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Biomedical subjects

H C Cheng

Publications and source records attributed to H C Cheng.

At least 55 records · Page 3Linked to original sources
PubMed1997

Relative blood volume measurements by magnetic resonance imaging facilitate detection of testicular torsion.

RATIONALE AND OBJECTIVES: The authors determine the utility of relative blood volume measurements (rBV) using a blood pool marker for magnetic resonance imaging (MRI) in detection of early testicular torsion. METHODS: Testicular torsion was induced in rats by counterclockwise 720 degrees rotation and fixation of the testis in the scrotum. MPEG-PL-DTPA-Gd enhanced MRI (30 mumol Gd/kg bolus injection) was performed 1 hour after torsion at 1.5 T using fat-suppressed three-dimensional fast spoiled gradient-recalled sequence for relative blood volume measurement and three-dimensional time-of-flight sequence for MR angiography (MRA). RESULTS: The rBV of the torqued testes was significantly lower (13.3% +/- 13.5%) than that of testes with sham operation (97.7% +/- 5.3%; P < 0.05). Rats with testicular torsion showed larger regions of ischemia than did animals with sham operation (63.4% +/- 13.0% versus 4.0% +/- 2.8% of all pixels in testis; P < 0.01). The MRA of testicular torsion showed engorgement of the distal testicular vein as a sign of venous compression or total disappearance of the testicular vein, indicating arterial insufficiency. CONCLUSIONS: The authors conclude that MPEG-PL-DTPA-Gd can be used to obtain functional (rBV), morphologic (tunica enhancement), and angiographic (venous engorgement, arterial compromise) findings that should improve the diagnosis of testicular torsion in the acute setting.

Animals↗
PubMed1997

Common and differential recognition of structural features in synthetic peptides by the catalytic domain and the Src-homology 2 (SH2) domain of pp60c-src.

The relative efficiencies of the catalytic domain of the src-family kinase pp60c-src in phosphorylating four peptide substrates including (i) src-optimal peptide (AEEEIYGEFEAKKKK), (ii) "-YEEI-peptide" (KKTHQEEEEPQYEEIPIYL), (iii) cdc2(6-20) (KVEKIGEGTYGVVYK), (iv) src-autophosphorylation site peptide (ADFGLARLIEDNEYTARG) and the relative efficiencies of its SH2 domain in binding the phosphorylated forms of these peptide substrates were compared. The results show that the src-optimal peptide, "-YEEI-peptide," cdc2(6-20) peptide were phosphorylated by the catalytic domain with high efficiency and that the phosphorylated form of all three peptides could bind the SH2 domain of the kinase, confirming the hypothesis proposed by Songyang and co-workers that the catalytic domain of pp60c-src phosphorylates sites which are recognized by its own SH2 domain (Songyang et al. (1995) Nature 373, 536-539). The four peptides were phosphorylated by the kinase with relative efficiencies in the order of Src-optimal peptide > "-YEEI-peptide" > cdc2(6-20) >> src-autophosphorylation site peptide. However, the Tyr(P)-Src-optimal peptide and [pY]15cdc2(6-20) bound to the SH2 domain of the kinase with an affinity at least an order of magnitude lower than that of the tight-binding peptide, "-pYEEI-peptide." Thus, our study suggests that the catalytic and SH2 domains of pp60c-src recognize overlapping but not identical determinants in the local structure around the tyrosine phosphorylation site of the substrate peptides.

Amino Acid Sequence↗
PubMed1996

Purification of bovine thymus cytosolic C-terminal Src kinase (CSK) and demonstration of differential efficiencies of phosphorylation and inactivation of p56lyn and pp60c-src by CSK.

