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M Cheung

Publications and source records attributed to M Cheung.

At least 37 records · Page 2Linked to original sources
PubMed1997

Probing the structural and functional domains of the CFTR chloride channel.

The cystic fibrosis transmembrane conductance regulator (CFTR) forms an anion-selective channel involved in epithelial chloride transport. Recent studies have provided new insights into the structural determinants of the channel's functional properties, such as anion selectivity, single-channel conductance, and gating. Using the scanning-cysteine-accessibility method we identified 7 residues in the M1 membrane-spanning segment and 11 residues in and flanking the M6 segment that are exposed on the water-accessible surface of the protein; many of these residues may line the ion-conducting pathway. The pattern of the accessible residues suggests that these segments have a largely alpha-helical secondary structure with one face exposed in the channel lumen. Our results suggest that the residues at the cytoplasmic end of the M6 segment loop back into the channel, narrowing the lumen, and thereby forming both the major resistance to ion movement and the charge-selectivity filter.

ATP-Binding Cassette Transporters↗
PubMed1997

Locating the anion-selectivity filter of the cystic fibrosis transmembrane conductance regulator (CFTR) chloride channel.

The cystic fibrosis transmembrane conductance regulator forms an anion-selective channel; the site and mechanism of charge selectivity is unknown. We previously reported that cysteines substituted, one at a time, for Ile331, Leu333, Arg334, Lys335, Phe337, Ser341, Ile344, Arg347, Thr351, Arg352, and Gln353, in and flanking the sixth membrane-spanning segment (M6), reacted with charged, sulfhydryl-specific, methanethiosulfonate (MTS) reagents. We inferred that these residues are on the water-accessible surface of the protein and may line the ion channel. We have now measured the voltage-dependence of the reaction rates of the MTS reagents with the accessible, engineering cysteines. By comparing the reaction rates of negatively and positively charged MTS reagents with these cysteines, we measured the extent of anion selectivity from the extracellular end of the channel to eight of the accessible residues. We show that the major site determining anion vs. cation selectivity is near the cytoplasmic end of the channel; it favors anions by approximately 25-fold and may involve the residues Arg347 and Arg 352. From the voltage dependence of the reaction rates, we calculated the electrical distance to the accessible residues. For the residues from Leu333 to Ser341 the electrical distance is not significantly different than zero; it is significantly different than zero for the residues Thr351 to Gln353. The maximum electrical distance measured was 0.6 suggesting that the channel extends more cytoplasmically and may include residues flanking the cytoplasmic end of the M6 segment. Furthermore, the electrical distance calculations indicate that R352C is closer to the extracellular end of the channel than either of the adjacent residues. We speculate that the cytoplasmic end of the M6 segment may loop back into the channel narrowing the lumen and thereby forming both the major resistance to current flow and the anion-selectivity filter.

Anions↗
PubMed1997

Inability of physicians to use metered-dose inhalers.

The effectiveness of antiasthmatic therapy with metered-dose inhalers (MDIs) is usually limited by defective inhalational technique. This study surveyed the MDI inhalation performance and knowledge of 100 physicians whose performance (demarcated into six stages) and knowledge scores correlated (r = 0.33, p < 0.001). Inhalation performance scores for shaking the canister, full expiration prior to using the MDI, breath holding thereafter, and positioning of the mouth-piece correlated with their corresponding knowledge scores (p < or = 0.02). Training of physicians entailing actual instruction and supervision of inhalational maneuvers may enhance the ability of doctors to use MDIs and facilitate better patient supervision.

Administration, Inhalation↗
PubMed1996

Identification of cystic fibrosis transmembrane conductance regulator channel-lining residues in and flanking the M6 membrane-spanning segment.

The cystic fibrosis transmembrane conductance regulator (CFTR) forms a chloride channel that is regulated by phosphorylation and ATP binding. Work by others suggested that some residues in the sixth transmembrane segment (M6) might be exposed in the channel and play a role in ion conduction and selectivity. To identify the residues in M6 that are exposed in the channel and the secondary structure of M6, we used the substituted cysteine accessibility method. We mutated to cysteine, one at a time, 24 consecutive residues in and flanking the M6 segment and expressed these mutants in Xenopus oocytes. We determined the accessibility of the engineered cysteines to charged, lipophobic, sulfhydryl-specific methanethiosulfonate (MTS) reagents applied extracellularly. The cysteines substituted for Ile331, Leu333, Arg334, Lys335, Phe337, Ser341, Ile344, Arg347, Thr351, Arg352, and Gln353 reacted with the MTS reagents, and we infer that they are exposed on the water-accessible surface of the protein. From the pattern of the exposed residues we infer that the secondary structure of the M6 segment includes both alpha-helical and extended regions. The diameter of the channel from the extracellular end to the level of Gln353 must be at least 6 A to allow the MTS reagents to reach these residues.

