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Biomedical subjects

L J Yang

Publications and source records attributed to L J Yang.

At least 19 recordsLinked to original sources
PubMed2001

Preparation and activity of conjugate of monoclonal antibody HAb18 against hepatoma F(ab')(2) fragment and staphylococcal enterotoxin A.

AIM: To prepare the conjugate of staphylococcal enterotoxin A (SEA) protein which is a bacterial SAg and the F(ab')(2) fragment of mAb HAb18 against human hepatocellular carcinoma (HCC), and identify its activity in order to use SAg in the targeting therapy of HCC. METHODS: MAb HAb18 was extracted from the abdominal dropsy of Balb/c mice, and was purified through chromatography column SP 40HR with Fast protein liquid chromatography (FPLC) system. The F(ab')(2) fragment of mAb HAb18 was prepared by papainic digestion method. The conjugate of mAb HAb18 F(ab')(2) fragment and SEA was prepared with chemical conjugating reagent N succinimidyl 3 (2-pyridyldithio) propionate (SPDP) and purified through chromatography column Superose 12 with FPLC system. The molecular mass and purity of each collected peak were identified with SDS-PAGE assay. The protein content was assayed by Lowry's method. The antibody activity of HAb18 F(ab')(2) against HCC in the conjugate was identified by indirect immunocytochemical ABC method, and the activity of SEA in the conjugate to activate peripheral blood mononuclear cells (PBMC) was identified with MTT assay. RESULTS: The IgG mAb HAb18 was extracted, and purified successfully. Immunocytochemical staining demonstrated that it reacted with most of HHCC cells of human HCC cell line. There were two peaks in the process of purification of the prepared HAb18 F(ab')(2) SEA conjugate. SDS-PAGE assay demonstrated that the molecular mass of the first peak was about 130 ku, and the second peak was the mixture of about 45 ku and a little 100 ku proteins. The immunocytochemical staining was similar in HAb18 F(ab') (2) SEA conjugate and HAb18 F(ab') (2), i.e.the cytoplasm and/or cell membranes of most HHCC cells were positively stained. The MTT assay showed that the optical absorbance (A) value at 490 nm of HAb18 F(ab') (2) SEA conjugate was 0.182 +/- 0.012, that of negative control was 0.033 +/- 0.009, and there was significant difference between them (P < 0.05). CONCLUSION: SPDP is a good protein conjugating reagent and can be used in preparing protein conjugate. The conjugate of mAb HAb18 F(ab') (2) fragment and SEA protein was prepared successfully in present study and can be used in the experimental study of HCC targeting therapy with the conjugate of SAg and anti HCC mAbs or their fragments.

Animals↗
PubMed2001

Extraction and purification of TGFbeta and its effect on the induction of apoptosis of hepatocytes.

AIM: To extract and purify the transforming growth factor beta (TGF beta), and to demonstrate its biological activity in vivo and induction of apoptosis of hepatocytes in vitro. METHODS: TGF beta was isolated from fresh bovine platelets by acid/ethanol extraction method and purified with ion exchange and gel chromatography. The extracted TGF beta was injected subcutaneously to mice, and its biological activity in vitro was observed 72 hrs post-injection by HE staining. The morphological changes were observed by HE staining and the occurrence of apoptosis was detected by TUNEL method after the human normal hepatic cell line QZG was treated with 8 microg.(-1)TGFbeta for 12 hrs in vitro. RESULTS: The molecular mass 25 ku TGFbeta protein was successfully extracted. It was able to induce localized granulation tissue formation in vitro. TGF beta-treated hepatocytes showed obvious apoptotic morphological changes, including the pyknosis and dense-stained nuclei and cytoplasm, the fragmentary, annular or crescent nuclei, and the "bubbling" cytoplasm. Moreover, its apoptotic rate was significantly higher than that of the control group (P<0.05). CONCLUSION: Biological active TGF beta protein is extracted and purified successfully from bovine platelets, and it is able to induce the apoptosis of hepatocytes.

