102 results on '"Chu LL"'
Search Results
52. The inhibitory effects of a new cobalt-based polyoxometalate on the growth of human cancer cells.
- Author
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Wang L, Yu K, Zhou BB, Su ZH, Gao S, Chu LL, and Liu JR
- Subjects
- Antineoplastic Agents chemistry, Apoptosis drug effects, Caspase 3 metabolism, Cell Line, Tumor, Cell Proliferation drug effects, Cobalt chemistry, Comet Assay, DNA Damage, Humans, Tungsten Compounds chemistry, Antineoplastic Agents pharmacology, Cobalt pharmacology, Tungsten Compounds pharmacology
- Abstract
A new cobalt-based polyoxometalate, (Himi)2[Bi(2)W2(0)O(66)(OH)(4)Co2(H2O)(6)Na(4) (H2O)14] · 17H2O (imi = iminazole) (BWCN) has been synthesized and structurally characterized. The inhibitory activities against selected human cancer lines were also determined in this study. The cell viability and chemoresistance of BWCN on human colon carcinoma HT-29 cells were assessed by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazoliumbromide), cell morphology changes, a comet assay and western blot analysis. The typical morphologic changes of apoptosis and DNA damage indicated that BWCN could have a distinct proliferation inhibitory effect on cancer cells. BWCN as a chemotherapeutic agent also induced apoptosis on HT-29 cells and showed a significant expression of cleaved-caspase-3. These results suggested that the active site of BWCN is the polymeric anion based on the basic tectonic block {BiW(9)}, and the possible mechanism is related to the interference of DNA synthesis in cancer cells.
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- 2014
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- View/download PDF
53. The health promotion lifestyle of metabolic syndrome individuals with a diet and exercise programme.
- Author
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Lin YH and Chu LL
- Subjects
- Adult, Aged, Aged, 80 and over, Female, Humans, Male, Middle Aged, Diet, Exercise, Health Promotion, Life Style, Metabolic Syndrome physiopathology
- Abstract
The purpose of this study was to explore a health promotion lifestyle (HPL) with a diet and exercise programme (DEP) in metabolic syndrome adults. The study consisted of 207 individuals who followed a DEP and 185 who did not. The subjects were rural community adults. Their HPL was evaluated using the Chinese version of the Health Promotion Lifestyle Profile Short Form (HPLP-S). The average HPLP-S score was significantly higher in the DEP group (3.28 ± 0.36) than in the group without the DEP (2.05 ± 0.65). Stepwise regression analysis revealed that group, gender, smoking, alcohol use, marital status, religion and chronic disease were predictors of an HPL and accounted for 67.0% of the variance in the HPLP-S score. This study demonstrates that a DEP has positive effects on a health promotion lifestyle. The community-based DEP targeting health promotion behaviours should be presented as a strategy for metabolic syndrome in adults., (© 2013 Wiley Publishing Asia Pty Ltd.)
- Published
- 2014
- Full Text
- View/download PDF
54. Human cancer cells retain modest levels of enzymatically active matriptase only in extracellular milieu following induction of zymogen activation.
- Author
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Chu LL, Xu Y, Yang JR, Hu YA, Chang HH, Lai HY, Tseng CC, Wang HY, Johnson MD, Wang JK, and Lin CY
- Subjects
- Cell Line, Tumor, Enzyme Activation, Extracellular Space enzymology, Extracellular Space metabolism, Humans, Neoplasms enzymology, Enzyme Precursors metabolism, Neoplasms pathology, Serine Endopeptidases metabolism
- Abstract
The type 2 transmembrane serine protease matriptase is broadly expressed in human carcinomas and hematological cancers. The proteolytic activity of matriptase is a potential target of drugs and imaging probes. We assessed the fate of active matriptase following the induction of matriptase zymogen activation. Exposing eight human carcinoma cells to pH 6.0 buffer induced robust matriptase zymogen activation followed by rapid inhibition of the nascent active matriptase by hepatocyte growth factor activator inhibitor (HAI)-1. Consequently, no enzymatically active matriptase was detected in these cells. Some active matriptase is, however, rapidly shed to the extracellular milieu by these carcinoma cells. The lack of cell-associated active matriptase and the shedding of active matriptase were also observed in two hematological cancer lines. Matriptase shedding is correlated closely with the induction of matriptase activation, suggesting that matriptase activation and shedding are kinetically coupled. The coupling allows a proportion of active matriptase to survive HAI-1 inhibition by rapid shedding from cell surface. Our study suggests that cellular free, active matriptase is scarce and might not be an effective target for in vivo imaging and drug development.
- Published
- 2014
- Full Text
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55. CT of acute appendicitis: can diagnostic accuracy serve as a practical performance metric for readers specialized in abdominal imaging?
