Biomedical subjects
C T Lin
Publications and source records attributed to C T Lin.
Monoclonal antibodies to a brain dopamine binding protein: production, specificity, and immunohistochemistry.
A dopamine binding protein (DABP) has been purified from the rat brain synaptic membrane to homogeneity by affinity chromatography and gel electrophoresis. The monoclonal antibodies against the DABP were produced by the mouse-mouse hybridoma technique and characterized for their specificity to dopamine receptors by displacement of dopamine receptor binding. These monoclonal antibodies have been used to localize DABP in rat brain by immunohistochemistry. A specific linear structure of reaction product was seen in both caudate nucleus and cerebral cortex. This finding suggests that the DABP is present in the cerebral cortex and caudate nucleus as a membranous component of the neurons or their processes.
Taurine system in the normal and ischemic rat hippocampus.
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The solid-state decarboxylation of the diammonium salt of moxalactam.
This paper reports studies of the solid-state chemistry of the diammonium salt of moxalactam. The methods employed include X-ray crystallography, molecular mechanics calculations, thermogravimetric analysis, and high-pressure liquid chromatography. The crystal structure shows that the malonic acid amide functionality in crystals of the diammonium salt is not planar. If the common decarboxylation mechanism is operating, then considerable rotation would be required for this functionality to attain coplanarity. Simultaneous HPLC and thermogravimetric analysis studies indicate that the decarboxylation of the diammonium salt of moxalactam is preceded by desolvation. Molecular mechanics calculations indicate that the barrier to rotation of the malonic acid amide functionality is relatively small in the dehydrated crystals, perhaps explaining the facile decarboxylation of this antibiotic. Alternatively, the amorphous desolvated crystals may allow enough molecular freedom for the malonic acid amide functionality to attain coplanarity and decarboxylate.
Brain L-glutamate decarboxylase: purification and subunit structure.
Glutamate decarboxylase (GDCase; L-glutamate-1-carboxy-lyase, EC 4.1.1.15) was purified from whole rat brain approximately equal to 1300-fold to apparent homogeneity with a specific activity of 2.4 units per mg of protein by a combination of column chromatographies on DEAE-cellulose, hydroxylapatite, and gel filtration, and preparative nondenaturing polyacrylamide gel electrophoresis. The purified preparation contained a single protein band that comigrated with GDCase activity in three diverse analyses: nondenaturing regular (5%) and gradient (3.6-25%) polyacrylamide gel electrophoresis and isoelectric focusing at pH 4-7. The native molecular mass was calculated to be 120 +/- 10 kDa from gradient polyacrylamide gel electrophoresis and 110 +/- 10 kDa from gel filtration. Under the treatment with NaDodSO4 and 2-mercaptoethanol, GDCase dissociated into two subunits of 40 +/- 2 and 80 +/- 4 kDa, as estimated from NaDodSO4 gel electrophoresis. However, only a 40-kDa subunit was detected when GDCase was treated with 4 M urea plus NaDodSO4 and 2-mercaptoethanol, suggesting that the 80-kDa subunit is the dimer of the 40-kDa subunit. In immunoblotting, polyclonal antibodies against GDCase reacted with both 40- and 80-kDa subunits, while monoclonal antibody reacted with only 80-kDa subunits. The isoelectric point of the native enzyme was 5.4. The Km for glutamate was 1.59 X 10(-3) M. In addition to L-glutamate, cysteine sulfinic acid was also decarboxylated at approximately equal to 10% of the rate of glutamate. The pH optimum was fairly broad, with a maximum at approximately equal to 7.3. The enzyme was strongly inhibited by carbonyl-trapping agents, sulfhydryl reagents, thiol compounds, and beta-methylene-DL-aspartate.
Treatment of a patient with Papillon-Lefèvre syndrome. A case report.
A vital root submersion procedure was performed in a rare case of Papillon-Lefèvre syndrome (PLS). Although the etiology and pathogenesis are obscure, possible host genetic defects and known or unknown periodontal pathogens may play important roles in the progression of PLS. Since this patient did not respond to conventional periodontal therapy, a submersion procedure was employed to maintain the alveolar ridge and resulted in controlling the progression of the patient's severe periodontitis. The following is a report of the treatment and the rationale for therapy.
[Intussusception in adults--report of nine cases].
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[Clinical study of establishing the vertical dimension of occlusion with various methods].
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Taurine in hippocampus: localization and postsynaptic action.