Animals↗
PubMed2001

[Changes of human recombination bone morphogenetic protein-2 in bone and marrow in tail suspended rats].

OBJECTIVE: To study human recombination bone morphogenetic protein-2 (rhBMP-2) change in bone and marrow under simulated weightlessness. METHOD: 5 SD rats were tail suspended for 14, 28 d with another 5 freely active rats as control. Histological samples were in situ hybridized. RESULT: rhBMP-2 expression of bone and marrow were lower in tail suspended rats than control levels (P < 0.05). rhBMP-2 expression tail suspended rats in 14 day were higher than that in 28 day suspended group (P < 0.05). CONCLUSION: rhBMP-2 levels in rats bone and marrow were lower after tail suspension.

Animals↗
PubMed2001

[The effects of apoptosis during fracture healing].

OBJECTIVE: To elucidate the occurrence and significance of apoptosis during fracture healing of the mandible. METHODS: An experimental model of fracture was established to detect apoptosis in the different stages of healing by TdT-mediated dUTP nick end-labeling (TUNEL) method. RESULTS: Apoptosis was observed in the whole healing process. The apoptotic cells markedly increased in the stage of intramembranous bone formation (5d) and chondrogenesis (11d). CONCLUSION: Apoptosis is involved in fracture healing. The number of cells was controlled and the useless cells were eliminated by means of apoptosis. This process is helpful to the healing of tissues.

English Abstract↗
PubMed2000

Toxic epidermal necrolysis following combination of methotrexate and trimethoprim-sulfamethoxazole.

A 15-year-old boy with T-cell acute lymphoblastic leukemia (ALL) (FAB L1), diagnosed in 1995, received combination chemotherapy consisting of 6 weeks of induction (vincristine, epirubicin, L-asparaginase, prednisolone) and 2 weeks of consolidation (cytosine arabinosides, etoposide). After achieving remission, for further maintenance of remission, he was treated with 14 cycles of intensive chemotherapy consisting of 6-MP, 10 mg/kg orally on the first 4 days, and cyclophosphamide, 1200 mg/m2, vincristine, 1.5 mg/m2, epirubicin, 15 mg/m2, and cytosine arabinoside, 40 mg/m2, intravenously on days 4, 11, 39, and 40, respectively. On day 18 of each cycle, he received intravenous methotrexate (MTX) infusion in a total dose of 150 mg/m2 plus oral leucovorin (30 mg/m2 ) rescue 36 h after starting MTX therapy. In addition, oral trimethoprim-sulfamethoxazole was given regularly to prevent Pneumocystis carinii infection. The patient achieved remission during the first course of treatment, but 8 months later the disease relapsed. He then received four doses of MTX (800 mg intravenously) plus leucovorin rescue in the following 4 months. During the last MTX therapy, small hemorrhagic bullae were found on the lateral side of the right ankle, but subsided after a few days. Due to partial remission of the disease, he was admitted again in January 1999 for high-dose MTX therapy. An initial hemogram on admission revealed hemoglobin 7.2 g/dL, white cell count 15,200/mm3, platelet count 153/mm3, blood creatinine 0.5 mg/dL, and alanine leucine aminotransferase (ALT) 20 U/L. He received 8500 mg of MTX (5000 mg/m2 ) as a continuous intravenous infusion for 24 h. Thirty-six hours after the start of MTX infusion, leucovorin (30 mg, intravenous) rescue was initiated every 6 h for 3 days. Another preventive measure to cover MTX toxicity included aggressive intravenous fluid replacement (4 L/m2 /day) and the addition of 25 meq/L sodium bicarbonate to the intravenous fluid to alkalinize the urine. Concurrent medication included 6-MP (50 mg) once daily and trimethoprim-sulfamethoxazole (120 mg, 600 mg) twice daily every other day. Plasma MTX levels were 52.36 micromol/L 24 h after MTX infusion, 1.87 micromol/L after 48 h, 0.57 micromol/L after 72 h, and 0.41 micromol/L after 96 h. These indicated delayed MTX plasma clearance. The blood creatinine level was mildly elevated from 0.5 mg/dL to 0.7 mg/dL. Thirty-six hours after the administration of MTX, the patient developed an erythematous painful swelling on the right middle finger. The erythema, with subsequent large bulla formation, progressed to all the fingers, toes, palms, and the soles of the feet. Some erythematous to hemorrhagic papules also appeared on the bilateral elbows. Subsequently, diffuse tender erythema with extensive erosions and focal tiny pustules developed on the back, abdomen, proximal extremities, and face (Fig. 1a,b). A positive Nikolsky's sign was also present. A biopsy specimen of the right dorsal hand lesion revealed parakeratosis, detached acanthotic epidermis with scattered necrotic keratinocytes, dyskeratotic cells and nuclear atypia, neutrophilic exocytosis, and many neutrophils in the papillary dermis (Fig. 2). The skin condition deteriorated rapidly. Toxic epidermal necrolysis-like lesions involved 90% of the total body surface on the fifth day after MTX infusion. Mucositis, diarrhea, involuntary tremor, fever, and chills were noted. The patient was then sent to the burn unit for intensive skin care. Ten days after MTX therapy, profound agranulocytosis and thrombocytopenia (white cell count 100/mm3, platelets 14,000/mm3, and hemoglobin 5.6 g/dL) were found. The patient was then started on granulocyte colony stimulation factor (G-CSF, 5 microg/kg/day), but his general condition deteriorated rapidly and he died 6 days later due to septic shock and multiple organ failure.