- Author
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Chu LL, Webb EM, Stengel JW, Yeh BM, Lu Y, and Coakley FV
- Subjects
- Acute Disease, Dimensional Measurement Accuracy, Female, Humans, Male, Middle Aged, Quality Assurance, Health Care, Reproducibility of Results, Retrospective Studies, Sensitivity and Specificity, Tomography, X-Ray Computed methods, Appendicitis diagnostic imaging
- Abstract
Purpose: To investigate diagnostic accuracy for acute appendicitis at computed tomography (CT) as a performance metric for radiologists specialized in abdominal imaging., Materials and Methods: We retrospectively identified six attending abdominal imagers who each independently interpreted over 100 CT studies for suspected acute appendicitis., Results: The mean number of studies per reader was 311 (range, 129-386). Mean reader diagnostic accuracy was 95.0% (range, 91.4-97.1%). Only one had a diagnostic accuracy (91.4%) that was significantly lower than all others., Conclusion: Diagnostic accuracy for acute appendicitis at CT may be an impractical performance metric for radiologists specialized in abdominal imaging., (© 2014.)
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- 2014
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56. Novel antitumor agent, trilacunary Keggin-type tungstobismuthate, inhibits proliferation and induces apoptosis in human gastric cancer SGC-7901 cells.
- Author
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Wang L, Zhou BB, Yu K, Su ZH, Gao S, Chu LL, Liu JR, and Yang GY
- Subjects
- Adenocarcinoma pathology, Antineoplastic Agents chemistry, Cell Line, Tumor, Cell Proliferation drug effects, DNA Fragmentation drug effects, Humans, Models, Molecular, NF-kappa B analysis, Photoelectron Spectroscopy, Proto-Oncogene Proteins c-bcl-2 analysis, Stomach pathology, Stomach Neoplasms pathology, Tungsten Compounds chemistry, Adenocarcinoma drug therapy, Antineoplastic Agents pharmacology, Apoptosis drug effects, Stomach drug effects, Stomach Neoplasms drug therapy, Tungsten Compounds pharmacology
- Abstract
A new one-dimensional chain-like compound of tungstobismuthate, [(W(OH)2)2 (Mn(H2O)3)2(Na3(H2O)14)(BiW9O33)2](Himi)2·16H2O (1) (imi = iminazole), has been synthesized in aqueous solution. The structure of 1 was identified by elemental analysis, IR, thermogravimetry (TG), X-ray photoelectron spectroscopy (XPS), (183)W-NMR, and single crystal X-ray diffraction. To investigate the inhibitory effect of 1 on human gastric adenocarcinoma SGC-7901 cells, cell proliferation and apoptosis initiation were examined by MTT assay (MTT = 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazoliumbromide), flow cytometry, nuclear staining, transmission electron microscopy, single cell gel electrophoresis, DNA fragmentation, and Western blotting. The results showed that 1 inhibited cell proliferation and induced apoptosis in SGC-7901 cells in dose-dependent manner. In addition, 1 also decreased the expression of bcl-2 protein and nuclear factor-κB p65 protein in SGC-7901 cells. And expression of bcl-2 protein exhibits a decreasing trend with increase of concentration of 1. Thus, 1 possessed a potential antitumor activity in SGC-7901 cells. This suggests that polyoxotungstates will provide a promising and novel antitumor agent in prevention and treatment of gastric adenocarcinoma.
- Published
- 2013
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57. Lifestyle intervention in non-alcoholic fatty liver disease in Chengyang District, Qingdao, China.
- Author
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Sun WH, Song MQ, Jiang CQ, Xin YN, Ma JL, Liu YX, Ma L, Lin ZH, Li CY, Liu L, Zhang M, Chu LL, Jiang XJ, Wan Q, Zhou L, Ren R, and Meng LF
- Abstract
Aim: To evaluate the effect of a 6 and 12 mo lifestyle modification intervention in nonalcoholic fatty liver diseases (NAFLD) in Chengyang District of Qingdao., Methods: Participants with NAFLD who had resided in Chengyang District for more than 5 years were enrolled in this study. After the 6 and 12 mo lifestyle modification intervention based on physical activity, nutrition and behavior therapy, parameters such as body weight, body mass index (BMI), waist circumference, serum alanine aminotransferase (ALT), aspartate aminotransferase values, serum cholesterol, triglycerides, fasting glucose, fasting insulin and visceral fat area (VFA), the liver-spleen ratio and the homeostasis model assessment of insulin resistance (HOMA-IR) were evaluated and compared between participants with and without the intervention., Results: Seven hundred and twenty-four participants were assigned to the lifestyle intervention group (LS) and 363 participants were assigned to the control group (CON). After the intervention, body weights in the LS group were significantly decreased compared to those in the CON group at 6 mo (11.59% ± 4.7% vs 0.4% ± 0.2%, P = 0.001) and at 12 mo (12.73% ± 5.6% vs 0.9% ± 0.3%, P = 0.001). Compared with the CON group, BMI was more decreased in the LS group after 6 and 12 mo (P = 0.043 and P = 0.032). Waist circumference was more reduced in the LS group than in CON (P = 0.031 and P = 0.017). After the 6 and 12 mo intervention, ALT decreased significantly in the LS group (P = 0.003 and P = 0.002). After 6 and 12 mo, the metabolic syndrome rate had decreased more in the LS group compared with the CON group (P = 0.026 and P = 0.017). After 12 mo, the HOMA-IR score decreased more obviously in the LS group (P = 0.041); this result also appeared in the VFA after 12 mo in the LS group (P = 0.035)., Conclusion: Lifestyle intervention was effective in improving NAFLD in both 6 and 12 mo interventions. This intervention offered a practical approach for treating a large number of NAFLD patients in the Chengyang District of Qingdao.
