Physiologic regulation of the epithelial sodium channel by phosphatidylinositides

PURPOSE OF REVIEW: Epithelial sodium channel (ENaC) activity is limiting for sodium reabsorption in the distal nephron. Humans regulate blood pressure by fine-tuning sodium balance through control of ENaC. ENaC dysfunction causes some hypertensive and renal salt wasting diseases. Thus, it is critical to understand the cellular mechanisms controlling ENaC activity. RECENT FINDINGS: ENaC is sensitive to phosphatidylinositol 4,5-bisphosphate (PIP2), the target of phospholipase C-mediated metabolism, and phosphatidylinositiol 3,4,5-trisphosphate (PIP3), the product of phosphatidylinositide 3-OH kinase (PI3-K). PIP2 is permissive for ENaC gating possibly interacting directly with the channel. Activation of distal nephron P2Y receptors tempers ENaC activity by promoting PIP2 metabolism. This is important because gene deletion of P2Y2 receptors causes hypertension associated with hyperactive ENaC. Aldosterone, the final hormone in a negative-feedback cascade activated by decreases in blood pressure, increases ENaC activity. PIP3 sits at a critical bifurcation in the aldosterone-signaling cascade, increasing ENaC open probability and number. PIP3-effectors mediate increases in ENaC number by suppressing channel retrieval. PIP3 binds ENaC, at a site distinct from that important to PIP2 regulation, to modulate directly open probability. SUMMARY: Phosphoinositides play key roles in physiologic control of ENaC and perhaps dysregulation plays a role in disease associated with abnormal renal sodium handling.     

