Search PubMed⌕ Search

Biomedical subjects

F H Tsao

Publications and source records attributed to F H Tsao.

At least 19 recordsLinked to original sources
PubMed1998

The interaction of lung annexin I with phospholipid monolayers at the air/water interface.

Lung annexin I (LAI), a calcium-ion-dependent phospholipid-binding protein, has been shown earlier to cause aggregation and fusion of bilayered vesicles containing phospholipids found in lung surfactant, and to be a very likely factor in the assembly of lung surfactant into the lamellar bodies stored in the Type II cell. In this study, we have measured the accumulation of LAI into spread monolayers of some major lipid components of lung surfactant, dipalmitoyl-phosphatidylcholine (DPPC), dipalmitoyl-phosphatidylglycerol (DPPG), palmitoyl-oleyoyl-phosphatidylglycerol (POPG), and selected mixtures, as a function of calcium-ion concentration and surface concentration (degree of packing) of the phospholipid monolayer. The ability of LAI to significantly penetrate such monolayers was calcium-ion-dependent and only occurred in the presence of DPPG or POPG. The relative extent of penetration into DPPG and POPG was directly related to the available free area in the monolayer, penetration being greater with POPG. Fluorescence microscopy measurements revealed that DPPC mixed with either DPPG or POPG caused a change in surface phase behavior in a manner believed to be related to certain types of bilayer fusion. A chemical breakdown product of LAI, LAI-bp, previously found not to cause aggregation and fusion of bilayers, did not exhibit comparable monolayer penetration or surface phase separation to LAI.

Air↗
PubMed1998

Degradation of annexin I in bronchoalveolar lavage fluid from patients with cystic fibrosis.

Annexin I is a 36 kilodalton (kD) calcium-dependent phospholipid-binding protein which may have anti-inflammatory properties. Previous investigations which sampled lower respiratory tract epithelial lining fluid (ELF) via bronchoalveolar lavage (BAL) have demonstrated that annexin I can be degraded in inflammatory lung disease. We analyzed BAL fluid from patients with cystic fibrosis (CF) to determine the effects of lung inflammation on the structure and activity of annexin I. Intact annexin I was absent in 17 out of 20 BAL fluid samples from patients with CF, due largely to degradation to a 33 kD protein. The three CF BAL fluids in which annexin I was detectable had very little or no unopposed neutrophil elastase activity in contrast to the 17 in which no annexin I was detectable. Annexin I was present in all BAL fluid samples from 10 normal volunteer (NV) subjects and 12 patients with interstitial lung disease (ILD). The 33 kD annexin I breakdown product was not detectable in samples from NV, but was detectable only in ILD patients with relatively high percentages of neutrophils on BAL differential cell counts. Annexin I appeared to be cleaved by neutrophil elastase at the N-terminal portion between Val-36 and Ser-37 to yield the 33 kD protein. Cleavage of the N-terminal portion of annexin I was accompanied by a marked change in the annexin I isoelectric point (pI) value (from 6.0 to 8.5-9.0) and greatly diminished annexin I functional activity. Our findings demonstrate that annexin I degradation in epithelial lining fluid is closely related to lung inflammation.

Amino Acid Sequence↗
PubMed1997

Annexin I concentration and prostacyclin production in rat lung and alveolar macrophages following irradiation.

The purpose of this study was to gather additional evidence in irradiated rat lung on the relationship between annexin I and prostaglandin synthesis. The right hemithorax of the animal was exposed to a single dose of 0 or 30 Gy of X-rays, and the animals were killed 3 months postirradiation. Levels of annexin I and synthesis of prostacyclin (PGI2) were determined in lungs, in cell-free bronchoalveolar lavage (BAL) fluid, and in macrophages lavaged from those lungs. In addition, protein concentration, lactate dehydrogenase (LDH) activity and macrophage count in BAL fluid obtained from irradiated lung were compared with that from sham-irradiated (0 Gy) lung. Levels of annexin I, the putative inhibitor of phospholipase A2, in lung and cell-free BAL fluid were decreased in samples from irradiated animals. By contrast, the level of annexin I in macrophages lavaged from irradiated lung was higher than that in macrophages from sham-irradiated lung. The irradiated lung produced nearly 3.5 times more prostacyclin than did the control lung. However, prostacyclin synthesis by macrophages lavaged from irradiated lung was no different than that of macrophages from sham-irradiated lung. Protein, LDH and macrophage number in BAL fluid from irradiated lungs were significantly higher than in corresponding specimens from sham-irradiated lungs. These data demonstrate that reduced levels of annexin I, as well as increased protein concentration, LDH activity and macrophage numbers in irradiated rat lung are reflected in BAL fluid. Therefore, information obtained from BAL fluid, but not from BAL macrophages, reflects lung status, and may serve as a minimally invasive index of radiation pneumonitis in this model. In irradiated lung, increased PGI2 synthesis coupled with a decreased annexin I level are consistent with the hypothesis of an inhibitory role of annexin I in prostaglandin metabolism. However, this hypothesis is not supported by findings in BAL macrophages, where increased annexin I concentration is not accompanied by a decrease in PGI2 production. In view of the latter findings, and a previous study from our laboratory demonstrating that phospholipase activity in irradiated rat lung is in fact decreased, despite the reduction in annexin I concentration and the hyperproduction of prostanoids, it would seem unlikely that annexin I negatively modulates prostaglandin synthesis via inhibition of phospholipase in this model.

