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Peter R Smith

Publications and source records attributed to Peter R Smith.

14 recordsLinked to original sources
PubMed2006

Interregulation of proton-gated Na(+) channel 3 and cystic fibrosis transmembrane conductance regulator.

Proton-gated Na(+) channels (ASIC) are new members of the epithelial sodium channel/degenerin gene family. ASIC3 mRNA has been detected in the homogenate of pulmonary tissues. However, whether ASIC3 is expressed in the apical membranes of lung epithelial cells and whether it regulates cystic fibrosis transmembrane conductance regulator (CFTR) function are not known at the present time. Using reverse transcription-PCR, we found that the ASIC3 mRNA was expressed in the human airway mucosal gland (Calu-3) and human airway epithelial (16HBE14o) cells. Indirect immunofluorescence microscopy revealed that ASIC3 was co-segregated with CFTR in the apical membranes of Calu-3 cells. Proton-gated, amiloride-sensitive short circuit Na(+) currents were recorded across Calu-3 monolayers mounted in an Ussing chamber. In whole-cell patch clamp studies, activation of CFTR channels with cAMP reduced proton-gated Na(+) current in Calu-3 cells from -154 +/- 28 to -33 +/- 16 pA (n = 5, p < 0.05) at -100 mV. On the other hand, cAMP-activated CFTR activity was significantly inhibited following constitutive activation of putative ASIC3 at pH 6.0. Immunoassays showed that both ASIC3 and CFTR proteins were expressed and co-immunoprecipitated mutually in Calu-3 cells. Similar results were obtained in human embryonic kidney 293T cells following transient co-transfection of ASIC3 and CFTR. Our results indicate that putative CFTR and ASIC3 channels functionally interact with each other, possibly via an intermolecular association. Because acidic luminal fluid in the cystic fibrosis airway and lung tends to stimulate ASIC3 channel expression and activity, the interaction of ASIC3 and CFTR may contribute to defective salt and fluid transepithelial transport in the cystic fibrotic pulmonary system.

Acid Sensing Ion Channels↗
PubMed2006

Delta-subunit confers novel biophysical features to alpha beta gamma-human epithelial sodium channel (ENaC) via a physical interaction.

Native amiloride-sensitive Na+ channels exhibit a variety of biophysical properties, including variable sensitivities to amiloride, different ion selectivities, and diverse unitary conductances. The molecular basis of these differences has not been elucidated. We tested the hypothesis that co-expression of delta-epithelial sodium channel (ENaC) underlies, at least in part, the multiplicity of amiloride-sensitive Na+ conductances in epithelial cells. For example, the delta-subunit may form multimeric channels with alpha beta gamma-ENaC. Reverse transcription-PCR revealed that delta-ENaC is co-expressed with alpha beta gamma-subunits in cultured human lung (H441 and A549), pancreatic (CFPAC), and colonic epithelial cells (Caco-2). Indirect immunofluorescence microscopy revealed that delta-ENaC is co-expressed with alpha-, beta-, and gamma-ENaC in H441 cells at the protein level. Measurement of current-voltage that cation selectivity ratios for the revealed relationships Na+/Li+/K+/Cs+/Ca2+/Mg2+, the apparent dissociation constant (Ki) for amiloride, and unitary conductances for delta alpha beta gamma-ENaC differed from those of both alpha beta gamma- and delta beta gamma-ENaC (n = 6). The contribution of the delta subunit to P(Li)/P(Na) ratio and unitary Na+ conductance under bi-ionic conditions depended on the injected cRNA concentration. In addition, the EC50 for proton activation, mean open and closed times, and the self-inhibition time of delta alpha beta gamma-ENaC differed from those of alpha beta gamma- and delta beta gamma-ENaC. Co-immunoprecipitation of delta-ENaC with alpha- and gamma-subunits in H441 and transfected COS-7 cells suggests an interaction among these proteins. We, therefore, concluded that the interactions of delta-ENaC with other subunits could account for heterogeneity of native epithelial channels.

