Renal function is crucially dependent on renal microstructure which provides the basis for the regulatory mechanisms that control the transport of water and solutes between filtrate and plasma and the urinary concentration. This study provides new, detailed information on mouse renal architecture, including the spatial course of the tubules, lengths of different segments of nephrons, histotopography of tubules and vascular bundles, and epithelial ultrastructure at well-defined positions along Henle's loop and the distal convolution of nephrons. Three-dimensional reconstruction of 200 nephrons and collecting ducts was performed on aligned digital images, obtained from 2.5-m-thick serial sections of mouse kidneys. Important new findings were highlighted: (1) A tortuous course of the descending thin limbs of long-looped nephrons and a winding course of the thick ascending limbs of short-looped nephrons contributed to a 27% average increase in the lengths of the corresponding segments, (2) the thick-walled tubules incorporated in the central part of the vascular bundles in the inner stripe of the outer medulla were identified as thick ascending limbs of long-looped nephrons, and (3) three types of short-looped nephron bends were identified to relate to the length and the position of the nephron and its corresponding glomerulus. The ultrastructure of the tubule segments was identified and suggests important implications for renal transport mechanisms that should be considered when evaluating the segmental distribution of water and solute transporters within the normal and diseased kidney. T he renal architecture is arranged elaborately to fulfill the physiologic demands for reabsorption of filtered substances and for urine concentration. The architecture includes the formation of the renal zones, the population of short-looped nephrons (SLN) and long-looped nephrons (LLN), the distribution of the tubule segments of nephrons, the tubular-vascular histotopography in medulla, and the epithelial configurations of the different tubule segments.From the 1960s to the 1980s, Kriz and others studied the renal microstructure, including the distribution of nephrons, the tubular-vascular relations in the medullary zones, and the ultrastructure of the epithelia along Henle's loop. Simultaneously, the diversities in renal structure were demonstrated between different species, including rat, mouse, hamster, and rabbit (1-3). In the late 1980s, a standard nomenclature for the kidney structure, which has been widely adopted, was published (4). With the development of digital techniques, three-dimensional (3D) representations of the tubule segments of nephrons were performed to a limited extent at the proximal tubules (PT) (5), distal tubule (6), and at the thin limbs (TL) of the LLN in the inner medulla (IM) (7) of rats. Two mathematical regionalbased models combining morphologic and immunohistochemical findings have been set up to simulate the urine concentration mechanism in rat medulla (8).At present, a large body of basic nephrol...

Folic acid- (FA-) induced kidney injury is characterized by the tubule damage due to the disturbance of the antioxidant system and subsequent interstitial fibrosis. FG-4592 is an inhibitor of prolyl hydroxylase of hypoxia-inducible factor (HIF), an antioxidant factor. The present study investigated the protective role of FG-4592 pretreatment at the early stage of the kidney injury and long-term impact on the progression of renal fibrosis. FG-4592 was administrated two days before FA injection in mice. On the second day after FA injection, the mice with FG-4592 pretreatment showed an improved renal function, compared with those without FG-4592 pretreatment, indicated by biochemical and histological parameters; meanwhile, the cellular content of iron, malondialdehyde, and 4-hydroxynonenal histologically decreased, implying the suppression of iron accumulation and lipid peroxidation. Simultaneously, upregulation of HIF-1α was found, along with Nrf2 activation, which was reflected by increased nuclear translocation and high-expression of downstream proteins, including heme-oxygenase1, glutathione peroxidase4, and cystine/glutamate transporter, as well as ferroportin. Correspondingly, the elevated levels of antioxidative enzymes and glutathione, as well as reduced iron accumulation, were observed, suggesting a lower risk of occurrence of ferroptosis with FG-4592 pretreatment. This was confirmed by reversed pathological parameters and improved renal function in FA-treated mice with the administration of ferrostatin-1, a specific ferroptosis inhibitor. Furthermore, a signal pathway study indicated that Nrf2 activation was associated with increased phosphorylation of Akt and GSK-3β, verified by the use of an inhibitor of the PI3K that phosphorylates Akt. Moreover, FG-4592 pretreatment also decreased macrophage infiltration and expression of inflammatory factors TNF-α and IL-1β. On the 14th day after FA injection, FG-4592 pretreatment decreased collagen deposition and expression of fibrosis biomarkers. These findings suggest that the protective role of FG-4592 pretreatment is achieved mainly by decreasing ferroptosis at the early stage of FA-induced kidney injury via Akt/GSK-3β-mediated Nrf2 activation, which retards the fibrosis progression.

