Biomedical subjects

T M Chiang

Publications and source records attributed to T M Chiang.

At least 37 records · Page 2Linked to original sources

PubMed1996

Effect of monoclonal antibody on the adhesion of platelets to aortic segments.

A purified monoclonal antibody raised against a purified platelet 65 kDa protein that binds to type I collagen receptor was used to see if the antibody can block the adhesion of platelets to damaged aortic segments. The purified antibody binds to the surface of human platelets by flow cytometry. The adhesion of platelets to the damaged aortic segments was significantly reduced by the pre-incubation of the platelet-rich plasma with the antibody, suggesting that the antibody binds to platelets and blocks the adhesion of platelets to the aortic segments.

Antibodies, Monoclonal↗

PubMed1996

Upregulation of nitric oxide synthase in cultured human keratinocytes after ultraviolet B and bradykinin.

Ultraviolet B (UVB) irradiation of the skin has been reported to upregulate nitric oxide synthase (NOS) activity with enhancement of nitric oxide (NO) formation. Bradykinin, a known stimulator of NO production, is produced in the skin within minutes of UVB irradiation. The combined effect of UVB and bradykinin on NOS was therefore examined in a cultured human keratinocyte (KC) line. Activity was determined in KC homogenates by the recovery of [3H]L-citrulline using labeled L-arginine as the substrate in the presence of mM NADPH. Monoclonal antibodies to specific isoforms of NOS that cross-react with their human counterparts were used to determine the isoform(s) in control, UVB, bradykinin treated and UVB and bradykinin treated KC. Human KC express NOS activity which is lowest at confluence and highest during proliferation. UVB increased NOS activity when a set dose of irradiation was administered from 32.2-48.3 mJ/cm2 but was inhibitory after 64.4 and 80.5 mJ/cm2. Thirty min after 10(-6) M bradykinin, NOS activity nearly doubled followed by return of activity to control levels at 60 min. Activity after UVB and bradykinin was only slightly higher than that observed with bradykinin alone. Immunochemically, an isoform of M(r) 155 kDa was detected in control cells with the antibody for the constitutive brain enzyme, bNOS. Recovery of this isoform increased after UVB treatment as well as after bradykinin which was time dependent. When both stimulants were used, the recovery of the 155 kDa enzyme was markedly enhanced, unlike the enzyme activity findings. These data indicate that the expression of NOS activity under unstimulated conditions in human KC in culture is due to the constitutive NOS found in neuronal tissue, bNOS. The recovery of bNOS increased after UVB and after bradykinin while the combination of both resulted in the synergistic increase in bNOS protein with only a marginal further increase in NOS activity.

Antibodies, Monoclonal↗

PubMed1994

Activation of phospholipase D in human platelets by collagen and thrombin and its relationship to platelet aggregation.

Stimulation of phospholipase D after activation of cell surface receptors has been reported in many cell types. We have investigated the mechanism of activation of this enzyme by collagen in the human platelet by assaying the release of [3H]methylcholine from [3H]methylphosphatidylcholine. Results from these studies suggest that phospholipase D activity is regulated by reversible phosphorylation. Phospholipase D activity was stimulated when platelet-rich plasma was preincubated with collagen and was not inhibited by aspirin. Among various aggregating agents tested, collagen and thrombin but not ADP activated phospholipase D activity (2- to 3-fold). The addition of sphingosine inhibited phospholipase D activity. Preincubation of platelet-rich plasma with sphingosine inhibited collagen- and thrombin-induced platelet aggregation and the release of ATP. The inhibitory effect of sphingosine on collagen- and thrombin- induced platelet aggregation and release of ATP was dose-dependent. The functional significance of phospholipase D activation was also tested by examining the effect of the product, phosphatidic acid, on collagen-induced platelet aggregation and release of ATP. Platelet shape change and the reversibility of platelet aggregation resulted by the addition of phosphatidic acid to platelet-rich plasma. Furthermore, the simultaneous addition of phosphatidic acid and collagen shortened the latency period but had no effect on platelet aggregation. Two platelet proteins (47 kDa and 22 kDa) increased in phosphorylation after the addition of 1 microM phosphatidic acid which did not cause platelet aggregation. These results suggest that collagen stimulates phospholipase D activity which plays a secondary role in platelet aggregation and the release reaction.

