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PubMed1996

Cloning and functional characterization of a system ASC-like Na+-dependent neutral amino acid transporter.

A cDNA was isolated from mouse testis which encodes a Na+-dependent neutral amino acid transporter. The encoded protein, designated ASCT2, showed amino acid sequence similarity to the mammalian glutamate transporters (40-44% identity), Na+-dependent neutral amino acid transporter ASCT1 (57% identity; Arriza, J. L., Kavanaugh, M. P., Fairman, W. A., Wu, Y.-N., Murdoch, G. H., North, R. A., and Amara, S. G.(1993) J. Biol. Chem. 268, 15329-15332; Shafqat, S., Tamarappoo, B. K., Kilberg, M. S., Puranam, R. S., McNamara, J. O., Guadano-Ferraz, A., and Fremeau, T., Jr. (1993) J. Biol. Chem. 268, 15351-15355) and a mouse adipocyte differentiation-associated gene product AAAT (94% identity; Liao, K., and Lane, D.(1995) Biochem. Biophys. Res. Commun. 208, 1008-1015). When expressed in Xenopus laevis oocytes, ASCT2 exhibited Na+-dependent uptakes of neutral amino acids such as L-alanine, L-serine, L-threonine, L-cysteine, and L-glutamine at high affinity with Km values around 20 microM. L-Methionine, L-leucine, L-glycine, and L-valine were also transported by ASCT2 but with lower affinity. The substrate selectivity of ASCT2 was typical of amino acid transport system ASC, which prefers neutral amino acids without bulky or branched side chains. ASCT2 also transported L-glutamate at low affinity (Km = 1.6 mM). L-Glutamate transport was enhanced by lowering extracellular pH, suggesting that L-glutamate was transported as protonated form. In contrast to electrogenic transport of glutamate transporters and the other ASC isoform ASCT1, ASCT2-mediated amino acid transport was electroneutral. Na+ dependence of L-alanine uptake fits to the Michaelis-Menten equation, suggesting a single Na+ cotransported with one amino acid, which was distinct from glutamate transporters coupled to two Na+. Northern blot hybridization revealed that ASCT2 was mainly expressed in kidney, large intestine, lung, skeletal muscle, testis, and adipose tissue. Functional characterization of ASCT2 provided fruitful information on the properties of substrate binding sites and the mechanisms of transport of Na+-dependent neutral and acidic amino acid transporter family, which would facilitate the structure-function analyses based on the comparison of the primary structures of ASCT2 and the other members of the family.

Amino Acid Sequence↗
PubMed2004

PYNOD, a novel Apaf-1/CED4-like protein is an inhibitor of ASC and caspase-1.

Recently, a large subfamily of nucleotide-binding and oligomerization domain-containing proteins that have an N-terminal pyrin-like domain and C-terminal leucine-rich repeats has been described. In this study, we identified PYNOD, a novel member of this family that lacks the leucine-rich repeats. We found that human PYNOD mRNA is expressed in various tissues and at high levels in heart, skeletal muscle and brain. It is also expressed in various cell lines, including haematopoietic cell lines. PYNOD oligomerizes and binds to ASC, an adaptor protein that plays a role in apoptotic and inflammatory signal transduction, and to caspase-1 and IL-1beta. PYNOD inhibits apoptosis-associated speck-like protein containing a CARD (ASC)-mediated NF-kappaB activation and apoptosis, and caspase-1-mediated IL-1beta maturation, and it does so in the presence and absence of constitutively active mutants of CARD12 and PYPAF1, which are enhancers of these processes. Thus, PYNOD is a novel regulator of apoptosis and inflammation.

Adaptor Proteins, Signal Transducing↗
PubMed1993

ASC system activity is altered by development of cell polarity in trophoblast from human placenta.

Pathways of neutral amino acid uptake were investigated in vitro during differentiation of primary cultures of trophoblast isolated from full-term human placentas and a clone (b30) of the BeWo cell line. Inhibition of initial alanine (0.1 microM) uptake by 2-(methylamino)isobutyric acid and unlabeled alanine revealed two Na(+)-dependent systems and one Na(+)-independent transporter. Characterization of these transporters, by selective inhibition, suggested system A, ASC, and L-like transporters. Concomitant with formation of microvillous membrane and syncytium, system ASC activity decreased from 16.1 +/- 2.8 pmol.mg DNA-1.min-1 at 24 h to 2.4 +/- 1.1 pmol.mg DNA-1.min-1 at 72 h. Na(+)-independent alanine uptake increased from 6.0 +/- 2.0 to 12.9 +/- 0.9 pmol.mg DNA-1.min-1 at 24 and 72 h, respectively. Similarly, alpha-(methylamino)isobutyric acid-insensitive, Na(+)-dependent activity in b30 cells (100 microM alanine) decreased from 6.5 +/- 1.6 to 1.2 +/- 1.2 nmol.mg DNA-1.min-1 for control and forskolin-treated cells, respectively. We conclude that membrane specialization accompanying fusion and differentiation of the cytotrophoblast to form syncytiotrophoblast results in a polarization of neutral amino acid transport systems.