Purinergic Inhibition of ENaC Produces Aldosterone Escape

The mechanisms underlying “aldosterone escape,” which refers to the excretion of sodium (Na⁺) during high Na⁺ intake despite inappropriately increased levels of mineralocorticoids, are incompletely understood. Because local purinergic tone in the aldosterone-sensitive distal nephron downregulates epithelial Na⁺ channel (ENaC) activity, we tested whether this mechanism mediates aldosterone escape. Here, urinary ATP concentration increased with dietary Na⁺ intake in mice. Physiologic concentrations of ATP decreased ENaC activity in a dosage-dependent manner. P2Y₂^−/− mice, which lack the purinergic receptor, had significantly less increased Na⁺ excretion than wild-type mice in response to high-Na⁺ intake. Exogenous deoxycorticosterone acetate and deletion of the P2Y₂ receptor each modestly increased the resistance of ENaC to changes in Na⁺ intake; together, they markedly increased resistance. Under the latter condition, ENaC could not respond to changes in Na⁺ intake. In contrast, as a result of aldosterone escape, wild-type mice had increased Na⁺ excretion in response to high-Na⁺ intake regardless of the presence of high deoxycorticosterone acetate. These data suggest that control of ENaC by purinergic signaling is necessary for aldosterone escape.Renal sodium (Na⁺) excretion influences BP by affecting systemic Na⁺ balance. Consequently, negative feedback in response to changes in Na⁺ balance, perceived as changes in effective circulating volume (ECV) and BP, control renal Na⁺ handling. The fine-tuning of Na⁺ balance occurs in the aldosterone-sensitive distal nephron (ASDN), including the connecting tubule (CNT) and the collecting duct (CD). Here, Na⁺ reabsorption is highest when ECV and BP are low. Activity of the epithelial Na⁺ channel (ENaC) is limiting for discretionary Na⁺ reabsorption across the ASDN.^1–7 As ECV declines, activity of the renin-angiotensin-aldosterone system (RAAS) increases with the mineralocorticoid aldosterone, stimulating channel activity to decrease Na⁺ excretion in correction of falling ECV. Aldosterone increases the number and activity of ENaC in the apical membrane.^8–11 Under normal conditions, the opposite is also true; aldosterone and, thus, ENaC activity decline in response to elevations in BP.The thiazide-sensitive Na-Cl co-transporter (NCC) in the distal convoluted tubule (DCT) is also a target of aldosterone with the mineralocorticoid increasing NCC activity and possibly transporter abundance in the apical membrane of DCT cells.^12–16 This increase in NCC activity decreases Na⁺ excretion. Thus, aldosterone promotes distal nephron Na⁺ reabsorption by increasing the activity of ENaC and NCC.The loss of negative-feedback regulation of renal Na⁺ excretion mediated by ENaC and NCC leads to hypertension. For instance, gain-of-function mutations in ENaC cause the channel to be hyperactive irrespective of mineralocorticoid status and systemic Na⁺ balance, thus leading to inappropriate Na⁺ retention and hypertension; conversely, loss-of-function mutations in ENaC can cause decreases in BP and renal salt wasting.^{2,3,5,7,17–19}The aldosterone-promoted decrease in renal Na⁺ excretion is overridden in some circumstances.^20,21 This disruption of aldosterone action is termed aldosterone escape and results in avid Na⁺ excretion during high Na⁺ intake despite elevated mineralocorticoid levels. Aldosterone escape seems to be a protective mechanism to allow appropriate response to positive Na⁺ balance despite inappropriate mineralocorticoid levels. Such escape is important clinically, for example, in primary aldosteronism, in which it may ameliorate, to some degree, the hypertensive effects of high circulating levels of aldosterone.^22–24 The mechanism allowing aldosterone escape is uncertain.Aldosterone escape is known to be dependent on increases in renal vascular perfusion pressure rather than systemic factors, including changes in the levels of circulating hormones, such as renin and aldosterone, or renal nerve activity.²⁵ This led to the idea that aldosterone escape is a manifestation of a pressure natriuresis response.^13,25,26 This is associated with declining Na⁺ reabsorption in the distal nephron despite elevated levels of aldosterone and occurs independent of changes in GFR.^26,27 Indeed, early micropuncture measurements demonstrated increases in urine flow and Na⁺ delivery to the CD during escape.^26,27 Details about the specific site(s) along the distal nephron involved in decreased Na⁺ reabsorption during aldosterone escape and the exact transport proteins involved in this escape, though, largely remain obscure.A study by Wang et al.¹³ revealed that during aldosterone escape, NCC levels in the DCT decrease. This response was selective because the levels of other apical membrane transport proteins, including ENaC, were unaffected. This finding supports the idea that NCC is, at least, one target for regulatory processes that mediate aldosterone escape, whereby decreasing NCC abundance facilitates Na⁺ excretion despite high levels of mineralocorticoid. The cellular mechanism underpinning declines in NCC levels during aldosterone escape is unknown. Similarly, it is unclear whether other transport proteins are involved in aldosterone escape because Na⁺ reabsorption is a manifestation not only of the number of transport proteins in the apical membrane but also of their activity.Because ENaC is critical to aldosterone regulation of Na⁺ excretion, particularly in response to changes in Na⁺ balance, evidenced by the hypertension associated with gain of ENaC function,^3,5,7 we sought to investigate the role of this channel in aldosterone escape. We recently demonstrated that local purinergic tone in the ASDN exerts paracrine downregulation of ENaC activity specifically by affecting channel open probability.²⁸ This paracrine pathway is physiologically relevant because mice lacking the purinergic receptor P2Y₂, responsible for the bulk of paracrine regulation in this system, have facilitated Na⁺ reabsorption in the distal nephron and mild increases in BP.²⁹ Increases in NKCC2 and ENaC activity contribute to this elevated BP.¹¹ It does not seem a coincidence that aldosterone and P2Y₂ signaling target the same final effector proteins, possibly allowing one to compensate for the loss of the other. This idea suggests that purinergic regulation of ENaC may contribute to aldosterone escape, a possibility supported by another recent finding from our laboratories: Elevated BP in P2Y₂^−/− mice is not salt sensitive in the presence of normal-feedback regulation by RAAS but becomes salt sensitive and associated with inappropriately active ENaC when negative-feedback regulation by aldosterone and local purinergic signaling both are disrupted.^11,29To test the role of ENaC and its regulation by mineralocorticoids and purinergic signaling in aldosterone escape and to understand better the mechanism underpinning escape, we quantified the actions of mineralocorticoid on renal Na⁺ excretion, ENaC activity, and urinary ATP levels in wild-type (WT) and P2Y₂^−/− mice stressed with different dietary Na⁺ regimens. We found that urinary [ATP] increases with dietary Na⁺ intake such that physiologic [ATP] decrease ENaC activity, resulting in increased Na⁺ excretion. Increased Na⁺ excretion is greater in WT compared with P2Y₂^−/− mice and associated with an inability of ENaC, particularly when mineralocorticoid is clamped at high levels, to respond appropriately to changes in dietary Na⁺ intake in the latter animals. Because ENaC activity normally is sensitive to changes in Na⁺ balance even when mineralocorticoids are clamped at high levels, these results show that control of ENaC by purinergic signaling is necessary for complete aldosterone escape, consistent with the loss of aldosterone-escape in P2Y₂^−/− mice and their pronounced hypertension relative to normal mice in the presence of elevated mineralocorticoids and high Na⁺ intake.     