Adolescent↗
PubMed2000

The TITAN5 gene of Arabidopsis encodes a protein related to the ADP ribosylation factor family of GTP binding proteins.

The titan (ttn) mutants of Arabidopsis exhibit dramatic alterations in mitosis and cell cycle control during seed development. Endosperm development in these mutants is characterized by the formation of giant polyploid nuclei with enlarged nucleoli. Embryo development is accompanied by significant cell enlargement in some mutants (ttn1 and ttn5) but not others (ttn2 and ttn3). We describe here the molecular cloning of TTN5 using a T-DNA-tagged allele. A second allele with a similar phenotype contains a nonsense mutation in the same coding region. The predicted protein is related to ADP ribosylation factors (ARFs), members of the RAS family of small GTP binding proteins that regulate various cellular functions in eukaryotes. TTN5 is most closely related in sequence to the ARL2 class of ARF-like proteins isolated from humans, rats, and mice. Although the cellular functions of ARL proteins remain unclear, the ttn5 phenotype is consistent with the known roles of ARFs in the regulation of intracellular vesicle transport.

ADP-Ribosylation Factors↗
PubMed2000

Generalized granuloma annulare associated with granulomatous mycosis fungoides.

We describe a 68-year-old man with plaque stage mycosis fungoides (MF) for 8 years. He developed tumorous lesions of granulomatous MF (GrMF) and generalized granuloma annulare (GA) after a previously indolent clinical course. Since then, the clinical course was aggressive with involvement of the bone marrow and lymph nodes, and leukemic change occurred. Systemic chemotherapy was given, but the patient died 9 months later due to neutropenic fever and septic shock. GA in malignant lymphoma has been reported most frequently in association with Hodgkin's disease. To the best of our knowledge, GA associated with GrMF has never been reported in the English language literature. The prognostic significance of the association of granulomatous inflammation and malignancy is reviewed.

Aged↗
PubMed2000

[Observation and analyses on voice changes in the normal aged].

OBJECTIVE: To measure the voice samples of the normal aged in order to systemically study the features of the voice changes. METHOD: To collect and analyze 146 voice samples of the normal aged with sonogram. RESULT: The fundamental frequency of the voice of the aged decreases and rises in the male more than 80 years older. The low frequency harmonics are regulation and the intensity is strong in the formant of the aged. The difference reduces in voice between male and female. The harmonics to noise ratio tends downwards and the amplitude perturbation quotient tends upwards along with the growth of age in the aged male. The changes of the above-mentioned parameters are not significant in the aged female. CONCLUSION: The voice changes are normal physiological ones in the normal aged. The changes of the parameters are used to evaluate normal aged voice and abnormal one. The changes show that the function in the aged phonation tends to decline to a certain extent and it must be protected and be trained.