Amiloride↗
PubMed2006

A diagnostic and management dilemma: combined paraneoplastic myasthenia gravis and Lambert-Eaton myasthenic syndrome presenting as acute respiratory failure.

BACKGROUND: Neuromuscular junction disorders are usually categorized as either presynaptic or postsynaptic. The most frequently encountered disorder of the postsynaptic neuromuscular junction is acquired myasthenia gravis. Lambert-Eaton myasthenic syndrome is a well-known prototype of the presynaptic autoimmune disorders of neuromuscular transmission. These major disorders of neuromuscular transmission are relatively common and distinctly recognized, but co-occurrence of these disorders (overlap myasthenic syndrome) is rare and has so far attracted little attention. REVIEW SUMMARY: This report describes a patient with acquired myasthenia gravis and immunologic coexistence of Lambert-Eaton myasthenic syndrome (overlap myasthenic syndrome) in association with abdominal/uterine leiomyosarcoma. The patient presented with acute respiratory failure, making identification and management of her illness challenging. A general overview of the complexities associated with overlap between myasthenia gravis and Lambert-Eaton myasthenic syndrome is provided and this patient's complicated clinical course and response to therapy are discussed. CONCLUSION: To our knowledge, this is the first report of overlap myasthenic syndrome in conjunction with abdominal leiomyosarcoma. The immunologic coexistence of acquired myasthenia gravis and Lambert-Eaton myasthenic syndrome in a patient with a malignant smooth-muscle tumor is intriguing and suggests that a common paraneoplastic process targeting 2 different onconeural antigens was the underlying pathogenic mechanism in this patient.

Aged↗
PubMed2005

The carboxyl terminus of the alpha-subunit of the amiloride-sensitive epithelial sodium channel binds to F-actin.

The activity of the amiloride-sensitive epithelial sodium channel (ENaC) is modulated by F-actin. However, it is unknown if there is a direct interaction between alpha-ENaC and actin. We have investigated the hypothesis that the actin cytoskeleton directly binds to the carboxyl terminus of alpha-ENaC using a combination of confocal microscopy, co-immunoprecipitation, and protein binding studies. Confocal microscopy of Madin-Darby canine kidney cell monolayers stably transfected with wild type, rat isoforms of alpha-, beta-, and gamma-ENaC revealed co-localization of alpha-ENaC with the cortical F-actin cytoskeleton both at the apical membrane and within the subapical cytoplasm. F-actin was found to co-immunoprecipitate with alpha-ENaC from whole cell lysates of this cell line. Gel overlay assays demonstrated that F-actin specifically binds to the carboxyl terminus of alpha-ENaC. A direct interaction between F-actin and the COOH terminus of alpha-ENaC was further corroborated by F-actin co-sedimentation studies. This is the first study to report a direct and specific biochemical interaction between F-actin and ENaC.

Actins↗
PubMed2005

Complex character analysis of heart rate variability following brain asphyxia.

In the present study Renyi entropy and L-Z complexity were used to characterize heart rate variability (HRV) of rats that were suffered from brain asphyxia and ischemia. Two groups of rats were studied: (a) rats (n=5) injected with NAALADase inhibitor, 2-PMPA, which has been proven neuroprotective in asphyxia injury and (b) control subjects (n=5) without medication. Renyi entropy and L-Z complexity of the R-R intervals (RRI) at different experiment stages were investigated in the two groups. The results show that both measures indicate less injury and better recovery in the drug injection group. The dynamic change of 90 min RRI signal after the asphyxia was investigated. The sudden reduction of the two parameters shows their sensitivity to the asphyxia insult.

Animals↗
PubMed2005

Protein kinase C-alpha regulation of gallbladder Na+ transport becomes progressively more dysfunctional during gallstone formation.

