Aims/hypothesis: The α subunit of the amiloride-sensitive epithelial sodium channel (α ENaC) is critical for the expression of functional channels. In humans and rats, non functional alternatively spliced forms of α ENaC have been proposed to act as negative regulatory components for ENaC. The purpose of this study was to examine the presence and consequently investigate the mRNA expression levels of alternatively spliced forms of α ENaC in kidney cortex of Dahl salt-resistant rats (R) versus Dahl salt-sensitive rats (S) on high salt and normal diets. Methods:Using quantitative RT-PCR strategy, we examined the mRNA expression levels of previously reported α ENaC-a and -b alternatively spliced forms in kidney cortex of Dahl S and R rats on normal and four-week high salt diet and compared their corresponding abundance to wildtype α ENaC mRNA levels. We identified 2 novel non-coding Cterminus spliced forms and examined their mRNA expression in Dahl R versus S rat kidney cortex. We also tested the presence of five previously reported lung-specific α ENaC spliced forms in Dahl rat kidney cortex (CK479583, CK475461, CK364785, CK475819, and CB690980).Results: Previously reported α ENaC-a and -b alternatively spliced forms are present in Dahl rat kidney cortex and are significantly higher in Dahl R versus S rats (P < 0.05). Four-week high salt diet significantly increases α ENaC-b (P < 0.05), but not α ENaC-a transcript abundance in Dahl R, but not S rats. Two non-coding α ENaC spliced forms -c and -d are newly identified in the present study, whose levels are comparable in Dahl R and S rats. Compared to α ENaC-wt, α ENaC-a, -c and -d are low abundance transcripts (4 +/-2, 110 +/-20, and 10 +/-2 fold less respectively), in contrast to α ENaC-b abundance that exceeds α ENaC-wt by 32 +/-3 fold. We could not identify any of the five previously reported lung-specific α ENaC spliced forms (CK479583, CK475461, CK364785, CK475819, and CB690980) in Dahl rat kidney cortex.Conclusion/interpretation: α ENaC alternative splicing might regulate α ENaC by the formation of coding RNA species (α ENaC-a and -b) and non-coding RNA species (α ENaC-c and -d). α ENaC-a and -b mRNA levels are significantly higher in Dahl R versus S rats. Additionally, α ENaC-b is a salt-sensitive transcript whose levels are significantly higher 4-weeks post high salt diet compared to normal salt diet in Dahl R rats. Among the four α ENaC transcripts (-a, -b, -c and -d), α ENaC-b is a predominant transcript that exceeds α ENaC-wt abundance by ~32 fold. α ENaC-a and -b spliced forms, particularly, α ENaC-b, might potentially act as dominant negative proteins for ENaC activity, thereby rescuing Dahl R rats from developing salt-sensitive hypertension on high salt diet. On the other hand, non-coding α ENaC-c and -d might assist alternative splicing, facilitate RNA processing, or regulate α ENaC as well as each other.

Insulin resistance has been extensively investigated during the past decade because of its proposed role in initiating a cluster of cardiovascular risk factors including hypertension. Insulin resistance is an inherited genetic trait that precedes hypertension in Dahl salt-sensitive (S) rats, and is not present in Dahl salt-resistant (R) rats. Owing to the co-existence of insulin resistance and salt sensitivity of blood pressure in Dahl S, but not R rats, Dahl S rats are used to elucidate the role of dietary salt as a potential link in exacerbating both phenotypes (insulin resistance and salt sensitivity). In light of available data, examining the impact of dietary salt on insulin resistance in Dahl S rats in terms of salt concentration and duration of exposure helps answer the following question: What percentage of dietary salt and for what duration of exposure would we expect an enhanced insulin resistance in Dahl S rats? This commentary gathers all available research done on insulin resistance in Dahl S rats in an attempt to unravel dietary salt contribution to insulin resistance in Dahl S rats. CommentaryOver the past decades, insulin resistance and compensatory hyperinsulinemia have been proposed as the common disorder underlying a constellation of risk factors (hypertension, diabetes, and hypertriglyceridemia) referred to as the metabolic syndrome or syndrome X [1]. Insulin resistance and salt-sensitivity of blood pressure coexist in Dahl salt-sensitive (S), but not salt-resistant (R) rats which in turn raises the following questions: a) Does insulin resistance in Dahl S rats predict their salt-sensitivity? b) Is insulin resistance in Dahl S rats a genetic abnormality or is it a metabolic consequence of salt-induced hypertension in this model? c) Does salt play a role in enhancing insulin resistance in Dahl S rats as it does with hypertension in this model? d) Does salt exert its action at a particular concentration and duration of exposure?To be able to address the above questions, Dahl S rats have been examined at various stages (weanlings and fully developed hypertensive rats) and with various dietary salt regimens (low, medium and high salt diet for durations of 1 to 4 weeks and up to 3 months of sodium loading). Weanling Dahl S rats when examined before developing hypertension and prior to sodium loading were found to be significantly insulin resistant as evidenced by their higher plasma insulin levels compared to weanling Dahl R rats [2]. This implies that insulin resistance in Dahl S rats is a genetic trait that is inherited and manifested prior to salt-sensitive hypertension in this model. When fully hypertensive Dahl S rats were examined (Dahl S rats become spontaneously hypertensive after 2-3 months [2], but can develop stable hypertension after 4 weeks of 8% NaCl [3], they remained insulin resistant compared to Dahl R rats [2]. Insulin resistance in hypertensive Dahl S