The C-terminal src kinase (CSK) is a ubiquitously expressed, cytosolic enzyme capable of phosphorylating and inactivating several plasma membrane-bound src-family protein tyrosine kinases in vitro [Nada, S., Okada, M., MacAuley, A., Cooper, J.A., & Nakagawa, H. (1990) Nature 351, 69-72; Bergman, M., Mustelin, T., Oetken, C., Partanen, J., Flint, N.A., Amrein, K.E., Autero, M., Burn, P., & Alitalo, K. (1992) EMBO J. 11, 2919-2924]. We purified CSK to apparent homogeneity from bovine thymus cytosol to study in vitro how the purified enzyme recognizes the various src-family kinases as its substrates. A novel assay method was developed for assaying the ability of CSK to inactivate src-family tyrosine kinases. With this assay method, we demonstrated that CSK inactivated p56lyn with a significantly higher efficiency than pp60c-src. Phosphopeptide mapping of CSK-phosphorylated p56lyn and pp60c-src shows that the consensus tyrosine residue (also termed tail tyrosine) in the C-terminal regulatory domain of p56lyn was phosphorylated by CSK with an efficiency much higher than that of pp60c-src. Thus, the higher efficiency of inactivation of p56lyn by CSK is a result of the ability of p56lyn to serve as a better substrate of CSK. The synthetic peptides derived from the C-terminal portion of p56lyn and pp60c-src were much poorer substrates than the intact src-family kinases for CSK, indicating that the local structure around the tail tyrosine is not sufficient to direct efficient phosphorylation of p56lyn by CSK. Nevertheless, the slightly higher efficiency displayed by CSK in phosphorylating the peptide derived from the C-terminal portion of p56lyn than that from pp60c-src suggests that the structural differences between the C-terminal portions of p56lyn and pp60c-src contribute to the differential efficiencies displayed by CSK in phosphorylating the two kinases. Determination of the CSK-phosphorylation site in the src-C-terminal peptide by phosphopeptide mapping reveals that the whole C-terminal regulatory domain and an adjacent part of the protein kinase domain contain some of the structural determinants directing CSK to phosphorylate the consensus tail tyrosine of the src-family kinases.

Amino Acid Sequence↗
PubMed1996

Purification and characterization of endogenous digoxin-like immunoreactive factors in chicken blood.

Studies have been performed to determine whether an endogenous material capable of binding to digoxin antibodies is present in the chicken plasma. In the blood of 12 chickens without feed control, endogenous digoxin-like immunoreactive factors (DLIF) binding of digoxin antibodies in enzyme immunoassays amounted to 866 / 302 pg digoxin equivalents/mL of plasma (mean +/- SEM). Immunoreactivity of DLIF increased to 1848***331 pg/mL with a double value of control after boiling and acid pretreating the plasma. The major purification steps employed in this report were gel filtration column chromatography, high performance liquid chromatography (HPLC) and isoelectric focusing (IEF). Using HPLC for the separation, at least 10 chicken DLIFs with different molecular weight (MW) have been found. The MW of the smallest is 300 daltons (Da) while the largest is 100 kDa. The value of the isoelectric point of the most abundant type of DLIF from untreated chicken plasma is 6.3 as determined by IEF. The partially purified DLIF inhibits Na+, K(+)-ATPase from a porcine cerebral cortex as well as three human red blood cell membrane preparations in a dose-response fashion.

Animals↗
PubMed1996

Substantial decrease of heat shock protein 90 in ventricular tissues of two sudden-death pigs with hypertrophic cardiomyopathy.

We recently developed a pig model with naturally occurring hypertrophic cardiomyopathy (HCM). To further characterize the biochemical features of high frequency of sudden death affected by this disease, the protein profiles of ventricular tissues were analyzed on normal, HCM, and HCM with sudden-death pigs. By sodium dodecylsulfate-polyacrylamide gel electrophoresis, a protein corresponding to an apparent molecular mass of 90 kDa (p90) was found to be markedly decreased in the HCM-affected tissues of sudden-death pigs, but not in HCM or normal pigs. Further study showed that the primary decrease of p90 in HCM pigs with sudden death was located mainly in the interventricular septum. As determined by the molecular mass and isoelectric point on 2-dimensional gels and Western immunoblot with a specific 90 kDa heat shock protein (HSP90) monoclonal antibody, the unknown protein was identified as HSP90. Our findings indicate for the first time that substantially decreased heart HSP90 is associated with HCM pigs with sudden death. Although the role that HSP90 may play in protecting pigs with HCM from sudden death is still nuclear, the model itself may provide further insight into understanding the role of heat shock proteins in cardiac sudden death.-Lee, W.-C., Lin, K.-Y., Chiu, Y.-T., Lin, J.-H., Cheng, H.-C., Huang, H.-C., Yang, P.-C., Liu, S.-K., Mao, S. J. T. Substantial decrease of heat shock protein 90 in ventricular tissues of two sudden-death pigs with hypertrophic cardiomyopathy.

Animals↗
PubMed1996

Viability of keratocytes in epikeratophakia lenticules.