Alanine↗
PubMed2001

Electrogenic Na(+)-dependent L-alanine transport in the lizard duodenum. Involvement of systems A and ASC.

L-Alanine transport across the isolated duodenal mucosa of the lizard Gallotia galloti has been studied in Ussing chambers under short-circuit conditions. Net L-alanine fluxes, transepithelial potential difference (PD), and short-circuit current (Isc) showed concentration-dependent relationships. Na(+)-dependent L-alanine transport was substantially inhibited by the analog alpha-methyl aminoisobutyric acid (MeAIB). Likewise, MeAIB fluxes were completely inhibited by L-alanine, indicating the presence of system A for neutral amino acid transport. System A transport activity was electrogenic and exhibited hyperbolic relationships for net MeAIB fluxes, PD, and Isc, which displayed similar apparent K(m) values. Na(+)-dependent L-alanine transport, but not MeAIB transport, was partially inhibited by L-serine and L-cysteine, indicating the participation of system ASC. This transport activity represents the major pathway for L-alanine absorption and seemed to operate in an electroneutral mode with a negligible contribution to the L-alanine-induced electrogenicity. It is concluded from the present study that the active Na(+)-dependent L-alanine transport across the isolated duodenal mucosa of Gallotia galloti results from the independent activity of systems A and ASC for neutral amino acid transport.

Alanine↗
PubMed1995

Medicare program; update of ambulatory surgical center (ASC) payment rates effective for services on or after October 1, 1995-HCFA. Notice.

This notice implements section 1833(i)(2)(C) of the Social Security Act, which mandates an automatic inflation adjustment to Medicare payment amounts for ambulatory surgical center (ASC) facility services during the years when the payment amounts are not updated based on a survey of the actual audited costs incurred by ASCs.

Ambulatory Surgical Procedures↗
PubMed2000

The new ASC safe harbor: the triumph of public policy over legal consistency.

This Article traces the history of the OIG's use of safe harbors to allow certain activities that arguably violate a literal reading of the Anti-Kickback Statute. The author focuses particular attention upon the development of the ASC Safe Harbor from its genesis through its final issuance on November 19, 1999. The analysis finds legal inconsistency in the regulation in this area, which it attributes to the development of the ASC Safe Harbor being driven primarily by nonlegal, public policy concerns. The Article concludes by suggesting that OIG publicly embrace those policy goals and adopt broader investment-related safe harbors that focus on limiting returns to those attainable in a "fair market."

Fraud↗
PubMed2005

A poxvirus-encoded pyrin domain protein interacts with ASC-1 to inhibit host inflammatory and apoptotic responses to infection.

Proinflammatory caspases play an essential role in innate immune responses to infection by regulating the cleavage and activation of proinflammatory cytokines. Activation of these enzymes requires the assembly of an intracellular molecular platform, termed the inflammasome, which is comprised of members of the pyrin domain (PYD)-containing superfamily of apoptosis and inflammation-regulatory proteins. We report here the identification and characterization of a poxvirus-encoded PYD-containing protein that interacts with the ASC-1 component of the inflammasome and inhibits caspase-1 activation and the processing of IL-1beta and IL-18 induced by diverse stimuli. Knockout viruses that do not express this protein are unable to productively infect monocytes and lymphocytes due to an abortive phenotype and are markedly attenuated in susceptible hosts due to decreased virus dissemination and enhanced inflammatory responses at sites of infection. Thus, modulation of inflammasome function constitutes an important immunomodulatory strategy employed by poxviruses to circumvent host antiviral responses.

Amino Acid Sequence↗
PubMed1983

Red-cell amino acid transport. Evidence for the presence of system ASC in mature human red blood cells.

