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Biomedical subjects

F H Tsao

Publications and source records attributed to F H Tsao.

At least 19 recordsLinked to original sources
PubMed1995

Annexin I in female rabbit reproductive organs: varying levels in relation to maturity and pregnancy.

The level of annexin I, a 36 kDa calcium-dependent phospholipid-binding protein (36 kDa PLBP) in the reproductive organs of young, mature, and pregnant rabbits was determined immunologically with antibodies raised against purified rabbit lung annexin I. In the cytosolic fractions of the ovary, fallopian tube, uterus, and placenta, annexin I was the only major immunoreactive protein. The reproductive organs appeared to have higher annexin I levels than most nonreproductive organ tissues, except the lung and the spleen which were also rich in annexin I. A small amount of annexin I and a nearly equal amount of its hydrolytic product, a 33 kDa polypeptide, were detected in the amniotic fluid between 21 and 27 days gestation. Structural similarity of annexin I in the reproductive organs and in the lung was suggested by their identical isoelectric point values. Annexin I in the ovary of adult rabbits was 70% higher than that in the respective organ of immature rabbits. The uterus of pregnant rabbits had about 84% higher annexin I contents than that of the nonpregnant rabbits. The placenta had more annexin I per mg cytosolic protein than either the ovary or the uterus during pregnancy. The high concentration of annexin I in the reproductive organs may reflect specific functions of these organs in the reproductive years and during the reproductive cycle.

Amniotic Fluid↗
PubMed1992

Identification of calcium-dependent phospholipid-binding proteins (annexins) from guinea pig alveolar type II cells.

A new group of calcium-regulating proteins, called annexins or Ca(++)-dependent phospholipid-binding proteins (PLBP), have been detected in different species, organs and cell types. In the present study, we have identified and quantitated PLBP from guinea pig lung, lavage fluid and alveolar type II cells to elucidate the possible role of PLBP in lung surfactant biogenesis and secretion. Lungs were lavaged and type II cells from lavaged lung were isolated by elastase digestion and purified by centrifugal elutriation. For the quantitative identification of PLBP, we performed ELISA assays and Western blot analysis by using an antiserum raised in guinea pigs against a pure rabbit lung 36 kDa PLBP. The lavage fluid, cytosol from lung and type II cells contained 784, 167 and 435 ng per mg protein, respectively, of PLBP. The SDS-PAGE electrophoretic pattern and Western blot confirmed that all lung samples have band corresponding to a 36 kDa protein. This indicates that both alveolar type II cells and lavage fluid have higher levels of PLBP than whole lung cytosol.

Animals↗
PubMed1992

Purification, characterization and substrate specificity of rabbit lung phospholipid transfer proteins.

Three phospholipid transfer proteins, namely proteins I, II and III, were purified from the rabbit lung cytosolic fraction. The molecular masses of phospholipid transfer proteins I, II and III are 32 kilodaltons (kDa), 22 kDa and 32 kDa, respectively; their isoelectric point values are 6.5, 7.0 and 6.8, respectively. Phospholipid transfer proteins I and III transferred phosphatidylcholine (PC) and phosphatidylinositol (PI) from donor unilamellar liposomes to acceptor multilamellar liposomes; protein II transferred PC but not PI. All the three phospholipid transfer proteins transferred phosphatidylethanolamine poorly and showed no tendency to transfer triolein. The transfer of [14C]PC from unilamellar liposomes to multilamellar liposomes facilitated by each protein was affected differently by the presence of acidic phospholipids in the PC unilamellar liposomes. In an equal molar ratio of acidic phospholipid and PC, phosphatidylglycerol (PG) reduced the activities of proteins I and III by 70% (P = 0.0004 and 0.0032, respectively) whereas PI and phosphatidylserine (PS) had an insignificant effect. In contrast, the protein II activity was stimulated 2-3-times more by either PG (P = 0.0024), PI (P = 0.0006) or PS (P = 0.0038). In addition, protein II transferred dioleoylPC (DOPC) about 2-times more effectively than dipalmitoylPC (DPPC) (P = 0.0002), whereas proteins I and III transferred DPPC 20-40% more effectively than DOPC but this was statistically insignificant. The markedly different substrate specificities of the three lung phospholipid transfer proteins suggest that these proteins may play an important role in sorting intracellular membrane phospholipids, possibly including lung surfactant phospholipids.

1,2-Dipalmitoylphosphatidylcholine↗
PubMed1992

Use of quantitative amniotic fluid phosphatidylglycerol as a criterion for fetal lung maturation.