6-Ketoprostaglandin F1 alpha↗
PubMed1997

Annexin I in fibrotic rat lung and cultured lung fibroblasts following irradiation.

Radiation-induced lung fibrosis is a result of collagen accumulation in the interstitium, partly due to increased collagen synthesis by fibroblasts. One feature of active collagen synthesis is increased membrane trafficking in the fibroblasts. A group of proteins called annexins is believed to play a regulatory role in membrane fusion and exocytosis. Therefore, increased annexin activity might be expected in the fibrotic lung. We tested this hypothesis by measuring annexin I levels, hydroxyproline content and ultrastructural changes in radiation-induced pulmonary fibrosis in rat. Three months after a single exposure to 30 Gy of X-rays to the right hemithorax, the right lung of the rat was atrophied and fibrotic with a concomitant increase in size of the shielded left lung. Electron micrographs revealed that the irradiated lung was ladened with interstitial collagen fibrils, with increased number of fibroblasts amongst them. Hydroxyproline concentration in the irradiated lung was nearly twice that in the sham-irradiated lung. Annexin I in the irradiated lung, on the other hand, was markedly reduced, and barely detectable on immunoblots. Since increased annexin I might precede enhanced collagen production, we also measured annexin I levels in rat lungs 3 days after 30 Gy irradiation and correlated that with hydroxyproline concentration. We found no appreciable difference in annexin I levels and hydroxyproline content between sham-irradiated and irradiated lungs at 3 days. To determine whether annexin I levels in cultured fibroblasts were altered by irradiation, we assayed annexin I in cultured rat lung fibroblasts 3 days after 0.10 Gy exposure, with concomitant measurement of 14C-proline incorporation. The annexin I level in fibroblasts irradiated with 10 Gy X-rays was 55% higher than in sham-irradiated fibroblasts. However, incorporation of 14C-proline into collagenase-sensitive macromolecules in the culture medium and extracellular matrix was not different between these two groups of cells. These data demonstrate a radiation-induced increase in immunoreactive annexin I in cultured lung fibroblasts, but fail to support the hypothesis of a positive correlation between annexin I concentration and fibrosis in irradiated rat lung.

Animals↗
PubMed1996

Isolation and sequencing of the cDNA encoding phosphatidylinositol transfer protein from rabbit lung.

The cDNA clones encoding rabbit lung phosphatidylinositol transfer protein (PI-TP) were isolated and sequenced. The putative polypeptide consisted of 270 amino acid (aa) residues, the same as human PI-TP, but one aa residue less than the PI-TP of rat and mouse. PI-TP RNA expression in various tissues of a pregnant rabbit was analyzed by Northern blot. Brain, placenta and fallopian tube had the highest PI-TP RNA expression. PI-TP RNA expression in alveolar epithelial type-II cells isolated from rabbit lung markedly increased after a 24-h culture, suggesting that PI-TP RNA expression in type-II cells can be modified by ambient factors.

Amino Acid Sequence↗
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↗
PubMed1995

Annexin I concentration, phospholipase activity and thromboxane synthesis in irradiated rat lung.