The epithelial sodium channel [ENaC] is critical for the maintenance of sodium balance, extracellular fluid volume and long term blood pressure control. Monogenic disorders causing ENaC hyperactivity have led to a severe form of hereditary hypertension in humans, known as Liddle's syndrome. Similarly, in animal models, ENaC hyperactivity has been well documented in kidneys of salt-sensitive [S] Dahl rats [a genetic model of salt-sensitive hypertension] versus their normotensive control [Dahl salt-resistant [R] rats]. The purpose of the present review is to highlight the differential regulation of ENaC in kidneys of Dahl S versus R rats. A systematic overview of the putative role of alternative splicing of the main α subunit of ENaC [α ENaC] in modulating ENaC expression in kidneys of Dahl rats will be discussed. Finally, a better understanding of the meaningful contribution of ENaC in the pathogenesis of salt-sensitive hypertension will be achieved upon completion of this review. Salt-sensitive hypertensionOver one-fifth of Canadian adults are diagnosed with hypertension http://www.statcan.com and over 50% of primary hypertensive patients are salt-sensitive [1]. Despite the fact that hypertension is the primary risk factor for stroke and heart disease, and has been labeled by the "silent killer disease", yet 42% of Canadians are still unaware of their increased blood pressures http:// www.heartandstroke.com/site/c.ikIQLcMWJtE/ b.3484023/.These above statistics, combined with the realization that salt-sensitive hypertension exacerbates mortality rates [2], worsens manifestations of target organ damage [3,4] and is a common finding in aging populations, emphasize the importance of identifying novel targets for prevention and treatment of salt-sensitive hypertension.The major contributor to the pathogenesis of salt-sensitive hypertension is dietary salt [5]. Dietary sodium, in turn, has sodium chloride [NaCl] as its major constituent. The sodium ion [Na + ] is transported into the superficial cells of several organs (see below) primarily via the amiloridesensitive Epithelial Sodium Channel [ENaC]. Owing to the fact that inadequate Na + excretion is a risk factor for hypertension, ENaC represents an attractive therapeutic target to study in salt-sensitive hypertension and α ENaC regulation by alternative splicing will be the focus of the present review.

Cervical cancer is considered a common yet preventable cause of death in women. In this report, we studied the role of the NF-kappaB gene family in HeLa human cervical cancer cells, using the Xrel3 c-Rel homologue of Xenopus laevis. The expression of Xrel3/c-Rel slowed cell growth 6-fold, consistent with an upregulated expression of the cell cycle inhibitor p21. The activated PARP apoptosis effector was significantly increased (P<0.01). Based on cell viability assays Xrel3 provided an anti-apoptotic effect in 1 microM cisplatin, and this was associated with significantly lower levels of the apoptotic proteins Bax and MDM-2 (P<0.05). Furthermore, there was a 3-fold drop in the level of the tumor suppressor protein p53. In 5 microM cisplatin, expression of HeLa Xrel3 enhanced apoptosis by significantly increasing the expression of the apoptotic proteins Bax and MDM-2 (P<0.05). However, the tumor suppressor protein p53 showed a significant decrease (P<0.05) relative to the control. Thus, c-Rel/NF-kappaB may potentially be of clinical significance, especially in tumors exhibiting resistance to high-level chemotherapy.

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