Gallbladder Na+ absorption and biliary Ca2+ are both increased during gallstone formation and may promote cholesterol nucleation. Na+/H+ exchange (NHE) is a major pathway for gallbladder Na+ transport. Ca2+-dependent second messengers, including protein kinase C (PKC), inhibit basal gallbladder Na+ transport. Multiple PKC isoforms with species- and tissue-specific expression have been reported. In this study we sought to characterize Ca2+-dependent PKC isoforms in gallbladder and to examine their roles in Na+ transport during gallstone formation. Gallbladders were harvested from prairie dogs fed either nonlithogenic chow or 1.2% cholesterol-enriched diet for varying periods to induce various stages of gallstone formation. PKC was activated with the use of phorboldibutyrate, and we assessed gallbladder NHE regulation by measuring unidirectional Na+ flux and dimethylamiloride-inhibitable 22Na+ uptake. We measured gallbladder PKC activity with the use of histone III-S phosphorylation and used Gö 6976 to determine PKC-alpha contributions. Gallbladder PKC isoform messenger RNA and protein expression were examined with the use of Northern- and Western-blot analysis, respectively. Prairie dog and human gallbladder expresses PKC-alpha, betaII, and delta isoforms. The PKC activation significantly decreased gallbladder J(Na)(ms) and reduced baseline 22Na+ uptake by inhibiting NHE. PKC-alpha mediated roughly 42% of total PKC activity under basal conditions. PKC-alpha regulates basal gallbladder Na+ transport by way of stimulation of NHE isoform NHE-2 and inhibition of isoform NHE-3. PKC-alpha blockade reversed PKC-induced inhibition of J(Na)(ms) and 22Na+ uptake by about 45% in controls but was progressively less effective during gallstone formation. PKC-alpha contribution to total PKC activity is progressively reduced, whereas expression of PKC-alpha mRNA, and protein increases significantly during gallstone formation. We conclude that PKC-alpha regulation of gallbladder NHE becomes progressively more dysfunctional and may in part account for the increased Na+ absorption observed during gallstone formation.

Animals↗
PubMed2004

Characterization of Na+-permeable cation channels in LLC-PK1 renal epithelial cells.

In this study, the presence of Na(+)-permeable cation channels was determined and characterized in LLC-PK1 cells, a renal tubular epithelial cell line with proximal tubule characteristics derived from pig kidney. Patch-clamp analysis under cell-attached conditions indicated the presence of spontaneously active Na(+)-permeable cation channels. The channels displayed nonrectifying single channel conductance of 11 pS, substates, and an approximately 3:1 Na(+)/K(+) permeability-selectivity ratio. The Na(+)-permeable cation channels were inhibited by pertussis toxin and reactivated by G protein agonists. Cation channel activity was observed in quiescent cell-attached patches after vasopressin stimulation. The addition of protein kinase A and ATP to excised patches also induced Na(+) channel activity. Spontaneous and vasopressin-induced Na(+) channel activity were inhibited by extracellular amiloride. To begin assessing potential molecular candidates for this cation channel, both reverse transcription-PCR and immunocytochemical analyses were conducted in LLC-PK1 cells. Expression of porcine orthologs of the alphaENaC and ApxL genes were found in LLC-PK1 cells. The expression of both gene products was confirmed by immunocytochemical analysis. Although alphaENaC labeling was mostly intracellular, ApxL labeled to both the apical membrane and cytoplasmic compartments of subconfluent LLC-PK1 cells. Vasopressin stimulation had no effect on alphaENaC immunolabeling but modified the cellular distribution of ApxL, consistent with an increased membrane-associated ApxL. The data indicate that proximal tubular LLC-PK1 renal epithelial cells express amiloride-sensitive, Na(+)-permeable cation channels, which are regulated by the cAMP pathway, and G proteins. This channel activity may implicate previously reported epithelial channel proteins, although this will require further experimentation. The evidence provides new clues as to potentially relevant Na(+) transport mechanisms in the mammalian proximal nephron.

Adenosine Triphosphate↗
PubMed2004

Cardiac troponin I predicts myocardial dysfunction and adverse outcome in septic shock.