The properties of Na+-dependent L-alanine transport in human erythrocytes were investigated using K+ as the Na+ substitute. Initial rates of Na+-dependent L-alanine uptake (0.2 mM extracellular amino acid) for erythrocytes from 22 donors ranged from 40 to 180 mumol/litre of cells per h at 37 degrees C. Amino acid uptake over the concentration range 0.1-8 mM was consistent with a single saturable component of Na+-dependent L-alanine transport. Apparent Km and Vmax. values at 37 and 5 degrees C measured in erythrocytes from the same donor were 0.27 and 0.085 mM respectively, and 270 and 8.5 mumol/litre of cells per h respectively. The transporter responsible for this uptake was identified as system ASC on the basis of cross-inhibition studies with a series of 42 amino acids and amino acid analogues. Apparent Ki values for glycine, L-alpha-amino-n-butyrate, L-serine and L-leucine as inhibitors of Na+-dependent L-alanine uptake at 37 degrees C were 4.2, 0.12, 0.16 and 0.70 mM respectively. Reticulocytes from a patient with inherited pyruvate kinase deficiency were found to have a 10-fold elevated activity of Na+-dependent L-alanine uptake compared with erythrocytes from normal donors. Separation of erythrocytes according to cell density (cell age) established that even the oldest mature erythrocytes retained significant Na+-dependent L-alanine transport activity. Amino acid transport was, however, a more sensitive indicator of cell age than acetylcholinesterase activity. Erythrocytes were found to accumulate L-alanine against its concentration gradient (distribution ratio approx. 1.5 after 4 h incubation), an effect that was abolished in Na+-free media. Na+-dependent L-alanine uptake was shown to be associated with L-alanine-dependent Na+ influx, the measured coupling ratio being 1:1.

Alanine↗
PubMed2002

Caspy, a zebrafish caspase, activated by ASC oligomerization is required for pharyngeal arch development.

The pyrin domain was identified recently in multiple proteins that are associated with apoptosis and/or inflammation, but the physiological and molecular function of these proteins remain poorly understood. We have identified Caspy and Caspy2, two zebrafish caspases containing N-terminal pyrin domains. Expression of Caspy and Caspy2 induced apoptosis in mammalian cells that were inhibited by general caspase inhibitors. Biochemical analysis revealed that both Caspy and Caspy2 are active caspases, but they exhibit different substrate specificity. Caspy, but not Caspy2, interacted with the zebrafish orthologue of ASC (zAsc), a pyrin- and caspase recruitment domain-containing protein identified previously in mammals. The pyrin domains of both Caspy and zAsc were required for their interaction. Furthermore, zAsc and Caspy co-localized to the "speck" when co-transfected into mammalian cells. Enforced oligomerization of zAsc, but not simple interaction with zAsc, induced specific proteolytic activation of Caspy and enhanced Caspy-dependent apoptosis. Injection of zebrafish embryos with a morpholino antisense oligonucleotide corresponding to caspy resulted in an "open mouth" phenotype associated with defective formation of the cartilaginous pharyngeal skeleton. These studies suggest that zAsc mediates the activation of Caspy, a caspase that plays an important role in the morphogenesis of the jaw and gill-bearing arches.

Amino Acid Sequence↗
PubMed1992

Nucleotide sequence, function, activation, and evolution of the cryptic asc operon of Escherichia coli K12.

The cryptic asc (previous called "SAC") operon of Escherichia coli K12 has been completely sequenced. It encodes a repressor (ascG); a PTS enzyme IIasc for the transport of arbutin, salicin, and cellobiose (ascF); and a phospho-beta-glucosidase that hydrolyzes the sugars which are phosphorylated during transport (ascB). ascG and ascFB are transcribed from divergent promoters. The cryptic operon is activated by the insertion of IS186 into the ascG (repressor) gene. The ascFB genes are paralogous to the cryptic bglFB genes, and ascG is paralogous to galR. The duplications that gave rise to these paralogous genes are estimated to have occurred approximately 320 Mya, a time that predates the divergence of E. coli and Salmonella typhimurium.

Amino Acid Sequence↗
PubMed1999

Effects of pH changes on systems ASC and B in rabbit ileum.

Influx of D-aspartate (D-Asp), L-glutamate (L-Glu), and serine (Ser) across the brush-border membrane of the intact mucosa from rabbit ileum has been examined. L-Glu influx is chloride independent and completely sodium dependent. D-Asp and L-Glu share a transport system with a maximum transport rate of 1 micromol. cm-2. h-1 and an apparent affinity constant (K1/2) of approximately 0.3 mM. The function of this transport system is pH insensitive between pH 5.65 and 8.2, and bipolar amino acids do not affect the way in which the transport system handles D-Asp and L-Glu. The characteristics of this transport system match those of system X-AG. L-Glu and Ser share a transporter for which the inhibitor constant (Ki) of L-Glu against Ser decreases from 54 to 10 mM when pH is reduced from 7.2 to 5.65, while the maximum rate of transport remains unaffected at approximately 10 micromol. cm-2. h-1. The Ki values (5 mM) of Ser against L-Glu influx and the L-Glu-sensitive contribution to Ser influx (0.8 micromol. cm-2. h-1 at 1 mM Ser) are the same at both pH values. The L-Glu-sensitive transport of Ser together with the contribution of system bo,+ account for approximately 50% of Ser influx at pH 7.2. The remaining 50% can be ascribed to system B. Transport of Ser by system B is reduced by >95% at pH 5.65. At pH 7. 2 Ki of Ser against transport of leucine (Leu) by system B is 18 mM and Ki of Leu against transport of Ser is 1.7 mM. The low-affinity transport of L-Glu and the L-Glu-sensitive transport of Ser are performed by an equivalent of system ASC. Supplementary experiments using the jejunum confirm the validity of these results for a major portion of the rabbit small intestine.