AIM: To study the influence of cryoprotectant, cooling rate, and warming rate on recovery and viability of keratocytes from corneas for cryolathing. METHODS: Corneas were frozen at -50 degrees C for 2 minutes either after exposure to 10% dimethyl sulphoxide in Eagle's MEM for 15 minutes at room temperature (about 22 degrees C), or without earlier exposure to the cryoprotectant. Corneas were cooled either rapidly (20 degrees C/min) or slowly (1 degree C/min), and they were warmed either rapidly (> 50 degrees C/min) by direct transfer into medium at 22 degrees C or slowly (< 20 degrees C/min) in air at 22 degrees C. The cryoprotectant was removed by dilution in medium containing 0.5 mol/l sucrose. Recovery of keratocytes was determined by using collagenase digestion to release the cells from the stroma and trypan blue staining. Viability was assessed by the outgrowth of cells from stromal explants in primary tissue culture. RESULTS: The use of a cryoprotectant before freezing was beneficial, irrespective of the different cooling and warming regimens. Both collagenase digestion and tissue culture revealed that keratocyte survival was improved when corneas were warmed rapidly rather than slowly. The collagenase digestion assay showed an apparently higher recovery of keratocytes after slow cooling (54.3%) than after rapid cooling (34.1%), but no differences in cell viability could be demonstrated by primary tissue culture. CONCLUSION: Although in these experiments slow cooling apparently provided the best recovery of keratocyte numbers (though not viability), previous work had revealed some disruption of the epithelial basement membrane after slow cooling. If viable keratocytes and good preservation of epithelial basement membrane are considered to be prerequisites for epikeratophakia lenticules then it is suggested that corneas should be prepared for cryolathing by freezing rapidly after exposure to 10% dimethyl sulphoxide and, following cryolathing, they should be warmed rapidly.

Animals↗
PubMed1995

Autophosphorylation induces autoactivation and a decrease in the Src homology 2 domain accessibility of the Lyn protein kinase.

Lyn is a member of the Src family of protein-tyrosine kinases that can readily undergo autophosphorylation in vitro. The site of autophosphorylation is Tyr397 which corresponds to the consensus autophosphorylation site of other Src family tyrosine kinases. The rate of autophosphorylation is concentration-dependent, indicating that the reaction follows an intermolecular mechanism. Autophosphorylation results in a 17-fold increase in protein-tyrosine kinase activity. Kinetic analysis demonstrates that phosphorylation of a substrate peptide by Lyn following autophosphorylation occurs with a 63-fold decrease in Km but no significant change in Vmax, suggesting that autophosphorylation relieves the conformational constraint that prevents binding of the substrate peptide to the active site of the kinase. Using a phosphotyrosine-containing peptide (pYEEI) that has previously been shown to bind to the Src homology 2 (SH2) domain of Src family tyrosine kinases with high affinity, we found that autophosphorylation results in a significant decrease in accessibility of the Lyn SH2 domain, indicating that conformational changes in the protein kinase domain induced by autophosphorylation can be propagated to the SH2 domain. Our study suggests that autophosphorylation plays an important role in regulating Lyn by modulating both its kinase activity and its interaction with other phosphotyrosine-containing molecules.

Amino Acid Sequence↗
PubMed1995

Characterization of two activated mutants of human pp60c-src that escape c-Src kinase regulation by distinct mechanisms.

Two activated transforming mutants of human pp60c-src were found to possess single point mutations within the regulatory carboxyl terminus (E527K in CY CST201) and the kinase domain (E381G in WO CST1), respectively, that do not directly interfere with either the regulatory c-Src kinase (CSK) phosphorylation site (Tyr530) or the SH2/3 domains. In vivo, both mutant proteins are hypophosphorylated on their carboxyl-terminal regulatory tyrosines and are hyperactive. In an in vitro Src kinase inactivation assay, both mutant Src proteins exhibited resistance to inactivation by CSK relative to wild-type Src. Under these in vitro conditions, E381G c-Src was found to be phosphorylated by CSK to wild-type levels, while E527K c-Src was not detectably phosphorylated. The ability of CSK to phosphorylate a carboxyl-terminal peptide modelled against E527K c-Src was also impaired, suggesting that CSK is unable to recognize E527K c-Src as an efficient substrate. In the case of E381G c-Src, examination of whether its SH2/3 domains were accessible to the carboxyl-terminal regulatory phosphotyrosine revealed a highly reduced ability of autophosphorylated E381G c-Src to bind to a synthetic phosphopeptide modelled from the SH2-binding region of polyoma middle-T antigen which binds to Src SH2 with high affinity. This suggests that the E381G c-Src mutation results in an altered or reduced accessibility of the SH2 domain of the autophosphorylated form of E381G c-Src and may represent a previously undescribed mode of Src activation. Further study of these and other Src mutants may offer additional new insights into the regulation of "Src family" kinases.

Amino Acid Sequence↗
PubMed1995

Skull morphology affected by different sleep positions in infancy.