Both immunocytochemical and electrophysiological methods have been employed to determine whether the localization of the taurine synthetic enzyme, cysteine sulfinic acid decarboxylase, (CSAD) and the postsynaptic action of taurine in the CA1 region of rat hippocampus are consistent with the hypothesis that taurine may be used as a neurotransmitter by some hippocampal neurons. At the light microscopic level, CSAD-immunoreactivity (CSAD-IR) was found in the pyramidal basket cells, and around pyramidal cells in stratum pyramidale and stratum radiatum. At the electron microscopic level, CSAD-IR was seen most often in the soma and the dendrites and was rather infrequent in the axon or the nerve terminals. Electrophysiological observations on the in vitro hippocampal slice demonstrated that pyramidal neurons respond to artificially applied taurine with inhibition that depended in large part upon an increased chloride conductance. Although electrophysiological observations are consistent with a neurotransmitter role for taurine, results from immunocytochemical studies suggest a minor role for taurine as a neurotransmitter. In fact, immunocytochemical observations suggested that taurine may be used as a neurotransmitter only by a small number of pyramidal basket interneurons, the vast majority of CSAD-positive neurons may use taurine for other functions.
Production and characterization of polyclonal and monoclonal antibodies to rat brain L-glutamate decarboxylase.
Specific monoclonal and polyclonal antibodies to rat brain glutamate decarboxylase (GAD) were produced and characterized. Polyclonal antibodies against GAD were raised in rabbits by injecting a total of 70-210 micrograms of purified GAD i.m. The specificity of anti-GAD serum was established from a variety of tests including Ouchterlony immunodiffusion, immunoelectrophoresis, immunoprecipitation, dot immunoassay, ELISA tests and Western immunoblottings. In immunodiffusion and immunoelectrophoresis tests using partially purified GAD preparations and anti-GAD serum a single, sharp precipitin line corresponding to GAD activity was obtained. Quantitative immunoprecipitation of GAD activity was achieved using anti-GAD IgG and Staphylococcus aureus. Specificity of the antiserum was further indicated from a dot immunoassay and ELISA tests in which the intensity of the reaction product was proportional to the amount of GAD protein present. In the Western immunoblotting experiments using partially purified GAD preparations only two protein bands corresponding to the position of the two subunits of GAD were stained by anti-GAD IgG, further supporting the specificity of polyclonal antibodies against GAD. In addition to polyclonal antibodies, several specific GAD-antibodies-producing clones were also obtained by the hybridoma technique. The specificity of monoclonal antibodies against GAD were established from the following criteria: positive on ELISA test using homogeneous GAD as antigen; formation of GAD--anti-GAD IgG complex as indicated from gel filtration chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis; and specific recognition of GAD subunit in a partially purified GAD preparation in Western immunoblotting test. Monoclonal antibodies were further characterized by immunohistochemical localization of known GABAergic neurons and their processes in the cerebellum and retina.
Morphological localization of apolipoproteins and their mRNA by immunocytochemistry and in situ nucleic acid hybridization.
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Immunoultrastructural demonstration of prostatic acid phosphatase isoenzyme 2 in prostatic carcinoma.
Human prostatic acid phosphatase isoenzyme 2 (HPAcP-2) was isolated from semen. This purified enzyme was immunized to rabbit to produce polyclonal antibodies. The specificity of the antibodies was tested by Western blot transfer method. Rabbit IgG-peroxidase conjugate was prepared from the antiserum and used to localize HPAcP-2 in prostatic carcinoma. It was found that in the tumor glandular acinus the normal basal cells were replaced by tumor cells containing reaction product. In the tumor cells, the reaction product was seen in the cisternae of rough endoplasmic reticulum (ER) and Golgi apparatus. The secretory vesicles which contained reaction product-stained granules and some amorphous material were seen to fuse with the apical plasma membrane and discharged their content into the glandular lumen. On the other hand, some secretory vesicles in the tumor cells facing to the basement membrane also discharged their similar content into the extracellular spaces. Reaction product-stained granules were found in the interstitial spaces surrounding the tumor cells. These findings suggest that HPAcP-2 is synthesized on the bound ribosomes and discharged into the cisternae of rough ER. The molecules are transported to the Golgi cisternae. After concentration and packaging, HPAcP-2 molecules are then transferred to the secretory vesicles, and discharged into the glandular lumen and to the extracellular spaces. The isoenzyme released in the extracellular space may reach the blood stream through the interstitial spaces or the lymphatic system, resulting in the elevation of serum HPAcPase level in some prostatic cancer patients.
Immunoreactive apolipoprotein E is a widely distributed cellular protein. Immunohistochemical localization of apolipoprotein E in baboon tissues.