ATP-Binding Cassette Transporters↗
PubMed1995

Predictive evidence for a porin-type beta-barrel fold in CHIP28 and other members of the MIP family. A restricted-pore model common to water channels and facilitators.

Water channels are the subject of much current attention, as they may be central for cell functions in a host of tissues. We have analyzed the possible field of facilitators and water channels of the MIP family based on structural predictions, on findings about the topology of CHIP28, and on the biophysical characteristics of water channels. We developed predictions for the following proteins: MIP26, NOD26, GLP, BIB, gamma-TIP, FA-CHIP, CHIP28k, WCH-CD1, and CHIP28. We utilized Kyte Doolittle hydrophobicity, Eisenberg's amphiphilicity, Chou-Fasman-Prevelige propensities, and our own Union algorithm. We found that hydrophobic amphiphilic segments likely to be transmembrane were consistently shorter than required for alpha-helical segments, but of the correct length for beta-strands. Turn propensity was high at frequent intervals, consistent with transmembrane beta-strands. We propose that these proteins fold as porin-like 16-stranded antiparallel beta-barrels. In water channels, from the size of molecules excluded, an extramembrane loop(s) would enter the pore and restrict it to a bottleneck with a width 4 A < or = w < or = 5 A. A similar but more mobile loop(s) would act as gate and binding site for the facilitators of the MIP family.

Amino Acid Sequence↗
PubMed1994

Are most transporters and channels beta barrels?

Given the sequence of transporters or channels of unknown secondary structure, it is usual to predict their putative transmembrane regions as alpha-helical. However, recent evidence for a facilitative glucose transporter (GLUT1) appears inconsistent with such predictions, which has led us to propose an alternative folding model for GLUTs based on the 16-stranded antiparallel beta-barrel of porins. Here we apply the same predictive algorithms we used for GLUTs to several other membrane proteins. For some of them, a high-resolution structure has been derived (beta-barrels: Rhodobacter capsulatus and Escherichia coli porins; multihelical: colicin A, bacteriorhodopsin, and reaction center L chain); we use them to test the prediction procedures. The other proteins we analyze (GLUT1, CHIP28, acetylcholine receptor alpha subunit, lac permease, Na(+)-glucose cotransporter, shaker K+ channel, sarcoplasmic reticulum Ca(2+)-ATPase) are representative of classes of similar membrane proteins. As with GLUTs, we find that the predicted transmembrane segments of these proteins are consistently shorter than expected for transmembrane spanning alpha-helices, but are of the correct length and number for the proteins to fold instead as porin-like beta-barrels.

Algorithms↗
PubMed1993

Evidence that facilitative glucose transporters may fold as beta-barrels.

A widely accepted model for the structure of the facilitative glucose transporters (GLUTs) predicts that they form 12 transmembrane alpha-helices and that the highly conserved sequence Ile-386-Ala-405 in GLUT1 is intracellular. We raised a polyclonal antibody against a synthetic peptide encompassing this conserved sequence and found that antibody treatment increased 2-deoxy-D-glucose (DOG) uptake in Xe-nopus oocytes expressing GLUT1, GLUT2, or GLUT4 only when applied to the extracellular side. This effect was dose dependent and was specifically blocked by competition with the peptide Ile-386-Ala-405; it was due to a decrease in the Km for the transport of DOG. To ascertain GLUT orientation, we raised anti-peptide antibodies against the last 21 and 25 C-terminal amino acids of GLUT1 and GLUT4, respectively, which were previously shown to be intracellular. These antibodies increased DOG uptake when injected into oocytes expressing GLUT1 and GLUT4, but not when added extracellularly. Prompted by the noted discrepancy, we found sequence similarity between GLUTs and porins, two of which are known from crystallography to form 16-stranded transmembrane antiparallel beta-barrels. Analysis of the hydrophobicity, amphiphilicity, and turn propensity of GLUT1 leads us to propose that GLUTs fold as porin-like transmembrane beta-barrels. This model is consistent with the results of the present antibody studies and also with previously published experimental evidence inconsistent with the 12-helix model.

Amino Acid Sequence↗
PubMed1993

Poor outcome of on-site CPR in a multi-level geriatric facility: three and a half years experience at the Baycrest Centre for Geriatric Care.