In infancy, prior to cranial suture and fontanel calcification, the craniofacial skeleton can be easily deformed by an externally exerted force. In this study, the relationship between the sleep position and skull morphology was investigated. A group of 81 cleft lip and/or palate infants without other systemic anomalies was first seen in the craniofacial center at approximately 1 month of age. The sleep position of each infant was recorded as supine, prone, or mixed type. The body and skull growth were longitudinally measured at 1, 3, and 6 months of age. Infants sleeping in the supine sleep position tended to have a wider head width, shorter head length, and a larger cephalic index by 6 months of age. The opposite phenomena were observed in the prone sleep group. The mixed sleep group tended to have head width, head length, and cephalic index between those of the supine sleep group and the prone sleep group. During the first 3 months of life, the sleep position could mold the skull primarily in the dimension of head width. In conclusion, the supine sleep position may promote brachycephaly and the prone sleep position dolichocephaly.

Age Factors↗
PubMed1995

Mechanism of c-SRC activation in human melanocytes: elevated level of protein tyrosine phosphatase activity directed against the carboxy-terminal regulatory tyrosine.

Normal human melanocytes, and some human melanoma cell lines, contain c-SRC which is constitutively activated by hypophosphorylation of tyrosine 530. We investigated the possibility that the activation of c-SRC in melanocytes might be attributable to elevated levels of tyrosine 530-directed protein tyrosine phosphatase activity in these cells. We found three times more of this phosphatase activity in cell extracts from melanocytes compared to fibroblasts. The majority of the tyrosine 530-dephosphorylating activity was present in the particulate fraction of cell homogenates, where c-SRC is also located. Treatment of melanocytes with the protein tyrosine phosphatase inhibitor, sodium orthovanadate, caused inactivation of c-SRC. From these results, we conclude that activation of c-SRC in human melanocytes may be attributed to an elevated level of protein tyrosine phosphatase activity directed against tyrosine 530.

Amino Acid Sequence↗
PubMed1995

Establishment of in vivo hepatoma models in rat and mouse from rodent hepatoma cell lines.

BACKGROUND: Hepatoma is one of the most common cancers in Southeast Asia and African countries. In Taiwan, it is the leading cause of death in male cancer patients. In order to examine the effect of various factors on the growth of hepatoma, in vivo hepatoma models such as carcinogen-induced hepatoma and subcutaneous implantation of hepatoma in nude mice have been used. However, there are disadvantages in these models. METHODS: Rats and mice were anesthetized by ketamine or ether, respectively. After a midline incision was made, N1S1 rat hepatoma cells were injected intrasplenically to partially hepatectomized or sham-operated rats, while BALB/c mice received intrasplenic injection of ML-2 and ML-3 mouse hepatoma cells. For direct tumor implantation, a 1mm3 N1S1 tumor piece was implanted in liver of Sprague-Dawley (SD) rat using a trocar. Animals were sacrificed at specific times after tumor implantation. Tumor incidence and the number of tumor nodules on the liver surface were recorded. Tumor samples were fixed and embedded for histological examination. RESULTS: After intrasplenic implantation of ML-2 cells, no tumor was observed on the liver in any of the 10 mice 40 days later. In comparison, rapid growth of hepatic ML-3 tumors was observed in all animals. Rat hepatoma cells RH-35, McA-RH7777 and McA-RH8994 cells did not form tumors in SD rats. The tumorigenicity of N1S1 cells in SD rats was dose-dependent on implanted tumor cells. In addition, hepatic N1S1 tumors could be obtained within a few weeks by homograft. CONCLUSIONS: We have successfully established in vivo hepatoma models in both the rat and the mouse. The murine ML-3 cells generated hepatoma in syngeneic BALB/c mice while the tumorigenicity of N1S1 cells in partially hepatectomized SD rats was dose-dependent on implanted tumor cells. These in vivo rodent models will be valuable tools for future studies of hepatoma.

Animals↗
PubMed1994

Maxillary dental arch affected by different sleep positions in unilateral complete cleft lip and palate infants.

The development of the dental arch is well designed for adaptive and compensatory growth. In this study, the relationship between the sleep position and dental arch development was investigated. A group of 42 infants with unilateral complete cleft lip and palate with either prone (16) or supine (26) sleep position were seen in the craniofacial center. All infants were less than 1 month of age at the initial visit. Dental impressions of the maxillary arch were taken at the initial visit and just before cheiloplasty. Ten arch dimensions were measured in each dental cast and the longitudinal change in each dimension was compared between the prone sleep group and the supine sleep group. Statistically significant changes were detected in the growth rate of the following dimensions: intercanine width, intertuberosity width, alveolar cleft width, anterior cleft width, and posterior cleft width. This study indicated that sleep position affected maxillary arch development. Infants sleeping in the prone sleep position tended to have narrower arch width and cleft width.

