Proximal tubular cysts in fetal human autosomal recessive polycystic kidney disease.
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Standard texts describe human autosomal recessive polycystic kidney disease (ARPKD) as a cystic kidney disease in which lesions are localized to collecting tubules. Murine models of ARPKD consistently demonstrate an early phase of proximal tubular (PT) cystic involvement, which disappears shortly after birth. This is followed by a phase of collecting tubular (CT) cyst formation and progressive enlargement leading to compromise of renal function and death. Because the description of cystic lesions in human ARPKD has been largely based on postnatal specimens, PT cyst formation was hypothesized to be a characteristic feature of fetal human, as well as murine, ARPKD. This study examines nephron segment-specific cyst localization histochemically by lectin binding in 11 human ARPKD specimens obtained at different fetal and postnatal ages. PT cysts were found in human fetal specimens from gestational age 14 wk to 26 wk. The percentage of cysts involving PT segments ranged from 2 to 41%. The cystic index of PT cysts ranged from 2 to 5. In all specimens in which PT cysts were found, both the percentage of CT cysts and their cystic index were equal to or greater than the percentage of PT cysts and the associated PT cystic index. PT cysts were absent in all kidney specimens older than 34 wk gestational age. It is concluded that human ARPKD, like murine ARPKD, has a transient phase of PT cyst formation during early fetal development. (+info)
The sgk, an aldosterone-induced gene in mineralocorticoid target cells, regulates the epithelial sodium channel.
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The sgk, an aldosterone-induced gene in mineralocorticoid target cells, regulates the epithelial sodium channel. Aldosterone increases sodium reabsorption in tight epithelia. The early phase of this stimulatory effect is thought to involve activation of apical sodium channels. To identify immediate-early genes that initiate this effect, we used a combination of polymerase chain reaction-based subtractive hybridization and differential display techniques. This review summarizes our recent findings. Aldosterone rapidly increases mRNA levels of a putative Ser/Thr kinase, sgk (or serum- and glucocorticoid-regulated kinase), in the native mineralocorticoid target cells, that is, in cortical collecting duct (CCD) cells. The induction of sgk mRNA occurs within 30 minutes of the addition of aldosterone and does not require de novo protein synthesis, indicating that sgk is an immediate/early aldosterone-induced gene. Induction of sgk by aldosterone is mediated through mineralocorticoid receptors (MRs), since it is prevented by ZK91857, an MR antagonist, but not by RU486, a glucocorticoid antagonist. In addition to aldosterone, RU28362, a pure glucocorticoid receptor agonist, also induced sgk mRNA, both in primary cultures of rabbit CCD cells and in the M-1 mouse CCD cell line. Sgk mRNA levels are also influenced by changes in the osmolality of the medium. In M-1 cells, incubation of cells for one hour in a mildly hypotonic medium decreased sgk mRNA levels, whereas incubation in hypertonic medium brought about opposite changes. To determine whether sgk is involved in the regulation of the epithelial sodium channel (ENaC), we coexpressed the full-length sgk cRNA in Xenopus oocytes with the three ENaC subunits. Expression of sgk resulted in a significant increase in the amiloride-sensitive Na current, suggesting that this protein kinase plays an important role in the early phase of aldosterone-stimulated Na transport. These results indicate that sgk is an aldosterone-induced immediate/early gene in native MR target cells, and is involved in the regulation of ion transport and possibly cell volume. (+info)
Physiologic resistance to the action of aldosterone.
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The collecting duct is one of the major targets for aldosterone's action. Experiments conducted several years ago suggested that the major site of action on Na+ and K+ transport was the cortical portion, the cortical collecting duct (CCD). Subsequent studies have shown that the entire collecting duct is capable of responding to aldosterone, but does so differently according to the region. The inner medullary collecting duct (IMCD), while exhibiting a relatively low rate of Na+ transport in isolated, perfused tubules, can develop substantial rates of Na+ transport when put in primary culture. The IMCD, in contrast to the CCD, usually secretes little K+. Investigations into the mechanisms for the lower rates of Na+ transport have revealed that transforming growth factor-beta (TGF-beta), which is endogenously produced in the inner medulla, can markedly reduce the natriferric action of aldosterone. This action of TGF-beta is not apparent within the first few hours of exposure, but its effects, even after removal, last for over 48 hours. The mechanism of this antagonism appears to involve pathways that are parallel and independent of the major transcriptional effects of aldosterone. (+info)
Aldosterone and potassium secretion by the cortical collecting duct.
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BACKGROUND: : Aldosterone has been implicated in the regulation of both Na and K concentrations in the plasma. Release of the hormone is known to be stimulated by high plasma K, and infusion of aldosterone lowers plasma K. However, the correlation between changes in mineralocorticoid levels and rates of K secretion is not perfect, suggesting that other factors may be involved. METHODS: : Patch-clamp recordings were made of K-channel activity in the split-open cortical collecting tubule of the rat. Estimates of channel density were made in cell-attached patches on the luminal membrane of principal cells of this segment. RESULTS: : Most of the K conductance of the apical membrane is mediated through low-conductance "SK" channels. The number of conducting SK channels is increased when animals are placed on a high-K diet. However, increasing plasma aldosterone levels by infusion of the hormone or by sodium restriction failed to change the number of active channels. CONCLUSIONS: : At least two circulating factors are required for the regulation of renal K secretion by the cortical collecting tubule. Aldosterone mainly stimulates secretion by increasing the driving force for K movement through apical channels. A second, as yet unidentified, factor increases the number of conducting K channels. (+info)
Nongenomic effects of aldosterone on Ca2+ in M-1 cortical collecting duct cells.