Aged↗
PubMed1999

Epidermal growth factor and angiotensin II regulation of extracellular signal-regulated protein kinase in rat liver epithelial WB cells.

Activation of extracellular signal-regulated protein kinase (ERK) is considered essential for mitogenesis. In the present study, rat liver epithelial WB cells were used to investigate the relative roles of Ca2+, protein kinase C (PKC), and protein tyrosine phosphorylation in mitogenesis and activation of the ERK pathway stimulated by epidermal growth factor (EGF) and angiotensin II (Ang II). The sensitivity of the ERK pathway to Ca2+ was studied by using 1,2-bis (O-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA) to chelate intracellular Ca2+ and a low extracellular Ca2+ concentration to prevent Ca2+ influx. Agonist-induced PKC activation was diminished by inhibition of PKC by GF-109203X (bisindolylmaleimide) or by down-regulation of PKC by long-term treatment of the cells with phorbol myristate acetate (PMA). Our results show that although activation of PKC was critical for mitogenesis induced by Ang II or EGF, the initial activation of ERK by both agonists in these cells was essentially independent of PKC activation and was insensitive to Ca2+ mobilization. This is in contrast to the findings in some cell types that exhibit a marked dependency on mobilization of Ca2+ and/or PKC activation. On the other hand, an obligatory tyrosine phosphorylation step for activation of ERK was indicated by the use of protein tyrosine kinase inhibitors, which profoundly inhibited the activation of ERK by EGF, Ang II, and PMA. Additional experiments indicated that tyrosine phosphorylation by a cytosolic tyrosine kinase may represent a general mechanism for G-protein coupled receptor mediated ERK activation.

Angiotensin II↗
PubMed1999

Tumor necrosis factor-alpha and ceramide induce cell death through different mechanisms in rat mesangial cells.

It has been proposed that ceramide acts as a cellular messenger to mediate tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis. Based on this hypothesis, it was postulated that resistance of some cells to TNF-alpha cytotoxicity was due to an insufficient production of ceramide on stimulation by TNF-alpha. The present study was initiated to investigate whether this was the case in mesangial cells, which normally are insensitive to TNF-alpha-induced apoptosis. Our results indicate that although C2 ceramide was toxic to mesangial cells, the cell death it induced differed both morphologically and biochemically from that induced by TNF-alpha in the presence of cycloheximide (CHX). The most apparent effect of C2 ceramide was to cause cells to swell, followed by disruption of the cell membrane. It is evident that C2 ceramide caused cell death by necrosis, whereas TNF-alpha in the presence of CHX killed the cells by apoptosis. C2 ceramide did not mimic the effects of TNF-alpha on the activation of c-Jun NH2-terminal protein kinase and nuclear factor-kappaB transcription factor. Although mitogen-activated protein kinase [extracellular signal-related kinase (ERK)] was activated by both C2 ceramide and TNF-alpha, such activation appeared to be mediated by different mechanisms as judged from the kinetics of ERK activation. Furthermore, the cleavage of cytosolic phospholipase A2 during cell death induced by C2 ceramide and by TNF-alpha in the presence of CHX showed distinctive patterns. The present study provides evidence that apoptosis and necrosis use distinctive signaling machinery to cause cell death.

Animals↗
PubMed1999

Induction of reparative dentin formation in dogs with combined recombinant human bone morphogenetic protein 2 and fibrin sealant.