Analysis of Variance↗
PubMed1994

[Physiological changes in the metabolism of football trainees following a match under high temperature].

The effects of a football match on the muscle metabolism of long-term football players were studied. Twenty-two senior high school students (Age: 16.95 +/- 0.82 y/o) were divided into two groups. Group A, included the subjects with longer histories of playing football (7 years) and group B were those with a shorter histories (6 years). There was no statistical difference between the two groups (p > 0.05). The temperature of all muscles tested showed statistical differences after exercise (p < 0.05). After 24 hours, all muscles except the gluteus maximus returned to the normal state while the skin temperature of the latissimus dorsi did not have any change. Body temperature, pulse rate, respiratory rate, and diastolic pressure all significantly increased immediately after the match (p < 0.05) and they all returned to their normal resting rate within one hour, except the body temperature, which required two hours to return to normal. A comparison between groups A and B showed a significant difference (p < 0.01) in creatine kinase level immediately and one hour after the match. It is inferred that energy is derived from the aerobic system.

Adolescent↗
PubMed1993

The N-terminal hydrophobic domain of P450c21 is required for membrane insertion and enzyme stability.

Microsomal cytochromes P-450 are known to be integrated into smooth endoplasmic reticulum through their hydrophobic sequences located at the N termini. The length requirement of the membrane insertion signal was determined by the generation of six plasmids encoding mutant P450c21 that lacked various portions of the N-terminal hydrophobic domains. When they were transcribed and translated in vitro in the presence of endoplasmic reticulum membranes, mutant protein lacking more than a third of the first hydrophobic domain gradually lost the ability to insert into the membrane and stayed mostly in the soluble fraction when the first N-terminal hydrophobic domain was removed. The steady-state amount of the truncated proteins was progressively reduced in parallel to the extent of their N-terminal deletions, due to their fast degradation. This process was accompanied by a decrease in the enzymatic activity. Therefore, the first hydrophobic domain of P450c21 not only serves as a membrane targeting and anchoring domain, but it is also important for the in vivo protein stability.

Animals↗
PubMed1993

In vitro substrate specificity of protein tyrosine kinases.

Synthetic peptides such as P60src autophosphorylation site peptides and angiotensin are indiscriminately phosphorylated by protein tyrosine kinases. The observation has led to the general belief that protein tyrosine kinases are highly promiscuous, displaying little in vitro site specificity. In recent years, evidence has been accumulating to indicate that such a belief requires close examination. Synthetic peptides showing high substrate activity for specific groups of protein tyrosine kinases have been obtained. Systematic modification of certain substrate peptides suggests that kinase substrate determinants reside with specific amino acid residues proximal to the target tyrosine. A number of protein kinases have been shown to be regulated by tyrosine phosphorylation at specific sites by highly specific protein tyrosine kinases. These and other selected biochemical studies that contribute to the evolving view of in vitro substrate specificity of protein tyrosine kinases are reviewed.

Amino Acid Sequence↗
PubMed1993

Zonation of cholesterol and glycerolipid synthesis in regenerating rat livers.

Hepatic zonation of cholesterol and glycerolipid synthesis was investigated in regenerating rat livers 24 hr after partial hepatectomy. Tritiated acetate and [U-14C]glycerol were injected into rats' peritoneal cavities for a short-term labeling study. Periportal and perivenous hepatocytes were isolated by digitonin collagenase perfusion. Cholesterol synthesis was significantly higher in periportal hepatocytes of the sham-operated livers (periportal/perivenous = 1.67; p < 0.05). Twenty-four hours after partial hepatectomy, cholesterol synthesis was selectively decreased by 40% (p < 0.01) in periportal hepatocytes. Consequently, hepatic zonation of cholesterol synthesis was abolished in regenerating livers. To study the cholesterol homeostasis on a long-term basis, we substituted deuterated water (25% enriched) for drinking water for 5 days to label newly synthesized cholesterol in a steady state. This procedure clearly demonstrated the net negative cholesterol balance 24 hr after partial hepatectomy. However, the newly synthesized cholesterol contributed equally to the cellular cholesterol pool in both zones. The synthesis of glycerolipids, whether measured from tritiated acetate or [U-14C]glycerol, was significantly increased without apparent zonation in the regenerating livers (twofold increase in phospholipid, and threefold to sevenfold increase in triacylglycerol). We concluded that hepatic zonation of cholesterol synthesis is caused by higher de novo synthesis in periportal hepatocytes, which is abolished in regenerating livers. No zonation of glycerolipid synthesis exists in normal and regenerating livers.

Acetates↗