OBJECTIVE: The objectives of this study were to determine myocardial injury in patients with septic shock by measuring serum cardiac troponin I (cTnI), to evaluate relationship between elevated cTnI and myocardial dysfunction and to determine if cTnI is a predictor of outcome in these patients. METHODS: Thirty-seven consecutive patients with septic shock were included in the study. Serum cTnI was measured at study entry and after 24 and 48 h. Transthoracic echocardiogram, electrocardiogram and regular biochemical and hemodynamic assessments were performed. RESULTS: Sixteen (43%) patients had elevated serum cTnI. These patients had higher need for inotropic/vasopressor support (94% vs. 53%, p=0.018), higher APACHE II score (28 vs. 20, p=0.004), higher incidence of regional wall motion abnormalities on echocardiography (56% vs. 6%, p=0.002), lower ejection fraction (46% vs. 62%, p=0.04) and higher mortality (56% vs. 24%, p=0.04) compared to normal cTnI patients. By multiple logistic regression analysis, serum cTnI and APACHE II score were independent predictor of death and length of stay in intensive care unit. Serum cTnI, APACHE II score, anion gap and serum lactate were independent predictor of need for inotropic/vasopressor support. Receiver-operating characteristics of serum cTnI as a predictor of death in septic shock were significant. The elevated serum level of cTnI correlated with the lower left ventricular ejection fraction (p<0.001). CONCLUSIONS: Myocardial injury can be determined in patients with septic shock by serum cTnI. Serum cTnI concentration correlates with myocardial dysfunction in septic shock. High serum cTnI predicts increased severity of sepsis and higher mortality. A close monitoring of patients with septic shock and elevated levels cTnI is warranted.

Aged↗
PubMed2004

Prognostic factors in lactic acidosis syndrome caused by nucleoside reverse transcriptase inhibitors: report of eight cases and review of the literature.

We conducted a retrospective study to identify prognostic factors in the lactic acidosis syndrome (LAS) caused by nucleoside reverse transcriptase inhibitors (NRTIs) in patients with HIV/AIDS. Fifty-eight cases of LAS were included in our analysis, 8 from our hospital spanning the years 1992-2002, and 50 reported in the English language literature from 1986 through 2002. Peak venous lactate level was the best predictor of mortality. Zidovudine was associated with higher lactate levels and higher mortality than stavudine and lamuvidine. Mortality declined progressively after 1986 when the first cases of NRTI-related LAS were described. Increased mortality with zidovudine in this study appears due in part to its greater use prior to 1990 when LAS was not widely recognized as a potential complication of NRTI therapy.

Acidosis, Lactic↗
PubMed2002

Origins of follicular cells and ontogeny of steroidogenesis in ovine fetal ovaries.

Using fetal sheep as the experimental model, we have elucidated some of the key events that culminate in the formation of primordial follicles. A special effort was made to determine the source of the somatic cells that ultimately become granulosa cells of primordial follicles. Between gestational days 38-100: (1) light and electron microscopy was used to characterize changes in ovarian histoarchitecture; (2) incorporation of BrdU was used to identify populations of proliferating cells within fetal ovaries before, during and after, follicular formation; and (3) in situ hybridisation was used to determine the cell-specific and temporal patterns of expression of mRNAs encoding for selected steroidogenic enzymes. At day 38 somatic (pregranulosa) cells were in contact with oogonia and easily distinguished from endothelial and mesenchymal cells. Between days 38 and 45, pregranulosa cell-oogonia complexes progressively coalesced to form 'tube-like' structures referred to as ovigerous cords. These cords consisted of pregranulosa cells and oogonia arranged such that pregranulosa cells formed the outer wall of the cords. Ovigerous cords were avascular, enveloped in a prominent basal lamina, open-ended where they interfaced with the ovarian surface epithelium, and formed a separate compartment whereby oogonia/oocytes were segregated from the surrounding stroma and vasculature until the time of follicular formation. The structural integrity of ovigerous cords was maintained through day 75, at which time primordial follicles (type 1 and type 1a) first emerged from the cords at the interface of the cortex and medulla. On the basis of the sequential structural changes that occurred during the differentiation and development of fetal ovaries and location of proliferating cells identified by the incorporation of BrdU, we conclude that the majority of the granulosa cells in primordial follicles are derived from mesothelial cells originating from the ovarian surface epithelium. In addition, from the cell-specific distribution and temporal pattern of expression of mRNAs for key steroidogenic enzymes we hypothesize that steroid hormones may play a pivotal paracrine/autocrine role in the formation and/or function of ovigerous cords as well as the development of the ovarian vascular network.

