactants, the rate constants of the reactions involved, the oxygen tension, and the compartmentalization of reactions determine the biological significance of the redox interactions between *NO and dopamine metabolism in the brain. The physiological relevance of *NO-driven chemical modifications of dopamine and its derivatives and the ensuing free radical production are discussed in connection with the neurodegeneration inherent in Parkinson's disease.     

Interferon-inducible guanylate binding protein (GBP2) is associated with better prognosis in breast cancer and indicates an efficient T cell response

Background

Recently, interferon-inducible guanylate binding protein (GBP2) has been discussed as a possible control factor in tumor development, which is controlled by p53, and inhibits NF-Kappa B and Rac protein as well as expression of matrix metalloproteinase 9. However, the potential role that GBP2 plays in tumor development and prognosis has not yet been studied.

Methods

We analyzed whether GBP2 mRNA levels are associated with metastasis-free interval in 766 patients with node negative breast carcinomas who did not receive systemic chemotherapy. Furthermore, response to anthracycline-based chemotherapy was studied in 768 breast cancer patients.

Results

High expression of GBP2 in breast carcinomas was associated with better prognosis in the univariate (P < 0.001, hazard ratio 0.763, 95 % CI 0.650–0.896) as well as in the multivariate Cox analysis (P = 0.008, hazard ratio 0.731, 95 % CI 0.580–0.920) adjusted to the established clinical factors age, pT stage, grading, hormone and ERBB2 receptor status. The association was particularly strong in subgroups with high proliferation and positive estrogen receptor status but did not reach significance in carcinomas with low expression of proliferation associated genes. Besides its prognostic capacity, GBP2 also predicted pathologically complete response to anthracycline-based chemotherapy (P = 0.0037, odds ratio 1.39, 95 % CI 1.11–1.74). Interestingly, GBP2 correlated with a recently established T cell signature, indicating tumor infiltration with T cells (R = 0.607, P < 0.001).

Conclusion

GBP2 is associated with better prognosis in fast proliferating tumors and probably represents a marker of an efficient T cell response.     

Y-chromosome diversity characterizes the Gulf of Oman

Arabia has served as a strategic crossroads for human disseminations, providing a natural connection between the distant populations of China and India in the east to the western civilizations along the Mediterranean. To explore this region's critical role in the migratory episodes leaving Africa to Eurasia and back, high-resolution Y-chromosome analysis of males from the United Arab Emirates (164), Qatar (72) and Yemen (62) was performed. The role of the Levant in the Neolithic dispersal of the E3b1-M35 sublineages is supported by the data, and the distribution and STR-based analyses of J1-M267 representatives points to their spread from the north, most likely during the Neolithic. With the exception of Yemen, southern Arabia, South Iran and South Pakistan display high diversity in their Y-haplogroup substructure possibly a result of gene flow along the coastal crescent-shaped corridor of the Gulf of Oman facilitating human dispersals. Elevated rates of consanguinity may have had an impact in Yemen and Qatar, which experience significant heterozygote deficiencies at various hypervariable autosomal STR loci.