Phosphatidylglycerol (PG) in amniotic fluid was quantitatively measured by thin-layer chromatography (TLC) in 941 amniotic fluid samples and related to the presence or absence of respiratory distress syndrome (RDS) in neonates born within 7 days of the amniocentesis. In 639 case tests with PG 1 nmol/ml or more of amniotic fluid, there was no RDS. However, there were six cases of RDS associated with 109 tests of PG from 0 to less than 1 nmol/ml. These 109 tests would all have been reported as PG positive if PG were only qualitatively assessed on the TLC plate by the naked eye. The highest incidence of RDS occurred when PG was absent (23 RDS of 193 patients). Hence, this study suggests that quantitative analysis of PG determines a cutoff point of PG that eliminates false-positive PG assessments (1 nmol/ml in our laboratory). There was no difference in the levels of PG between males and females at equal gestational ages, but the incidence of RDS among male neonates was 2.4 times higher (p less than 0.05) than female neonates in the group with an immature amniotic fluid lecithin to sphingomyelin ratio and PG less than 1.0 nmol/ml.

Amniotic Fluid↗
PubMed1991

Lung calcium-dependent phospholipid-binding proteins: structure and function.

Distinct peptide maps of two rabbit lung Ca2(+)-dependent phospholipid-binding proteins (PLBPs), 36,000 and 33,000, were generated by cyanogen bromide (CNBr) cleavage, trypsin or Staphylococcus aureus V8 proteinase digestion. The amino acid sequence of a CNBr-cleaved peptide of the 36,000 PLBP was aligned to the amino terminus of human lipocortin I with more than 77% identity, but had no identity with the known amino terminal sequence of other known annexins. Partial amino acid sequence of a 33,000 PLBP peptide demonstrated a close (56%) relationship to endonexin II, human placental anticoagulant protein, and porcine intestine protein II, but shared only 32% identity with lipocortin I, 30% with lipocortin II. Antiserum generated against purified 36,000 PLBP reacted strongly with the 33,000 PLBP, but did not react with any other rabbit lung cytosolic proteins. Both PLBPs inhibited the phospholipase A2 reaction when dioleoyl phosphatidylcholine and phosphatidylglycerol vesicles or monolayers were used as substrates. In the vesicle assay, the phospholipase A2 reaction was inhibited at lower substrate phospholipid concentrations but not at nearly saturating substrate concentrations. In the monolayer assay, the phospholipid-binding proteins did not inhibit phospholipase A2 at a low phospholipid surface concentration of 3.8.10(-3) molecules/A2, but they did at higher surface concentrations between 1.1 x 10(-2) and 3.8 x 10(-2) molecules/A2. The inhibition of phospholipase A2 by rabbit lung phospholipid-binding proteins is most likely due to the prevention of penetration by phospholipase A2 into the interface, a requirement for the enzyme to act on the substrate.

Amino Acid Sequence↗
PubMed1990

Purification and characterization of two rabbit lung Ca2(+)-dependent phospholipid-binding proteins.

Two Ca2(+)-dependent phospholipid-binding proteins (PLBPs) in rabbit lung cytosolic fraction have been purified to homogeneity. The apparent molecular weights of these two proteins are 36,000 and 33,000. Both the 36,000 and 33,000 PLBPs aggregated certain negatively charged unilamellar liposomes, but not the neutral phosphatidylcholine (PC) liposomes, in the presence of Ca2+. However, both PLBPs fused PC unilamellar liposomes to membrane acceptors. The 36,000 and 33,000 PLBPs had different specificities for phospholipid head groups, effects of Ca2+ and membrane charges and amino acid compositions. Both the PLBPs aggregated the surfactant membranes (lamellar bodies or from lung lavage) and nonsurfactant membranes (microsomes or mitochondria) to a level similar to that of the synthetic acidic phospholipid vesicles, but the proteins fused [14C]PC liposomes to the surfactant membranes 13- to 16-times more than to the synthetic phospholipid vesicles. The 36,000 PLBP fused [14C]PC liposomes to microsomes or mitochondria only half that of the fusion to the surfactant; the 33,000 PLBP fused [14C]PC liposomes to the surfactant and nonsurfactant alike. The PLBP's aggregate activity was not affected by the depletion of biological membrane proteins and the disruption of the native membrane integrity, but its fusion activity was greatly reduced. These results suggest that: (1) the 36,000 and 33,000 PLBPs are two different proteins; (2) the PLBPs may possess two catalytic reactions, one for the aggregation of small vesicles due to the binding of the protein to phospholipid vesicles and one for the fusion of small vesicles to acceptors; (3) the fusion activity was probably regulated by biological membrane proteins or structures; and (4) lung PLBP(s) might play a role in lung surfactant biogenesis.