Endoscopic therapy of dysplasia and early-stage cancers of the esophagus

Endoscopic treatments have become a viable alternative for some patients with early-stage esophageal neoplasia. Although esophagectomy remains the standard of care for high-grade dysplasia and superficial cancers, surgical morbidity and mortality may deter patients who are medically unfit or reluctant to undergo surgery. Photodynamic therapy (PDT) and endoscopic mucosal resection (EMR) are the best-studied nonsurgical approaches at present. PDT has been reported to eradicate high-grade dysplasia (HGD) and early Barrett's cancers at rates ranging from 75% to 100% and 17% to 100%, respectively, and a recent randomized controlled trial confirmed that PDT may prevent progression of HGD to cancer. Complete remission rates greater than 90% have also been reported with EMR and other mucosa-ablating interventions, although recurrence rates necessitate close endoscopic surveillance and retreatment in some patients. In addition to PDT and EMR, several emerging endoscopic treatment options for superficial esophageal neoplasia may provide attractive alternatives to surgery.     

Thalamic noradrenaline in Parkinson's disease: Deficits suggest role in motor and non‐motor symptoms

The thalamus occupies a pivotal position within the corticobasal ganglia‐cortical circuits. In Parkinson's disease (PD), the thalamus exhibits pathological neuronal discharge patterns, foremost increased bursting and oscillatory activity, which are thought to perturb the faithful transfer of basal ganglia impulse flow to the cortex. Analogous abnormal thalamic discharge patterns develop in animals with experimentally reduced thalamic noradrenaline; conversely, added to thalamic neuronal preparations, noradrenaline exhibits marked antioscillatory and antibursting activity. Our study is based on this experimentally established link between noradrenaline and the quality of thalamic neuronal discharges. We analyzed 14 thalamic nuclei from all functionally relevant territories of 9 patients with PD and 8 controls, and measured noradrenaline with high‐performance liquid chromatography with electrochemical detection. In PD, noradrenaline was profoundly reduced in all nuclei of the motor (pallidonigral and cerebellar) thalamus (ventroanterior: ?86%, P = .0011; ventrolateral oral: ?87%, P = .0010; ventrolateral caudal: ?89%, P = .0014): Also, marked noradrenaline losses, ranging from 68% to 91% of controls, were found in other thalamic territories, including associative, limbic and intralaminar regions; the primary sensory regions were only mildly affected. The marked noradrenergic deafferentiation of the thalamus discloses a strategically located noradrenergic component in the overall pathophysiology of PD, suggesting a role in the complex mechanisms involved with the genesis of the motor and non‐motor symptoms. Our study thus significantly contributes to the knowledge of the extrastriatal nondopaminergic mechanisms of PD with direct relevance to treatment of this disorder. © 2012 Movement Disorder Society     

Current applications of 3-d intraoperative navigation in craniomaxillofacial surgery: a retrospective clinical review

Intraoperative navigation is a tool that provides surgeons with real-time, interactive access to their patient's diagnostic imaging studies while in the operating room. This modality allows for anatomic localization and facilitates intraoperative planning and diagnosis. The application of intraoperative navigation to neurosurgery, otolaryngology, and orthopedic surgery has been well documented; however, only isolated reports have analyzed its potential in the field of craniomaxillofacial surgery. Advancements in 3-dimensional navigational systems have greatly improved the accuracy of the technology, further broadening its scope. In this article, we evaluate a series of 101 craniomaxillofacial cases in which intraoperative navigation was used. The most common application was for intraorbital cases, such as enophthalmos and acute orbital fracture repairs. Other applications included tumor resection, osteotomy design, pathology localization, and craniotomy design. The major limitations of this technology have been its cost and the fact that it cannot reliably be used for soft-tissue reconstruction currently.     