OBJECTIVE: To investigate the reparative dentin formation induced by the complex of recombinant human bone morphogenetic protein 2 (rhBMP2) and fibrin sealant (FS) in dogs. METHODS: Freshly exposed pulp of molars, premolars, and canines were treated, respectively, with the complex of rhBMP2-FS, rhBMP2, and Ca(OH)2 paste, which served as control. RESULT: Wound healing of exposed pulp treated with the complex of rhBMP2 and FS seemed better than that with rhBMP2 alone. Dentin bridge formation was observed at 1 week when pulp was treated with the complex and at 4 weeks with rhBMP2 and Ca(OH)2. At 9 weeks after operation, more amount of tubular dentin bridge formation was found in pulp treated with the complex than with rhBMP2. CONCLUSIONS: The results suggest that synergistic effects of fibrin and rhBMP2 existed and that rh-BMP2 with FS carrier can be used as bioactive pulp capping agent. Together, these agents can induce a large amount of dentin and enhance healing of exposed pulp.

Animals↗
PubMed1998

Correlation between sustained c-Jun N-terminal protein kinase activation and apoptosis induced by tumor necrosis factor-alpha in rat mesangial cells.

Rat mesangial cells are normally resistant to tumor necrosis factor-alpha (TNF-alpha)-induced apoptosis. In this report we show that the cells can be made susceptible to the apoptotic effect of TNF-alpha when pretreated with actinomycin D, cycloheximide, or vanadate. c-Jun N-terminal protein kinase (JNK) has been thought to mediate apoptotic processes elicited by some stimuli, but its involvement in TNF-alpha-induced apoptosis has been controversial. JNK activation was investigated under conditions where the mesangial cells were either resistant or susceptible to TNF-alpha-induced apoptosis. TNF-alpha alone stimulated a single transient JNK activity peak. However, when the cells were pretreated with actinomycin D or cycloheximide, TNF-alpha stimulated a second sustained JNK activity peak. When the cells were pretreated with the phosphatase inhibitor vanadate, TNF-alpha-induced JNK activation was greatly prolonged. In all three cases, a sustained JNK activation was associated with the initiation of apoptosis. Our data suggest that a sustained activation of JNK induced by these reagents may be associated with blocking the expression of a phosphatase that inactivates JNK. Further studies reveal that the expression of mitogen-activated protein kinase phosphatase-1 (MKP-1) was induced by TNF-alpha, indicating that MKP-1 may be involved in protecting the cells from apoptosis by preventing a prolonged activation of JNK under normal conditions. Additional studies showed that extracellular signal-regulated protein kinase activation stimulated by TNF-alpha was unlikely to contribute to the resistance of mesangial cells to TNF-alpha cytotoxicity.

Animals↗
PubMed1998

A novel approach to identify proteins associated with the inhibition of neurite growth.

In the present study, a novel combination of techniques was used to identify the genes that may be involved in the lack of axonal regeneration in the mammalian adult central nervous system (CNS). The key features of this approach are: (1) a functional assay that can be affected by antibody perturbation; (2) increased specificity of the polyclonal antiserum by adsorption; (3) the expression cloning of the genes from a lambdagt11 library; (4) amplification of the insert cDNA by PCR; and (5) the direct cycle sequencing of PCR products. In this culture assay system, neurons were plated directly on sections of the rat CNS. This assay system could be used to demonstrate the lack of neuronal attachment to or neurite extension over myelinated regions of the CNS (white matter). This prohibitive nature of the CNS sections could be masked by a rabbit polyclonal antiserum directed against rat CNS white matter. This data indicates that the anti-white matter antiserum recognizes and neutralizes inhibitory molecules on the surface of the sections. Making the assumption that the prohibitive antigen is associated with the cell membrane, the antiserum was adsorbed against a soluble protein fraction of the adult rat brain. This adsorption significantly increased the specificity of the antiserum as demonstrated by immunoblot methods. The adsorbed antiserum was then used to screen the cDNA library of the adult rat brain. The present report describes this novel combination of techniques allowing one to go from a functional tissue culture assay system to defining the molecular basis for the cellular interactions.

Adsorption↗
PubMed1998

Detection of potato virus Y P1 protein in infected cells and analysis of its cleavage site.