Amino Acids↗
PubMed1990

The use of a multichannel scaling method to detect radioactive species in spread monolayers.

This study describes a simple and inexpensive method for monitoring radioactive species spread as monolayers at the air/water interface. The combination of a discriminator and multichannel scalar counter with a personal computer functions to unify all measurements, to simplify the operational process and data acquisition, and to provide a real-time display of the data. Its use is demonstrated by following the hydrolysis of L-alpha-[1-14C]dioleoyl phosphatidylcholine by the enzyme, phospholipase A2, isolated from the porcine pancreas.

Animals↗
PubMed1989

Phosphatidylglycerol, lecithin/sphingomyelin ratio and respiratory distress syndrome in diabetic and non-diabetic pregnancies.

Samples of amniotic fluid from 514 non-diabetic and 69 diabetic patients were analyzed for phospholipid content. Results were correlated with incidence of respiratory distress syndrome (RDS) in the neonate. The incidence of RDS was 4.5% among diabetics and 5.3% among non-diabetics. In the presence of phosphatidylglycerol (PG), no infant developed RDS while in the absence of PG the incidence of RDS was 16.7% and 14.4%, respectively. In the presence of a mature lecithin/sphingomyelin (L/S) ratio the respective incidence of RDS was 1.6 and 1.8, while with an immature L/S ratio the incidence of RDS was 28.6% and 29%. The incidence of RDS after 37 weeks gestation was 0% among diabetics and 0.6% among non-diabetics. We conclude that amniotic fluid phospholipids are equally predictive of risk for RDS in diabetics as among non-diabetic patients. We suggest that in patients with accurate gestational dating, amniotic fluid analysis for phospholipids might not be necessary to establish fetal lung maturity.

Amniotic Fluid↗
PubMed1988

Retinyl ester synthesis by isolated adult rabbit lung type II cells.

Type II alveolar cells were isolated from adult rabbit lungs and then cultured on monolayers for 16 hours. These cells were then covered with buffered medium containing [3H]-retinol. After 30-120 minutes incubation, the cells were extracted with Hexane: Ethanol and the hexane extract analyzed by HPLC. A linear synthesis of [3H]-retinyl palmitate with time of incubation was demonstrated.

Animals↗
PubMed1986

Reversibility of cholinephosphotransferase in lung microsomes.

The effect of cytidine 5'-monophosphate (CMP) on the incorporation of cytidine 5'-diphosphate (CDP) [methyl-14C]choline or [1-14C]dipalmitoylglycerol into phosphatidylcholine (PC) catalyzed by rabbit lung microsomal CDPcholine:1,2-diacyl-sn-glycerol cholinephosphotransferase (EC 2.7.8.2) was studied. In the presence of 0.85 mM CMP and nonsaturating diacylglycerol concentration, the incorporation of CDP[14C]choline into PC was markedly stimulated, but the incorporation of [14C]dipalmitoylglycerol into PC was inhibited. This was due to the increase of endogenous diacylglycerol generated from microsomal PC by the cholinephosphotransferase reverse reaction. However, the newly synthesized PC was not readily hydrolyzed in the presence of CMP. The results of this study suggest that the endogenous membranous diacylglycerol is utilized more preferentially for PC synthesis than the exogenous diacylglycerol and that the newly synthesized PC could rapidly equilibrate with the endogenous membrane PC pool.

Animals↗
PubMed1986

Selective use of palmitic acid over stearic acid for synthesis of phosphatidylcholine and phosphatidylglycerol in lung.

The incorporation of [3H]palmitic acid and [14C]stearic acid into phospholipids in rabbit lung tissue was studied. Under equal molar concentrations of palmitate and stearate, palmitate was incorporated to the 1- and 2-positions of phosphatidylcholine (PC) and phosphatidylglycerol (PG) 2-3 times more than stearate. By contrast, palmitate was 30% less than stearate in phosphatidylethanolamine, phosphatidylinositol and phosphatidylserine. These results suggest that preferential utilization of palmitate over stearate, rather than substrate availability, determines the high content of palmitoyl at the 1- and 2-positions of PC and PG in lung.

Animals↗
PubMed1986

5'-Nucleotidase activity in adult and fetal rabbit lungs.

5'-Nucleotidase was assayed in the membrane fractions isolated from rabbit lung homogenate. The enzyme activity was measured from the rate of hydrolysis of [U-14C]cytidine 5'-monophosphate (CMP). The optimal pH of the rate of hydrolysis is around 8.0 and the enzyme activity is stimulated by Mg2+. The apparent Km and Vmax of the enzyme in the microsomal fraction for CMP were 3.33 mM and 1.43 mumol/min/mg protein, respectively. During lung development the enzyme activity increased moderately at late gestational ages and reached its maximum level in adults (p less than 0.001). The developing profile of 5'-nucleotidase activity is similar to the developing pattern of the CMP content in rabbit lungs.