Amino Acid Transport Systems↗
PubMed1996

Regulation of the ASC system and Na+/K + pump activities in brown trout (Salmo trutta) hepatocytes

The present study investigates the regulation of Na+/K+ pump activity and alanine uptake in trout hepatocytes. Pump activity increased when cells were incubated in an amino-acid-free medium, while it was reduced in cells from fasted animals. Short-term exposure (3 h) to glucagon modified the activity of the pump in a complex seasonally dependent pattern: in experiments carried out in autumn and winter there was some inhibition, while in spring the pump was activated by this hormone. Pharmacological modification of levels of two intracellular signal transducers, namely cyclic AMP and Ca2+, always led to a reduction in pump activity. These experiments were conducted in May, when activation of the pump by glucagon exposure occurred. There is no apparent explanation for the mechanism by which this hormone modifies the activity of the pump. Glucagon also regulates the activity of system ASC (a Na+-dependent amino acid carrier with short-chain neutral amino acids as preferred substrates). This regulation also showed a seasonally dependent pattern, although the pattern was opposite to that found for the regulation of Na+/K+ pump activity.

Journal Article↗
PubMed2001

ASCs spin medical event reporting into QI gold.

As reported in the September 2001 Data Strategies & Benchmarks, ambulatory surgery centers have taken the leap into medical events reporting. Learn the specifics of how some ASCs have turned what they found into concrete operational and clinical gains.

Disclosure↗
PubMed2004

Management of patients with two consecutive ASC-US smears.

INTRODUCTION: To determine whether aggressive or expectative management of patients after two consecutive smears with atypical squamous cells of undetermined significance is preferable. To determine whether triage with high-risk human papillomavirus will identify all patients with cervical intraepithelial neoplasia grade 2 and 3. METHODS: 140 of 282 patients referred for colposcopy with two consecutive smears with atypical squamous cells of undetermined significance were only treated when abnormalities suggestive of high-grade cervical intraepithelial neoplasia were present at colposcopy. The other 142 patients underwent excision of all detected colposcopic abnormalities. Both groups were compared regarding the final cytological follow-up, the number of diathermy loop excisions, and the detection of cervical intraepithelial neoplasia. Retrospectively, the outcome of triage with high-risk human papillomavirus in the first group was investigated. RESULTS: There was no significant difference in final cytological follow-up between patients managed by expectative or by aggressive colposcopic management. Significantly less diathermy loop excisions (p < 0.001) are performed in case of expectative management. The sensitivity, specificity, negative- and positive predictive values of triage with high-risk human papillomavirus detection were comparable with those of colposcopy alone. CONCLUSIONS: Patients referred with two consecutive ASC-US smears may be followed with an expectative colposcopic management and cytological follow-up. Triage with high-risk human papillomavirus will reduce the number of referrals and colposcopies, but (cytological) follow-up remains necessary in all high-risk human papillomavirus negative patients as well.

Colposcopy↗
PubMed1991

ORN role in an office based ASC.

This article outlines the role of an ophthalmic registered nurse (ORN) in an office based ambulatory surgical center (ASC). Advantages of this concept to both nurse and client will be outlined. In an office based ambulatory surgery center the opportunity exists for the professional nurse to follow the client throughout the preoperative, intraoperative and postoperative phases of the surgical procedure.

Ambulatory Surgical Procedures↗
PubMed2004

Microglandular hyperplasia has a cytomorphological spectrum overlapping with atypical squamous cells-cannot exclude high-grade squamous intraepithelial lesion (ASC-H).

We retrospectively evaluated cytological interpretations of conventional cervical smears in 48 cases of biopsy-proven microglandular hyperplasia and compared them with 15 negative controls. A glandular pattern was noted in all 48 cases but was predominant in 73% (35/48) of the cases. Immature metaplastic pattern was present in 71% (34/48) and was predominant in 27% (13/48). These features were not observed in negative control smears. 85% of cases (41/48) were interpreted as negative for epithelial cell abnormality. Two cases with predominantly glandular pattern (6%, 2/35) were interpreted as atypical glandular cells. Five cases with predominantly immature metaplastic pattern (38%, 5/13) showed checkerboard arrangement or rows of single cells with slightly larger atypical nuclei leading to interpretation as "atypical squamous cells-cannot exclude high-grade squamous intraepithelial lesion." The metaplastic cells with normoblast-like karyorrhectic apoptotic debris confined to the nuclear area were scattered among these cells in microglandular hyperplasia with metaplastic pattern.

Adult↗