Adenosine Triphosphate↗

PubMed1993

Collagen-platelet interaction: separate receptor sites for types I and III collagen.

We have isolated a platelet membrane protein of M(r) 47 kDa which is responsible for the interaction of platelets with type III collagen. The 47 kDa protein was purified to apparent homogeneity by type III collagen-Sepharose 2B column chromatography and preparative slab gel electrophoreses. The 47 kDa protein blocked the adhesion of platelets to type III but not to type I collagen. Polyclonal antibodies were obtained from rabbits immunized with the purified 47 kDa protein emulsified in complete Freund's adjuvant. The polyclonal antibodies inhibited the type III collagen but not type I collagen-induced platelet aggregation. The inhibitory effect of the antibodies on type III collagen-induced platelet aggregation was dose-dependent. Cross-inhibition on platelet aggregation studies showed that type I collagen receptor antibodies (M(r) 65 kDa) did not inhibit type III collagen-induced platelet aggregation and type III collagen receptor antibodies did not inhibit type I collagen induced platelet aggregation. These results suggest that type I and type III collagens interact with platelets at separate sites.

Animals↗

PubMed1993

The role of protein phosphatases 1 and 2A in collagen-platelet interaction.

Okadaic acid, a specific inhibitor of phosphoserine/threonine protein phosphatases 1 and 2A, was used to determine whether these protein phosphatases play a role in collagen-induced platelet aggregation and release reaction as measured by ATP release. Collagen-induced platelet aggregation and ATP release were inhibited by the addition of okadaic acid to platelet-rich plasma in a dose-dependent manner. The inhibitory effect of okadaic acid on collagen-induced platelet aggregation correlated with phosphorylation of proteins with M(r) 14.4, 25, 32, 36, 50, 60, and 80 kDa. The 14.4-kDa protein was purified to apparent homogeneity by electroelution from gel slices. This protein reacted with antibodies to phospholipase A2 (PLA2). Since okadaic acid inhibited PLA2 activity in platelet-rich plasma but not in the PLA2 assay mixture, the effect appears to be indirect. Furthermore, using a combination of immunoprecipitation and measurement of enzyme activity, PLA2 activity was inhibited in the presence of okadaic acid. The inhibited activity could not be restored by the addition of collagen. These results suggest that the phosphorylated form of PLA2 is inactive. Using [32P]glycogen phosphorylase a as substrate, protein phosphatase activity was inhibited by okadaic acid in a concentration-dependent manner. An immunoblot of platelet homogenates with anti-protein phosphatase 1 showed a band with M(r) 50 kDa reacting with the antibodies, suggesting that the 50-kDa protein is protein phosphatase 1. These data clearly show that okadaic acid increases the phosphorylation and indirectly decreases the activity of PLA2, but whether inhibition of PLA2 activity is related to collagen-induced platelet aggregation and release reaction remains to be determined.

Adenosine Triphosphate↗

PubMed1993

[Clinical observation and analysis of febrile children].

Fever is an indication of acute disease, and needs special attention. To better understand the causes of fever in children, we analysed the records of 100 febrile children who were admitted to this Hospital from June 1987 to December, 1989. In these 100 cases, 7 days' fever prior to admission included 32 cases (32%); 7 to 14 days' fever prior to admission, 43 cases (43%); over 14 days' pre-admission, 25 cases (25%). The age distribution was younger than 1 month, 3 cases; 1 month to 1 year, 52 cases; 1 year to 2 1/2 years, 20 cases; 2 1/2 years to 5 years, 16 cases; 5 to 10 years 6 cases; older than 10 years included 3 cases. Post-admission diagnoses of these children showed most had an infectious disease: 86 cases; "summer fever": 5 cases; perineal abscess: 1 case; cellulitis: 1 case; leukemia, 2 cases; 5 were from other causes. Fevers were more prevalent in summer. In conclusion, the most common cause of prolonged fever in children is infectious disease 86%, with the highest incidence of age distribution under 2 1/2 years old, with 73%. The case distribution showed a prevalence in summer 43%. Almost all prognoses were good.