5'-Nucleotidase↗
PubMed1983

Conversion of choline to phosphatidylcholine in the isolated-ventilated-perfused neonatal rabbit lung.

The isolated-ventilated-perfused neonatal rabbit lung model was used to study pulse dosed 14C-choline incorporation into 14C-phosphatidylcholine (PC) and 14C-disaturated phosphatidylcholine (DSPC). 14C-PC and 14C-DSPC synthesis were linearly dependent upon perfusion time to 30 min and upon the pulse dose of 14C-choline of 20-250 nmol at both 10 and 30 min of perfusion. 2-3% of the pulse-dosed choline was taken up by the lung. The water-soluble metabolites of choline found in lung after 10-30 min of perfusion were: choline, 50-60%; betaine 2.4-3.0%; phosphorylcholine, 26-41%, and CDP-choline, trace-10%. 1-day-old perfused lungs incorporated pulse-dosed 14C-choline into 14C-PC and 14C-DSPC at slightly higher rates than at 10 days old. Newborn rabbits receiving an intraperitoneal injection of 0.1 mg dexamethasone/100 g body weight 16 and 22 h prior to perfusion incorporated significantly more 14C-choline into 14C-PC and 14C-DSPC than saline-injected controls.

Animals↗
PubMed1982

Determination of phosphatidylglycerol in amniotic fluid by a simple one-dimensional thin-layer chromatography method.

Phosphatidylglycerol (PG) in amniotic fluid is the second important component of lung surfactant phospholipids and may be clinically useful in assessing fetal lung maturity in utero. Although methods for PG determination are available, there are shortcomings in clinical application. We developed an alternative reliable one-dimensional thin-layer chromatography(TLC) method for separating and quantitating PG in amniotic fluid. A mini-TLC plate (8 X 10 cm) was prepared from silica gel H containing 5% ammonium sulfate. The plate was first developed in tetrahydrofuron/dimethoxymethane/methanol/2N ammonium hydroxide (30.0:20.6:5.6:3.0 v/v) and then in chloroform/methanol (60.9, v/v) in the same direction. PG was clearly separated from other phospholipids and neutral lipids, even when large amounts of other phospholipids were present on the TLC plate. The density of the charred PG was directly proportional to the amount of PG up to 8 nmol. The content of PG in nmol in the specimen can be quantitated by comparing with a standard PG. Up to 10% of blood serum or 3% meconium showed no detectable PG, nor did these substances affect PG quantitation in amniotic fluid. This method is sensitive and accurate. It is also time-saving and economical.

Amniotic Fluid↗
PubMed1980

Specific transfer of dipalmitoyl phosphatidylcholine in rabbit lung.

Liposomes were used as a tool to probe the specific transfer of dipalmitoyl phosphatidylcholine in rabbit lung. After incubation of liposomes with lung slices, lamellar bodies took up 30-50% more radioactively labeled dipalmitoyl phosphatidylcholine from egg yolk phosphatidylcholine liposomes than the labeled dioleoyl phosphatidylcholine, whereas mitochondria and microsomes took up both species at relatively equal rates. Furthermore, when 50% egg yolk phosphatidylcholine was replaced by dipalmitoyl phosphatidylcholine, the uptake of [1-14C]dipalmitoyl phosphatidylcholine by lamellar bodies was greatly reduced due to the isotope-dilution effect of the unlabeled dipalmitoyl species. This competitive uptake was not observed in mitochondria and microsomes. Similar results were also observed when liposomes were incubated with the cell-free homogenate. On the other hand, distearoyl or dioleoyl phosphatidylcholine did not compete with the uptake of [1-14C]dipalmitoyl phosphatidylcholine. These results suggest an intracellular specific transfer of dipalmitoyl phosphatidylcholine from liposomes to lamellar bodies, presumably by a specific transfer protein. When liposomes were labeled with [1-14C]dioleoyl phosphatidylcholine, the addition of unlabeled dioleoyl phosphatidylcholine up to 50% of total phosphatidylcholine concentration did not compete with the uptake of [1-14C]dioleoyl phosphatidylcholine in all fractions including lamellar bodies. This indicates that the intracellular uptake of liposomal dioleoyl phosphatidylcholine by lamellar bodies and other subcellular organelles was probably dominated by the fusion mechanism.

Animals↗