Should all endoscopically excised rectal polyps be tattooed? A plea for localization

Background

More than 5–8?% of endoscopically removed rectal polyps presumed to be benign contain invasive carcinoma. Tattooing has been advocated for follow-up localization of the resection site. Despite proven benefits, the authors propose that tattooing is not routinely performed when benign-appearing rectal polyps are endoscopically excised, thereby confounding management when invasive cancer is found. The secondary goal of the study was to determine the frequency of localization, polyp characteristics, and accuracy of predicting malignant potential at the authors’ institution.

Methods

All patients with rectal neoplasia discovered during endoscopic polypectomy from 1 January 2003 to 1 August 2010 were retrospectively identified from Temple University Hospital’s Tumor Registry. Demographic and clinical data were extracted from medical records including polyp size, gross appearance, pathology, resection margins, location based on preoperative colonoscopy, initial removal technique, tattoo performance, and ensuing procedures.

Results

During the study period, 49 patients had colonoscopic excision of presumed benign rectal polyps with ensuing diagnosis of neoplasia in the specimen. The malignant histology included adenocarcinoma (n?=?5), carcinoma in situ (n?=?21), carcinoid (n?=?22), and composite carcinoid (n?=?1). Only two polyps were tattooed at the initial polypectomy. Three polyps were “suspicious for malignancy.” None of the suspicious polyps were tattooed. One of the suspicious lesions was an adenocarcinoma, and the remaining two were benign. The distance from the anal verge was noted in only seven patients. The predominant excision technique was hot snare polypectomy (n?=?29). None of the incomplete polyp excisions for 15 patients were “suspicious for malignancy” or tattooed. Several strategies were used to manage incomplete resections including surveillance (40?%), repeat colonoscopic polypectomy (27?%), and surgery (33?%).

Conclusions

Most malignant rectal polyps are neither diagnosed nor tattooed at initial colonoscopy. Moreover, the distance of the polyp from the anal verge is rarely measured, and gross characteristics are not well described. Tattooing of all endoscopically excised rectal polypectomy sites would avoid confounding of subsequent identification and management.     

Different effects of P. gingivalis LPS and E. coli LPS on the expression of interleukin-6 in human gingival fibroblasts

Objective. Gingival fibroblasts (GFs) produce pro-inflammatory cytokines in response to stimulation with lipopolysaccharide (LPS) of Porphyromonas gingivalis, which is thought to be mediated by activation of toll-like receptors (TLR)2 and TLR4. The present study investigated the expression of interleukin (IL)-6, TLR2, and TLR4 in GFs of seven different donors upon stimulation with P. gingivalis LPS. The effects of P. gingivalis LPS were compared with those of TLR4 agonist Escherichia coli LPS and TLR2 agonist Pam3CSK4. Materials and methods. GFs were stimulated with P. gingivalis LPS, E. coli LPS or Pam3CSK4 and the expression of IL-6, TLR2 and TLR4 was measured by qPCR. The surface expression of TLR2 and TLR4 was measured by flow cytometry. Results. In GFs from three donors, P. gingivalis LPS and Pam3CSK4 induced a markedly lower increase in IL-6 expression than E. coli LPS. This was accompanied by significant down-regulation of the TLR2 and TLR4 expression. In GFs from another four donors, an increase in IL-6 expression upon stimulation with P. gingivalis LPS and Pam3CSK4 was similar or even higher than that induced by E. coli LPS. In GFs of these donors, all stimuli induced an up-regulation of both mRNA and protein expression of TLR2 and did not influence that of TLR4. Conclusions. This study suggests that P. gingivalis LPS and E. coli LPS differently regulate cytokine production in human gingival fibroblasts. Regulation of the expression level of TLR2 and TLR4 by periodontal pathogens might be an important factor controlling the inflammatory response in GFs.