Our objective was to determine the outcome of onsite CPR in elderly patients receiving institutional care who had access to 24-hour on-site full ACLS capability. We used a retrospective chart and CPR protocol review of all patients who received CPR during the study period; these were patients or residents in a multi-level geriatric long-term care facility that provides various degrees of facility-based and ambulatory care. Immediate, short-term, and long-term survivorship were determined and correlated where possible with the category of patient seen (short-term care versus long-term care) and the relationship between the witnessing of the arrest and survival. Of the 41 patients who underwent CPR, there were only four survivors of 60 days or greater. Of these, three subjects were short-term, not long-term care patients of the Centre; they returned to their previous level of function. One fully dependent bed-ridden patient returned to that level of care for a survival period of 100 days. Of the four long-term survivors, three of the arrests were witnessed, and one was indeterminate. No unwitnessed arrests resulted in long-term survival. Our experience suggests that CPR in the elderly long-term care patient is unlikely to be successful even when it is available on-site. Unwitnessed arrests in this population were universally fatal. This information has helped Baycrest Centre for Geriatric Care develop protocols, guidelines, and policies for CPR that are suitable for our population.

Aged↗
PubMed1990

Isolation and immunohistochemical characterization of a pancreatic carcinoma-associated monoclonal antibody.

The aim of this study was to define a cellular antigen associated with human pancreatic ductal carcinoma, and to study its distribution in a large panel of malignant, benign, and normal tissues. For this purpose, monoclonal antibodies were generated against a postmicrosomal fraction of fresh human pancreatic cancer. One such antibody, LD-B1, reacted strongly with 95% of cases of primary and metastatic pancreatic ductal carcinomas. It also immunostained gallbladder carcinomas and cholangiocarcinomas. By contrast, it exhibited focal or weak reactivity to 10% of other types of common malignant tumors. On normal pancreas, staining was observed in ductal and centriacinar cells, but not in acinar or endocrine cells. In chronic pancreatitis, ductal staining intensity increased proportionally with the degree of cellular atypia. The antigen was also detected in gallbladder epithelium, bile ducts, ductal epithelium of sweat glands and salivary glands, and focally in a few other normal nonpancreatic tissues. These results suggest that LD-B1 MoAb can be used in immunohistochemical studies as a marker of pancreatic adenocarcinoma.

Adenocarcinoma↗
PubMed1990

The significance of skull fracture in acute traumatic intracranial hematomas in adolescents: a prospective study.

A prospective study was conducted to validate the retrospective finding that adolescents (11 to 15 years old) with skull fractures were prone to develop acute traumatic intracranial hematoma (ICH). Over a 4-year period, 1178 consecutive adolescents attended the emergency room directly, of whom 760 were discharged well and 418 were admitted. All underwent skull x-ray studies. Immediate computerized tomography (CT) scans were performed in patients with Glasgow Coma Scale (GCS) scores of less than 15, in those with radiological and/or clinical evidence of skull fracture, and whenever clinically indicated. Of the 418 admitted patients, only 26 had skull fractures; 13 of these developed ICH. Four patients without skull fracture developed diffuse brain swelling. The remaining 401 patients were discharged after observation periods of up to 48 hours. Of the 13 patients with ICH, 10 had admission GCS scores of 15; however, four deteriorated rapidly and required urgent operation, and four remained stable but were operated on due to their large ICH. Two required conservative treatment only and both made good recovery. Three patients were in coma (GCS score less than or equal to 8) on admission. One patient had an epidural hematoma and made good recovery after surgery. Two developed delayed ICH after operations for associated systemic injuries despite initial CT showing diffuse brain swelling only, and both died despite evacuation of the ICH. Multivariate analysis showed that skull fracture was the only independent significant risk factor in predicting ICH in adolescents (sensitivity of 100% and specificity of 97%). A routine skull x-ray study is therefore mandatory in all head-injured adolescents and, if a skull fracture is detected, immediate CT may be performed for early detection of ICH.

Acute Disease↗
PubMed1989

Isoflurane or halothane for cesarean section: comparative maternal and neonatal effects.

The maternal and neonatal effects of isoflurane and halothane combined with 50% N2O - 50% O2 were compared in 60 healthy parturients undergoing primary or repeat cesarean section. All patients had rapid sequence induction of anesthesia with sodium thiamylal 4 mg/kg followed by succinylcholine for tracheal intubation. Patients were randomly assigned to one of three groups of 20 each (inspired 0.5% isoflurane, 1% isoflurane or 0.5% halothane), combined with 50% N2O and O2. After delivery, 67% N2O in O2 was used, supplemented by butorphanol. Maternal blood loss did not differ significantly among the three groups and none of the patients developed intraoperative awareness. At the time of delivery, maternal plasma epinephrine levels were significantly above preinduction levels in the 0.5% isoflurane group but unchanged in the other two groups. Neonatal status as ascertained by Apgar scores, cord acid base status and the Neurologic and Adaptive Capacity Scores (NACS) was equally good in the three groups of patients. Serum inorganic fluoride concentrations in the mother after anesthesia were not significantly above preanesthetic levels in any of the groups and there was no biochemical evidence of renal toxicity. In all neonates fluoride ion concentrations in the first voided urine sample were less than 7 mumol/l, a value well below that associated with nephrotoxicity. It is concluded that isoflurane is a safe supplement to N2O - O2 mixture for cesarean section and is a safer alternative to halothane in situations when patients receiving beta-adrenergic therapy require cesarean section since halothane might potentiate arrhythmias caused by beta adrenergic agonists.