BACKGROUND AND PURPOSELiver fibrosis is commonly associated with obesity and most obese patients develop hyperleptinaemia. The adipocytokine leptin has a unique role in the development of liver fibrosis. Activation of hepatic stellate cells (HSCs) is a key step in hepatic fibrogenesis and sterol regulatory element-binding protein-1c (SREBP-1c) can inhibit HSC activation. We have shown that leptin strongly inhibits SREBP-1c expression in rat HSCs. Hence, we aimed to clarify whether the b-catenin pathway, the crucial negative regulator of adipocyte differentiation, mediates the effects of leptin on SREBP-1c expression in HSCs and in mouse liver fibrosis. EXPERIMENTAL APPROACHHSCs were prepared from rats and mice. Gene expressions were analysed by real-time PCR, Western blot analysis, immunostaining and transient transfection assays. KEY RESULTSLeptin increased b-catenin protein but not mRNA levels in cultured HSCs. Leptin induced phosphorylation of glycogen synthase kinase-3b at Ser 9 and subsequent stabilization of b-catenin protein was mediated, at least in part, by ERK and p38 MAPK pathways. The leptin-induced b-catenin pathway reduced SREBP-1c expression and activity but did not affect protein levels of key regulators controlling SREBP-1c activity, and was not involved in leptin inhibition of liver X receptor a. In a mouse model of liver injury, the b-catenin pathway was shown to be involved in leptin-induced liver fibrosis. CONCLUSIONS AND IMPLICATIONSThe b-catenin pathway contributes to leptin regulation of SREBP-1c expression in HSCs and leptin-induced liver fibrosis in mice. These results have potential implications for clarifying the mechanisms of liver fibrogenesis associated with elevated leptin levels. Abbreviationsa-SMA, alpha-smooth muscle actin; ER, endoplasmic reticulum; FFA, free fatty acid; GSK-3b, glycogen synthase kinase-3b; HSC, hepatic stellate cell; Insig, insulin-induced proteins; LXR, liver X receptor; SCAP, SREBP cleavage-activating protein; SREBP-1c, sterol regulatory element-binding protein-1c

Factor X a (FX a ) binding to factor V a (FV a ) on plateletderived membranes containing surface-exposed phosphatidylserine (PS) forms the "prothrombinase complex" that is essential for efficient thrombin generation during blood coagulation. There are two naturally occurring isoforms of FV a , FV a1 and FV a2 . These two isoforms differ by a 3-kDa polysaccharide chain ( ). The ability of soluble PS to trigger formation of a soluble prothrombinase complex suggests that exposure of PS molecules during platelet activation is likely the key event responsible for the assembly of an active membrane-bound complex.The final step in the blood coagulation cascade involves the activation of prothrombin to thrombin, which is the central enzyme of the coagulation system. This activation requires assembly of an enzyme complex, called prothrombinase (1), which consists of blood coagulation factors X a (a serine protease) and V a (a cofactor), Ca 2ϩ , and membranous vesicles derived from stimulated platelets (2). Several studies (3-5) have suggested that phosphatidylserine (PS) 1 might play a specific role in prothrombin activation. PS is asymmetrically distributed to the cytoplasmic surface of resting platelet membranes (6) but is exposed when human platelets are activated (7). It has become clear only very recently that PS regulates the structure and function of factors X a and V a (8, 10). 2 Here we explore further the extent of this regulation.Factor V exists in plasma as an inactive, single chain glycoprotein with a molecular mass of 330 kDa. The active form of factor V, FV a , has a central domain removed to yield a heterodimer composed of two chains, a heavy chain (M r ϭ 94,000 in the bovine species; 105,000 in human) and a heterogeneous light chain (M r ϭ 74,000 in FV a1 or 71,000 in FV a2 ). The heavy and light chains form a tight complex in the presence of a calcium ion (11). The heterogeneity in the light chain is seen in both the bovine and human molecules. In the human form, it appears to arise from glycosylation of Asn 2181 at the C-terminal end of the light chain (12). Prothrombinase complexes assembled from the two molecular species derived from human plasma are observed to have somewhat different cofactor activities (13,14). This has been attributed to substantially different affinities for binding to membranes (13). However, our lab has reported that the two forms of both bovine and human FV a bind to membranes with only ϳ3-fold different affinities (12,14). This suggests that differences in the ability to support prothrombinase activity must reflect either different binding between factors FX a and FV a1 versus FV a2 or different intrinsic activities of the FX a ⅐FV a1 and FX a ⅐FV a2 complexes. Although our results have favored the former possibility (14), it has been difficult to prove this unambiguously because it is difficult to measure precisely the interaction between FX a and FV a on a membrane surface (15).The presence of FV a in a reaction mixture is critical to obtaining a maximal and physiologicall...

Our studies show that genistein up-regulated HO-1 and GCLC expression through the EKR1/2 and PKC /Nrf2 pathways during oxidative stress.