Annexin I, a member of a family of Ca(2+)-dependent phospholipid-binding proteins (PLBP), has been suggested as a regulator of prostaglandin metabolism as a result of its inhibitory effect on phospholipase A2. Synthesis of prostaglandin is increased in irradiated tissue, but the mechanism underlying this increase has not been delineated. It is conceivable that a decrease in the level of annexin I resulting in increased phospholipase activity may be responsible for the enhanced synthesis of prostaglandin. Accordingly, we measured the level of a lung 36 kDa PLBP, which possesses characteristics of annexin I, as well as the activity of phospholipase and the synthesis of thromboxane A2 (TXA2) in irradiated rat lung. The right lung of rats was irradiated with 0, 15 or 30 Gy of X rays and the animals were sacrificed after 3 months. Phospholipid binding protein was assayed by its ability to transfer unilamellar liposomes to multilamellar liposomes and by immunoblotting against anti-36 kDa rabbit PLBP antisera. Production of TXA2 by minced lung tissue was determined by radioimmunoassay of its stable metabolite TXB2. Phospholipase activity was assayed by hydrolysis of [14C]dioleoylphosphatidylcholine. Our results showed that PLBP activity in the lungs irradiated with 30 Gy was lower than that in the lungs irradiated with 0 and 15 Gy (8.82 +/- 0.47 compared to 9.73 +/- 0.49 and 9.95 +/- 0.78 nmol phospholipid transferred/mg protein, respectively). Western blotting demonstrated a near total depletion of annexin I in the lungs irradiated with 30 Gy. Phospholipase activity was also lower in the lungs irradiated with 30 Gy compared to that in the lungs irradiated with 0 Gy (0.23 +/- 0.01 vs 0.32 +/- 0.01 nmol phosphatidylcholine liberated/mg protein/min, P < 0.001). Reduced phospholipase activity was observed not only in the cytosolic or soluble fraction of lung homogenate, but also in precipitates obtained after 21,000g and 100,000g centrifugation. Despite this decline in phospholipase activity, there was a 2.8-fold increase in the synthesis of thromboxane (367 +/- 65 compared to 1076 +/- 143 pg TXB2/mg tissue/10 min for lungs irradiated with 0 and 30 Gy, respectively). These results are not consistent with the hypothesis that increased synthesis of thromboxane A2 in irradiated rat lung is a direct result of elevated phospholipase activity. In fact, phospholipase activity is decreased in the irradiated lung, despite a decline in the concentration of annexin I, its putative inhibitor.

Animals↗
PubMed1994

Immunocharacterization and developmental regulation of rabbit lung calcium-dependent phospholipid-binding proteins.

The purpose of this work was to use the immunoblotting methods to study the 36 kDa calcium-dependent phospholipid-binding protein (PLBP) in the adult and fetal rabbit lungs to gain insight into the significance of this protein in lung development. The identity of the 36 kDa PLBP and the antigen specificity of the antiserum raised against this protein in the guinea pig were tested against known annexins and antibodies to the annexins. Our results showed that the rabbit lung 36 kDa PLBP contained only one protein which cross-reacted with antibodies against annexin 1. However, the 36 kDa PLBP was slightly smaller (36 vs. 37 kDa) and more acidic (pI 6.0 vs. 6.9) than the recombinant human annexin 1. The guinea pig antiserum only reacted with annexin 1, not with any of the other annexins tested. In the cytosolic fractions of the lung and the alveolar epithelial type II cells, and in the lung lavage fluid, the 36 kDa PLBP was by far the most prominent protein with minor presence of a 33 kDa protein recognized by the guinea pig antiserum. The amount of the 36 kDa PLBP of type II cells was 55% higher than that in the lung tissue and 2.6-times higher than that in the lung lavage (9.3 +/- 0.62, 6.0 +/- 0.31 and 3.6 +/- 0.04 micrograms/mg protein, respectively). The 36 kDa PLBP appeared in the fetal rabbit lungs as early as at 21 days gestation and increased 2-fold to reach the adult level at 27 days gestation (term 31 days). The high content of PLBP in type II cells and the rapid increase in this protein in the fetal lungs at late gestations suggest an important role of the 36 kDa PLBP in lung development and surfactant biogenesis.

Animals↗
PubMed1993

The surface properties of lung 36 kDa Ca(2+)-dependent phospholipid-binding protein.

The intrinsic surface activity of a 36 kDa rabbit lung calcium-dependent phospholipid-binding protein (PLBP), a member of the annexin family of such proteins, at the air/water interface has been determined from measurements of surface tension of aqueous solutions, and surface concentration of 14C-labeled PLBP adsorbed from aqueous solution in the absence and presence of Ca2+. It was also possible to spread insoluble monolayers of PLBP to determine surface pressure vs. surface concentration isotherms, as well as surface elasticity and surface viscosity as a function of frequency from electrocapillary wave diffraction measurements. PLBP has been shown to exhibit significant intrinsic surface activity at the air/water interface, comparable to a variety of other hydrophobic proteins known to be quite surface active. In all cases, surface properties were enhanced by the presence of Ca2+, particularly the degree of surface viscoelasticity at close-packing in the monolayer. This is believed to reflect changes in protein conformation at the surface.

Animals↗
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↗