Adult↗
PubMed1989

A randomized study of high-dose versus low-dose cis-platinum combined with cyclophosphamide in the treatment of advanced ovarian cancer. Hong Kong Ovarian Carcinoma Study Group.

A randomized clinical study of patients with advanced epithelial ovarian cancer after debulking surgery showed that high-dose (120 mg/m2) cis-platinum (DDP) in combination with cyclophosphamide (600 mg/m2) had a significantly higher response and survival rate than the low-dose DDP (60 mg/m2) and cyclophosphamide combination. The 3-year actuarial survival rate of the high-dose group was 60% and that of the low-dose group was 30%. Though moderate to severe marrow toxicity was evident in 80% of the patients in the high-dose group and 40% of the low-dose group, no serious sepsis or death developed as a result of the marrow depression. Mild neurotoxicity was observed in 55% of the patients in the high-dose group and only 20% in the low-dose group. Mild nephrotoxicity was seen in 25 and 17% of patients in the high- and low-dose groups, respectively. It was concluded that the 120 mg/m2 dose DDP and cyclophosphamide combination should be used in the treatment of carcinoma of the ovary in spite of its toxicities. However, it should only be used in institutions with supportive facilities in the management of patients with severe marrow depression.

Adolescent↗
PubMed1989

Lipoprotein, apolipoprotein, and lipolytic enzyme changes following estrogen administration in postmenopausal women.

To test whether estrogen can modulate the cholesterolemic response to an Occidental diet, six healthy postmenopausal women were studied for 84 days while ingesting a solid food diet of constant composition high in cholesterol content (995 mg/d). In the middle of the study, estrogen (17 alpha-ethinyl estradiol, 1 microgram/kg per day) was administered orally. Ingestion of the diet for the initial 28 days did not alter lipoprotein lipid or apolipoprotein (apo) levels. However, with just 4 days of estrogen use there were significant decreases in apoE (-36%), low density lipoprotein cholesterol (-26%), and postheparin plasma hepatic triglyceride lipase activity (HTGL) (-61%), and an increase in high density lipoprotein (HDL) triglyceride (72%). These changes persisted throughout the estrogen use. The percent change in HTGL with 4 days of estrogen correlated inversely with the percent change in HDL triglyceride (rs = -0.94). After 28 days of estrogen there were also significant increases in HDL cholesterol (21%), HDL2 cholesterol (42%), apoA-I (37%), and apoA-II (9%), and a decrease in apoB (-11%). The level of apoE at this juncture correlated inversely with the level of HDL cholesterol (rs = -0.90), and the levels of HTGL and apoA-I correlated with HDL2 cholesterol (rs = -0.89 and rs = 0.89, respectively). Thus, HTGL may play a role in both the early estrogen-related changes in HDL triglyceride and apoE and the late estrogen-related changes in HDL cholesterol, apoA-I, and apoA-II.

Aged↗
PubMed1988

Enzymatic properties of the 15-lipoxygenase of human cultured keratinocytes.

The arachidonic acid 15-lipoxygenase or linoleic acid omega-6 lipoxygenase of human neonatal foreskin cultured keratinocytes converts arachidonic acid to 15-hydroxy-eicosatetraenoic acid and linoleic acid to 13-hydroxy-linoleic acid. A mean of 93% of the 15-lipoxygenase activity in sonicates of cultured keratinocytes was recovered in the 400,000 X g supernatant, attesting to the cytosolic localization of this enzyme. Optimal 15-lipoxygenase activity in the 400,000 X g supernatant was expressed at pH 6.7-7.3 and in the presence of calcium at a concentration of 2 mM or higher. Keratinocyte 15-lipoxygenase metabolized arachidonic acid (Km = 10.6 microM) and linoleic acid (Km = 9.5 microM) with similar efficiency. Nordihydroguaiaretic acid and 5,8,11,14-eicosatetraynoic acid both inhibited the conversion of arachidonic acid to 15-HETE with respective 50% inhibitory concentrations of 2.0 microM and 0.9 microM, while ATP, GTP, and cyclic AMP had no effect on activity at pH 6.8-7.2. The enzymatic properties of human keratinocyte 15-lipoxygenase thus resemble those of PMN leukocyte 15-lipoxygenase and the mediators generated may contribute to the regulation of cutaneous sensation and inflammation.

Arachidonate 15-Lipoxygenase↗