Age Factors↗

PubMed1992

Okadaic acid and vanadate inhibit collagen-induced platelet aggregation; the functional relation of phosphatases on platelet aggregation.

The different specific inhibitors for phosphoserine/threonine and phosphotyrosine protein phosphatases were used to study the role of these protein phosphatases in collagen-platelet interaction. The collagen-induced platelet aggregation and the release reaction as measured ATP release were inhibited in a dose-dependent fashion by the addition of okadaic acid, a specific inhibitor of phosphoserine/threonine protein phosphatase 1 and 2A. The inhibition was also observed by the addition of a phosphotyrosine protein phosphatase inhibitor, vanadate. Suboptimal concentrations of both inhibitors together also inhibited collagen-induced platelet aggregation and release reaction in a concentration-dependent fashion. These results suggest that collagen-platelet interaction is modulated by both protein phosphatases.

Adenosine Triphosphate↗

PubMed1991

Activation mechanisms of platelet-activating factor in U937 cells: possible involvement of protein kinase C.

We have previously demonstrated that platelet-activating factor (PAF) binds specifically on cell membranes isolated from U937 cells. We now describe biological evidence showing that the effect of PAF on U937 cells is a receptor-mediated event. myo-[3H]Inositol-labeled U937 cells were used to investigate the possible role of phosphoinositide metabolism in these cells after binding of PAF. Formation of inositol phosphates (IP1, IP2, and IP3) in response to PAF was increased two- to threefold more than in vehicle control in U937 cells. The effect of PAF on endogenous protein phosphorylation was also studied by using 32PO4-labeled cells. PAF stimulates the phosphorylation of a 45-kDa protein in a time-dependent and dose-related fashion. Since the phospholipase C-generated diglyceride is an important activator of protein kinase C, the phosphorylated 45-kDa protein could be the substrate of protein kinase C. In this regard, we were able to demonstrate that phorbol ester enhances the phosphorylation of the same 45-kDa protein band. In addition, sphingosine, a protein kinase C inhibitor, inhibits the phosphorylation of the same 45-kDa protein band. Down-regulation of the protein kinase C also inhibits the 45-kDa protein phosphorylation. These results suggest that protein kinase C is involved in the PAF-U937 cell interaction.

Diglycerides↗

PubMed1991

Stimulation of phospholipase A2 activity in human platelets by trypsin and collagen.

Type I collagen enhanced human platelet phospholipase A2 activity whether added to platelet-rich plasma or washed platelets. The stimulatory effect of type I collagen on platelet membrane phospholipase A2 activity was also observed in a cell-free system utilizing platelet membranes. The release of arachidonic acid was enhanced by types I and III but not by type V collagen. The activation of platelet phospholipase A2 by type I collagen was inhibited by soybean trypsin inhibitor and mimicked by trypsin. However, type I collagen addition was not associated with any detectable changes in platelet membrane proteins while trypsin altered many proteins. These results point to acid soluble phospholipase A2 activity of platelets as an enzyme activated by type I collagen.

Blood Platelets↗

PubMed1991

[Comparison of Bactec radiometric system and Lowenstein-Jensen medium for detection of mycobacterium tuberculosis and the susceptibility of isolates to antituberculosis drugs].