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BACKGROUND: Aldosterone at physiological levels induces rapid (<5 min) increases in intracellular protein kinase C (PKC) activity and a rise in calcium and pH in mineralocorticoid hormone target epithelia, such as distal colon and sweat gland. The end targets of these rapid responses in epithelia are Na+/H+ exchange and K+ channels. METHODS: The mouse cortical collecting duct (CCD) M-1 cell line was grown to confluency and loaded with Fura-2 for spectrofluorescence measurements of intracellular free calcium at 37 degrees C bathed in Krebs solution. RESULTS: Aldosterone (1 nmol/L) produced a rapid, transient peak increase in [Ca2+]i in M-1 cells. This effect was abolished upon removal of extracellular Ca2+, but was unaffected by pretreatment with spironolactone (10 micromol/L) or actinomycin D (10 micromol/L). However, pretreatment with the specific PKC inhibitor chelerythrine chloride (1 micromol/L) prevented the aldosterone-induced rise in [Ca2+]i. Dexamethasone, at a concentration 10,000-fold higher than aldosterone (10 micromol/L), also produced a transient increase in [Ca2+]i, but this response was significantly smaller than that of aldosterone. In contrast, hydrocortisone had no effect on [Ca2+]i at either nmol/L or micromol/L concentrations. Both of the sex steroids, 17beta-estradiol (10 nmol/L) and progesterone (10 nmol/L), induced protein kinase C-dependent increases in [Ca2+]i. CONCLUSIONS: Aldosterone and sex steroid hormones activate intracellular calcium signaling in CCD cells via a nongenomic PKC-dependent pathway, which may have important implications for renal transport. (+info)
Activation of the amiloride-sensitive epithelial sodium channel by the serine protease mCAP1 expressed in a mouse cortical collecting duct cell line.
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This study examines whether serine proteases can activate the amiloride-sensitive sodium channel (ENaC) in mammalian kidney epithelial cells. The transepithelial sodium transport assessed by amiloride-sensitive short-circuit current appears to be sensitive to aprotinin, a protease inhibitor in a mouse cortical collecting duct cell line (mpkCCD(c14)). This result indicated that serine proteases may be implicated in the regulation of ENaC-mediated sodium transport. Using degenerated oligonucleotides to a previously isolated serine protease from Xenopus, xCAP1 (channel activating protease), a novel full-length serine protease (mCAP1), has been isolated and characterized. RNA analysis showed a broad pattern of expression in tissues (kidney, lung, colon, and salivary glands) expressing ENaC. Reverse transcription-PCR experiments also showed that mCAP1 was abundantly expressed in proximal tubule cells and was also expressed in intact and cultured collecting duct cells. Coexpression of the Xenopus, rat, or human alpha-, beta-, and gamma-ENaC subunits in Xenopus oocytes also showed that mCAP1 induces a significant increase in ENaC-mediated current accompanied by a decrease of channel molecules at the cell surface. It is proposed that this novel mouse channel activating protease may act as a regulator of ENaC within the kidney. (+info)
Protein-tyrosine phosphatase reduces the number of apical small conductance K+ channels in the rat cortical collecting duct.
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Previous studies have demonstrated that an increase in the activity of protein-tyrosine kinase (PTK) is involved in the down-regulation of the activity of apical small conductance K(+) (SK) channels in the cortical collecting duct (CCD) from rats on a K(+)-deficient diet (). We used the patch clamp technique to investigate the role of protein-tyrosine phosphatase (PTP) in the regulation of the activity of SK channels in the CCD from rats on a high K(+) diet. Western blot analysis indicated that PTP-1D is expressed in the renal cortex. Application of 1 microm phenylarsine oxide (PAO) or 1 mm benzylphosphonic acid, agents that inhibit PTP, reversibly reduced channel activity by 95%. Pretreatment of CCDs with PAO for 30 min decreased the mean NP(o) reversibly from control value 3.20 to 0.40. Addition of 1 microm herbimycin A, an inhibitor of PTK, had no significant effect on channel activity in the CCDs from rats on a high K(+) diet. However, herbimycin A abolished the inhibitory effect of PAO, indicating that the effect of PAO is the result of interaction between PTK and PTP. Addition of brefeldin A, an agent that blocks protein trafficking from Golgi complex to the membrane, had no effect on channel activity. Moreover, application of colchicine, a microtubule inhibitor, or paclitaxel, a microtubule stabilizer, had no effect on channel activity. In contrast, PAO still reduced channel activity in the presence of brefeldin A, colchicine, or paclitaxel. Furthermore, the effect of PAO on channel activity was absent when the tubules were bathed in 16% sucrose-containing bath solution or treated with concanavalin A. We conclude that PTP is involved in the regulation of the activity of SK channels and that inhibition of PTP may facilitate the internalization of the SK channels. (+info)
Normalization of hyperosmotic-induced inositol uptake by renal and endothelial cells is regulated by NF-kappaB.
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Hyperosmolarity is a stress factor that has been shown to cause an increase in the transcription of the Na(+)-dependent myo-inositol cotransporter (SMIT). However, regulation of the reversion of SMIT mRNA levels and transporter activity following removal of hyperosmotic stress is less understood. Previously we have shown that postinduction normalization of SMIT mRNA levels and myo-inositol accumulation following removal of hyperosmotic stress is inhibited by actinomycin D and cycloheximide, suggesting that normalization requires RNA transcription and protein synthesis. We now demonstrate that removal of hyperosmotic stress causes an activation of the transcription factor NF-kappaB in renal and endothelial cells. Inhibiting NF-kappaB activation with pyrrolidine dithiocarbamate (PD) blocks the normalization of SMIT mRNA levels and myo-inositol accumulation on removal of the cells from hyperosmotic medium. These studies demonstrate that the downregulation of the myo-inositol transporter following reversal of hyperosmotic induction is regulated via the activation of NF-kappaB. (+info)