Animals↗
PubMed2002

Lovastatin alters biliary lipid composition and dissolves gallstones: a long-term study in prairie dogs.

Lovastatin, an inhibitor of the rate-limiting enzyme in cholesterol biosynthesis, is widely used to treat hypercholesterolemia. We investigated the long-term effects of lovastatin alone and in combination with ursodeoxycholic acid on biliary lipid composition and gallstone dissolution. Forty-two prairie dogs were fed 1.2% cholesterol diet for 5 weeks, and cholecystectomy was performed on 6 animals to confirm gallstones. The remaining animals were maintained on a 0.4% cholesterol diet and were randomized to receive placebo, lovastatin (3.3 mg/g diet), ursodeoxycholic acid (10 mg/g), or combination of both drugs. After 10 weeks, animals underwent cholecystectomy. Dissolution response to therapy was determined, and serum and biliary lipids were measured. All treatment groups had significant reductions in serum cholesterol. Lovastatin treatment reduced both hepatic and gallbladder bile cholesterol, altered bile acid composition, and induced a 79% total response compared to placebo. Although ursodeoxycholic treatment induced a 44% response, long-term combination treatment elevated both gallbladder bile cholesterol and calcium and failed to produce an augmented response. These data suggest that lovastatin therapy alone may promote gallstone dissolution in humans.

Animals↗
PubMed2002

HIV infection is a risk factor for venous thromboembolism.

The reported incidence of venous thromboembolism (VTE) in patients with human immunodeficiency virus (HIV) infection has ranged from 0.25 to 0.96% in clinical studies, but up to 17% at autopsy. A preliminary analysis at our hospital suggested that the frequency of VTE among HIV-positive individuals might be higher than previously reported. To further evaluate this issue, we performed a retrospective study of patients with a diagnosis of VTE and/or HIV infection discharged from our hospital between July 1, 1998 and June 30, 1999. A total of 13,496 patients were discharged during the year of the study. There were 244 patients with VTE and 362 who were HIV-positive. Ten of the 244 patients with VTE were HIV-positive (4.1%). The frequency of VTE among HIV-positive individuals was 10/362 (2.8%) compared to 234/13134 (1.8%) in the non-HIV-positive group, but the difference is not statistically significant. However, in patients under age 50, the frequencies were significantly different: 10/302 (3.31%) versus 35/6594 (0.53%), respectively (p < 0.0001). The frequency of VTE in HIV-positive patients less than 50 years old (3.31%) was greater than in HIV-positive patients over 50 years of age (0/60), but the difference did not reach statistical significance. In contrast, in the non-HIV-positive group, VTE was significantly more frequent in those 50 and older compared to younger patients (3.04% versus 0.53%, p = 0.0001). Statistical analysis indicated that the direction of association between age and diagnosis of VTE differed for HIV-positive patients versus non-HIV-positive patients. Our results suggest that HIV-positive patients under age 50 are at increased risk for VTE compared with non-HIV-positive individuals.

Adolescent↗
PubMed2002

Diagnostic tests for CAP: current approaches and future perspectives.

Diagnostic testing in community-acquired pneumonia (CAP) serves three functions: firstly, to confirm the presence of CAP; secondly, to assess severity; and thirdly, to identify the causative pathogen. Available techniques are satisfactory to fulfill the first two roles but are seriously inadequate as regards the third. Accordingly, antibiotic therapy for CAP must be empirical, at least initially. This article reviews current diagnostic methods and provides suggestions for appropriate use of diagnostic tests in CAP. The shortcomings of the available methods for microbiologic diagnosis are discussed. The potential for PCR to become the much sought after 'ideal' test for microbiologic diagnosis in CAP is explored.

Blood↗