Tuberculosis is a chronic, durable but curable disease. Active cases are usually contagious and spread easily to the close intimates, especially in children causing catastrophic complications. Thereafter, early diagnosis and adequate treatment is critical for the control of the disease. Bactec 460 12B medium (Bactec system) provides a rapid, accurate and reliable method for the detection and differentiation of mycobacterium tuberculosis. It is better than the traditional method, Lowenstein-Jensen (L-J) medium. To prove this point, from Sept. 1990 to Jan. 1991, 341 specimens were sent to our laboratory for detection of mycobacterium tuberculosis. Each sample was cultured in both Bactec system and L-J medium. Meanwhile, each isolate was also cultured for the susceptibility to the antituberculosis drugs. In those 341 specimens, 45 samples were cultured as positive in either media. 43 samples were positive from Bactec system, the positive rate was 12.6% and the sensitivity rate was 95.6%. The positive cultures from L-J medium were 27 samples, the positive rate was 7.9% and the sensitivity rate was 60% (p less than 0.05). The positive culture duration in Bactec system was 5-33 days, the average was 13.05 days. But in L-J medium, it was 19-45 days and the average was 27.48 days (p less than 0.01). The susceptibility of the isolates to the common-use antituberculosis drugs was as follows: isoniazid, 47.8%; rifampin, 84.1%; ethambutol, 71%; streptomycin, 82.6%; (p less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Antitubercular Agents↗

PubMed1991

[Escherichia coli strains and their susceptibility to antibiotics in nosocomial infection].

Nosocomial infection may cause severe diseases and is sometimes life-threatening. Such infection may be caused by different bacteria strains, but in our pediatric ward, a large number of cases are infected by E. coli. We studied the cases hospitalized during the period from January 1986 to December 1988 in pediatric ward, sick baby room, baby room or intensive care unit. The most common microorganism of nosocomial infection was found to be Staphylococcus aureus (37 strains), followed by E. coli (32 strains). However, bacteremia, was more often caused by E. coli (13 strains) than by Staphylococcus aureus (4 strains). In patients with E. coli nosocomial infection the ratio of male to female was 2:1. Of them, 87.5% were below 1 month old, and 6.25% were from 1 month to 1 year or from 5 to 10 years of age. The antibiotics sensitivity in bacteremia strains of E. coli seemed to be the same as that in non-bacteremia strains. Both of them were less sensitive to aminoglycoside than the community-acquired strains. Their sensitivity to 3rd-generation cephalosporins was 100%.

Anti-Bacterial Agents↗

PubMed1991

[Pediatric bacteremia strains grow in blood culture media].

Bacteremia strains may influence the clinical therapy, the isolation of bacteremia strains is important for patients. In order to understand the distribution and the growth in the blood culture media of bacteremia strains, from January 1982 through December 1988 we studied the blood culture sent to us from the pediatric ward. These were three hundred and forty one bacterial strains (3.1%) isolated from patients. In the meantime we also studied the bacteremia strains growth in the blood culture media. In these positive cultures, there were 83 gram-positive, and 258 gram-negative strains. The 83 gram-positive included Staphylococcus aureus (38 strains), hemolytic streptococcus (22 strains), Enterococcus (13 strains), Streptococcus pneumoniae (8 strains) and others (2 strains). The 258 gram-negative strains included Escherichia coli (97 strains), Salmonella spp. (43 strains), Enterobacter spp. (31 strains). Klebsiella spp. (23 strains), Pseudomonas spp. (25 strains), Proteus spp. (17 strains), Citrobacter spp. (11 strains) and others (11 strains). The Growth of bacteremia strains in the blood culture medium, showed that first day isolation strains were 33.7%, second day strains 24.9%, third day were 19.9%, fourth day were 13.5%, fifth day were 3.2%, sixth day were 4.1% and seventh day were 0.6%. In conclusion, the bacteremia strains showed that the gram-negative, which were 75.7%, were more prevalent than the gram-positive which were 24.3%, the isolation of the strains was more in the first to fourth day and grew by 92%, on the fifth day it grew by 3.2% and on the seventh day by only 0.6%. On the fifth day the baby had recovered and we were able to discontinue antibiotic therapy.