The P1 protein is liberated from the N-terminal region of the potyviral polyprotein by cleavage depending on its own autoproteolytic activity. Existence of the 32-kDa P1 protein in tobacco plants infected with potato virus Y ordinary strain (PVY-O) was detected by an antiserum against a recombinant PVY-O P1 protein. In vivo analysis using tobacco protoplasts confirmed that the Phe284-Ser285 was the cleavage site separating the P1 protein from the PVY-O polyprotein. Phe284 was indispensable for proteolysis and Ser285 was needed for optimal cleavage susceptibility.

Amino Acid Sequence↗
PubMed1998

SEM study on microvascular changes following implantation of bone morphogenetic protein combined with hydroxyapatite into experimental bone defects.

BMP-HAP complex was implanted into a bone defect in the femur of Wistar-strain rats, and animals were allowed to heal for one to 8 weeks prior to sacrifice. Similar bone defects without BMP-HAP complex served as controls. Osseous healing and microvascular changes, as revealed by plastic microcorrosion castings, were subsequently examined under a scanning electron microscope. One week after implantation, sproutings and congregate sinusoidal capillary plexuses and primary bone trabeculae (woven bone) were observed around and between the BMP-HAP complexes. Control specimens revealed a fine and immature sinusoidal capillary plexus arising from sproutings and elongations of pre-existing blood vessels, but bone formation was not observed. At two weeks, newly-formed trabeculae were observed around and on the surface of the HAPs, and a network of thick, newly-formed vessels was observed in intervening space. At three weeks, networks of newly-formed vessels were observed on the surface of the HAPs, and surrounding newly-formed trabeculae had become thickened. At four weeks, bone defects were filled, and HAP was completely embedded in new bone. At eight weeks, the HAP was fused with new bone, and the boundary between the HAP and new bone was unclear. In comparison with comparable surgically created control bone defects without implanted BMP-HAP complex, BMP apparently stimulates new vascularization. Further, implanted BMP-HAP apparently stimulates undifferentiated mesenchymal cells to differentiate into angioblasts and osteoblasts, or vice versa (dedifferentiation).

Animals↗
PubMed1997

Sialic acid specificity of myelin-associated glycoprotein binding.

Myelin-associated glycoprotein (MAG), a nervous system cell adhesion molecule, is an I-type lectin that binds to sialylated glycoconjugates, including gangliosides bearing characteristic structural determinants (Yang, L. J.-S., Zeller, C. B., Shaper, N. L., Kiso, M., Hasegawa, A., Shapiro, R. E., and Schnaar, R. L. (1996) Proc. Natl. Acad. Sci. U.S.A. 93, 814-818). Two cell adhesion systems, COS-1 monkey kidney fibroblasts transiently transfected to express MAG and Chinese hamster ovary (CHO) cells stably transfected to express MAG, were used to probe the structural specificity of MAG-ganglioside binding. Both cell types bound to the same gangliosides: GQ1b alpha (IV3NeuAc,III6NeuAc,II3(NeuAc)2Gg4Cer) > GT1b = GD1a > GM3 > GM1, GD1b, and GQ1b (the latter do not support adhesion). Binding was enhanced by pretreatment of MAG-expressing cells with neuraminidase. MAG-expressing Chinese hamster ovary cells bound directly to gangliosides resolved on thin layer chromatograms, allowing detection of MAG binding species in a mixture. The simplest ganglioside ligand for MAG was GM3 bearing N-acetylneuraminic acid, whereas GM3 bearing N-glycolylneuraminic acid did not support adhesion. Chemical modifications of N-acetylneuraminic acid residues (on GD1a) abrogated MAG binding. Mild periodate oxidation of sialic acids to their corresponding seven-carbon (or eight-carbon) sialic acid aldehydes abolished MAG binding, as did further conversion to the corresponding primary alcohols. Eliminating the anionic charge by ethyl esterification, amidation, or reduction also abolished MAG-mediated cell adhesion. These data demonstrate that MAG-ganglioside binding is highly specific and defines key carbohydrate structural determinants for MAG-mediated cell adhesion to gangliosides.

Animals↗