Apolipoprotein (apo)E is an important protein determinant in cholesterol homeostasis in man. The protein is synthesized by the liver as well as by a number of extrahepatic tissues. In the present study, immunohistochemical techniques were used to identify apoE in specific cells in various baboon organs. In the 11 tissues studied, the following cell types have been found to harbor apoE immunoreactivity: cerebral astrocytes; thyroid follicular cells; alveolar type II pneumocytes; hepatocytes, and Kupffer cells; adrenocortical cells in zona fasciculata and zona reticularis; adrenal medullary cells; some renal tubular epithelia; some pancreatic islet cells; histiocytic macrophages in lymph nodes and the spleen; some gastric mucosal epithelia; and ovarian oocytes. These observations indicate the wide distribution of apoE in many organs and suggest that the protein might perform other important functions such as regulation of local hormonal homeostasis in addition to its role in cholesterol metabolism.
Calmodulin: localization in plant tissues.
Calmodulin was purified from bovine brain by preparative SDS-polyacrylamide gel electrophoresis. The denatured, purified calmodulin was used to immunize rabbits to produce antiserum. This antiserum was used to study the distribution of calmodulin in plant tissues by indirect immunohistochemistry. The root tips from corn seeds, oat seeds, peanuts, spaghetti squash seeds, and the terminal buds of spinach were investigated. A method for plant tissue sectioning and inhibition of endogenous peroxide activity was developed. In the corn root section, reaction product from anti-calmodulin was found mainly in the root cap cells. Lesser but significant amounts of calmodulin were localized in metaxylem elements, in some stele cells surrounding metaxylem elements, in apical initials, and in the cortical cells. Similar findings were also observed in other root tips from oat seeds, peanuts, and spaghetti squash seeds. In the terminal buds of the spinach, calmodulin-stained cells were highly concentrated in the apical meristem and leaf primordium. These findings suggest that the high concentration of calmodulin in the root cap may be important in relation to gravitropism and growth development.
Estrogen induction of very low density apolipoprotein II synthesis, a major avian liver yolk protein, involves the recruitment of hepatocytes.
The mechanism of steroid hormone action was studied in the cockerel liver. Very low density apolipoprotein II (apo-VLDL-II), a yolk protein, is a low molecular weight apolipoprotein that is inducible by estrogen. The intracellular apo-VLDL-II messenger RNA (mRNA) concentration under various hormonal conditions was examined by a dot-blot assay. The concentration was very low in untreated cockerels (approximately 0.5 molecule per cell). It increased to 8,000 molecules per cell within 24 h of estrogen treatment and reached a maximum level of approximately 70,000 molecules per cell after 14 daily doses of estrogen. The distribution of hepatocytes harboring apo-VLDL-II and its mRNA was studied by immunohistochemistry and by in situ nucleic acid hybridization to cloned [3H]apo-VLDL-II complementary DNA (cDNA). The number of cells containing the immunoreactive protein and the hybridizable mRNA increased from extremely low (0.3% and 0.27%, respectively) to substantial (11% and 10%, respectively) at 24 h after estrogen treatment and to extremely high (94% and 92%, respectively) in maximally treated animals. Our studies indicate that, in addition to enhanced transcription and stabilization of mRNA, the recruitment of liver cells previously not engaged in the synthesis of apo-VLDL-II is an important mechanism by which the hormone induces the hepatic production of this protein. The phenomenon of recruitment and the heterogeneity of the functional capacity of individual hepatocytes to respond to estrogen may be important to our understanding of estrogen action in the liver.
Isolation, purification, and immunochemical studies of benzodiazepine receptor(s) and its ligand(s) for mammalian brain.
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[Computed tomography and cerebral angiography in brain metastasis of choriocarcinoma].
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Alteration of acid phosphatase isoenzyme in a human prostatic cancer cell line.
The acid phosphatase (AcP) isoenzyme in a human prostatic cancer cell line was compared to that of prostatic tissue extract by electrophoresis. The major isoenzyme by prostatic tissue extract is the AcP isoenzyme 2, while only AcP isoenzyme 4 (AcP-4) was observed in the human prostatic cancer cell line. A monoclonal antibody specific to AcP-4 was used to investigate the ultrastructural distribution of AcP-4 in a prostatic cancer cell line. The peroxidase staining pattern indicates that AcP-4 is synthesized on bound ribosomes, discharged into the cisternae of rough endoplasmic reticulum, transported to the cisternae of Golgi apparatus for concentration and packaging, and transferred to the secretory vesicles for exocytosis. It is well known that synthesis and secretion of AcP-2 are the major characteristics of the highly differentiated prostatic epithelial cells. The present data demonstrate the loss of this specific function in the prostatic cancer cell line. Instead of AcP-2, the dedifferentiated cancer cell line synthesizes and secretes AcP-4, which is a common AcP isoenzyme of many nonprostatic tissues.