Blood↗

PubMed1990

Collagen-platelet interaction: type XI collagen-induced platelet aggregation.

Type XI collagen in its native fibrillar but not in soluble monomeric form mediates human platelet aggregation and release of adenosine triphosphate in a dose-dependent manner. Its action is inhibited by aspirin. Type XI collagen also increased radiolabelled phosphate incorporation into protein bands with molecular weights of 42 KDa and 22 KDa, respectively. In contract, these events were not observed in platelets incubated with type IX collagen. These results suggest that the fibrillar type XI collagen has the same ability as other types of collagen to induce human platelet aggregation.

Adenosine Triphosphate↗

PubMed1990

Activation of cyclic AMP phosphodiesterase by phorbol and protein kinase C pathway: differences in normal and diabetic tissue.

Diabetes mellitus is associated with high levels of adenosine 3',5'-cyclic monophosphate in tissue and plasma. Diabetes inhibits and insulin stimulates and restores low Km adenosine 3',5'-cyclic monophosphate phosphodiesterase activity. We recently reported that phorbol ester, a tumor promoting agent known to act through protein kinase C also stimulates phosphodiesterase. Here, we address the issue of whether or not the activation of phosphodiesterase by insulin and phorbol ester is different in streptozotocin diabetic adipose tissue. Rat adipose tissue was incubated with insulin, phorbol ester or other known components or effectors of the protein kinase C pathway, i.e. 1,2 dioleoyl-glycerol, 1- oleoyl, 2- acetylglycerol, Ca(++)-Ionophore A 23187, and nifedipine. After incubation, preparation and assay of adenosine 3',5'-cyclic monophosphate phosphodiesterase was made. As in previous data streptozotocin-diabetes inhibits basal phosphodiesterase by about 50% (P less than .02); insulin and phorbol ester each stimulate phosphodiesterase, in streptozotocin-diabetes less than normal (P less than .025); nifedipine inhibits phorbol stimulated phosphodiesterase in streptozotocin-diabetes but not normal (P less than .001); and nifedipine inhibits insulin stimulated phosphodiesterase in normal (84%) and diabetic (97%) (P less than .005). In normal and diabetic tissue, diacyl glycerol and oleoyl-acyl glycerol stimulate phosphodiesterase, are augmented by ionophore and inhibited by nifedipine. In addition 32P incorporation studies and measurements of tyrosine kinase activity are presented which support these differences between normal and diabetic. In summary then, these data suggest common pathways of activation for low Km adenosine 3',5'-cyclic monophosphate phosphodiesterase by insulin and phorbol ester; imply a relationship between two second messenger systems, phosphoinositides and adenosine 3',5'-cyclic monophosphate; and demonstrate a difference in activation of phosphodiesterase between normal and diabetic adipose tissue.

3',5'-Cyclic-AMP Phosphodiesterases↗

PubMed1989

The effect of low dose of 12-O-tetradecanoyl-phorbol-13-acetate on collagen platelet interactions.

We and others have reported that phorbol ester doses ranging from 0.25 to 25 micrograms/ml induce human platelets to aggregate and release serotonin. In this paper, we report the effect of low doses of phorbol (0.5 to 50 ng/ml) on subthreshold amounts of collagen induced platelet aggregation. The platelet aggregation induced by the addition of subthreshold amounts of collagen can be enhanced by low doses of phorbol. The combined low doses of phorbol and subthreshold amounts of collagen induced platelet aggregation can be inhibited by the addition of aspirin and imidazole. The increase in platelet aggregation induced by the combined low doses of phorbol and subthreshold amounts of collagen is probably mediated by the collagen fibril formation.

Aspirin↗

PubMed1989

Serine and tyrosine protein kinase activities in Streptococcus pyogenes. Phosphorylation of native and synthetic peptides of streptococcal M proteins.

Two forms of protein kinase activity were isolated from crude extracts of Streptococcus pyogenes and partially purified by ion exchange chromatography and affinity chromatography. The phosphorylation activities were shown to be insensitive to cAMP, required the presence of divalent cations, and eluted from a Sephadex G-200 column with approximate molecular masses of 60 and 45 kDa, respectively. Both enzymes were capable of phosphorylating eukaryotic proteins and synthetic polypeptides in addition to endogenous and heterologous prokaryotic proteins at serine and tyrosine residues. Firm evidence for tyrosine kinase activity was obtained by the use of a tyrosine kinase-specific substrate, a 4:1 glutamate:tyrosine copolymer. Both protein kinases phosphorylated HPr, a phosphocarrier protein of the phosphotransferase system isolated from S. pyogenes and Bacillus stearothermophilus, but failed to phosphorylate HPr isolated from Escherichia coli. Both also phosphorylated a native polypeptide fragment (pep M24) as well as synthetic peptide copies of M protein, the major virulence determinant of group A streptococci. These results indicate that prokaryotic protein kinases are capable of phosphorylating eukaryotic proteins and suggest that the protein kinases of streptococci may play an important role not only in the phosphotransferase system but also in the virulence properties of these organisms.

Amino Acid Sequence↗

PubMed1989

Collagen-platelet interaction: inhibition by a monoclonal antibody which binds a 90,000 dalton platelet glycoprotein.

Polyclonal antiserum prepared to purified type I collagen receptor, a 65,000 molecular weight protein, isolated from human platelets reacted with two other proteins with molecular weights of 90,000 and 58,000 in immunoblots of solubilized platelet membranes. The immunoreactive proteins were purified to homogeneity with molecular sieve chromatography and preparative gel electrophoreses. Periodic acid Schiff stain showed that both the 90,000 and 65,000 proteins were glycoproteins. These purified proteins reacted with immunoglobulin G (IgG) fractions isolated from antiserum raised against the 65,000 protein and poly- and monoclonal antibody specific for the glycoprotein IIb-IIIa suggesting that these three proteins are immunocross reactive with GPIIb-IIIa. To further examine the immunocross-reactivity of these proteins, a monoclonal antibody was raised against the 90,000 glycoprotein. This monoclonal antibody also reacted with all three proteins in enzyme-linked immunosorbent assays and transblot experiments suggesting that these three possess a common antigenic determinant. The monoclonal antibody prepared to the 90,000 glycoprotein also inhibited platelet aggregation induced by the addition of collagen and ADP but not alpha-thrombin and epinephrine-induced aggregation. This suggests that the monoclonal antibody binds a protein on the platelet surface which plays a role in platelet aggregation induced by the addition of these agonists.

Acetylglucosaminidase↗

PubMed1988

Phosphorylation of the outer surface of platelets enhances the effects of collagen on aggregation, ATP release, calcium translocation and phosphoinositide hydrolysis.

Phosphorylation of the outer surface of human platelets increases their functional responsiveness to subthreshold amounts of collagen. Collagen-stimulated platelet aggregation, release of ATP and changes in cytoplasmic ionized calcium levels were all enhanced by pretreatment with ATP plus a kinase purified from human plasma. [3H]-myo-inositol-labeled human platelets were used to investigate the possible role of phosphoinositide metabolism in mediating this enhanced responsiveness. Formation of inositol mono-, bis-, and trisphosphate in response to collagen was more pronounced in phosphorylated platelets than in control platelets. Collagen-stimulated inositol phosphate production in both control and phosphorylated platelets was completely inhibited by the addition of 20 microM indomethacin. Of a range of agents that stimulated phosphoinositide hydrolysis, only the response to collagen was enhanced by phosphorylation. The plasma protein kinase was shown to catalyse phosphorylation of the collagen receptor. These data indicate that the enhanced phosphoinositide hydrolysis resulting from phosphorylation of the collagen receptor might be linked to the increased functional responses to collagen.

Adenosine Triphosphate↗