- Published
- 2012
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58. Colorectal cancer screening in women: an underutilized lifesaver.
- Author
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Chu LL, Weinstein S, and Yee J
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- Adenocarcinoma diagnosis, Adenocarcinoma prevention & control, Adenocarcinoma secondary, Adenoma diagnosis, Adenoma prevention & control, Advisory Committees, Aged, Canada epidemiology, Colonic Polyps diagnosis, Colonic Polyps prevention & control, Colonography, Computed Tomographic economics, Colorectal Neoplasms epidemiology, Early Detection of Cancer economics, Female, Health Knowledge, Attitudes, Practice, Humans, Lipoma diagnosis, Lipoma epidemiology, Lipoma prevention & control, Male, Middle Aged, Patient Compliance statistics & numerical data, Reimbursement Mechanisms, United States epidemiology, Colonography, Computed Tomographic methods, Colorectal Neoplasms diagnosis, Colorectal Neoplasms prevention & control, Early Detection of Cancer statistics & numerical data, Women's Health
- Abstract
Objective: Colorectal cancer (CRC) represents the third most common cancer diagnosed and a major cause of cancer-related deaths in women. Despite strong evidence that early screening decreases colorectal cancer incidence and mortality rates, colorectal cancer screening rates in women still lag significantly behind screening rates for breast and cervical cancers. Additionally, women have been found to be less likely than men to undergo CRC screening. This is despite the fact that the overall lifetime risk for the development of colorectal carcinoma is similar in both sexes. Barriers to screening have been found to be different for women compared with men. Screening adherence in women also appears to be associated with various social and demographic factors., Conclusion: CT colonography (CTC) is an accurate, minimally invasive, and well-tolerated examination that is newly endorsed by the American Cancer Society, U.S. Multisociety Task Force, and the American College of Radiology. Improved screening compliance may occur in women with further dissemination of CTC.
- Published
- 2011
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59. In vivo validation of PAX2 as a target for renal cancer therapy.
- Author
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Hueber PA, Iglesias D, Chu LL, Eccles M, and Goodyer P
- Subjects
- Animals, Apoptosis drug effects, Base Sequence, Carcinoma, Renal Cell metabolism, Carcinoma, Renal Cell pathology, Caspase 3 metabolism, Cell Line, Tumor, Drug Resistance, Neoplasm, Enzyme Activation, Gene Silencing, Humans, Kidney Neoplasms metabolism, Kidney Neoplasms pathology, Mice, Mice, Nude, Molecular Sequence Data, Neoplasm Transplantation, PAX2 Transcription Factor genetics, RNA, Small Interfering genetics, Transplantation, Heterologous, Antineoplastic Agents therapeutic use, Carcinoma, Renal Cell therapy, Cisplatin therapeutic use, Kidney Neoplasms therapy, PAX2 Transcription Factor metabolism
- Abstract
PAX genes are frequently overexpressed in human cancer tissue and appear to contribute to the tumor phenotype, suggesting that they may be potential targets for cancer therapy. In particular, aberrant PAX2 expression has been reported in a high proportion of primary tumors, including the majority of renal cell carcinomas (RCC). We recently demonstrated that PAX2 suppresses cisplatin-induced apoptosis in cultured RCC cells. We hypothesized that silencing of PAX2 expression might partially overcome the notorious resistance of renal cell carcinomas to chemotherapy in vivo. In this report, we show that a PAX2 shRNA successfully knocks down PAX2 mRNA and protein levels in an RCC cell line (ACHN). ACHN cells stably transfected with shRNAs targeted against the PAX2 homeodomain are 3-6-fold more susceptible to cisplatin-induced caspase-3 activation than control ACHN cells line. Furthermore, growth of subcutaneous ACHN/shPAX2 xenografts in nude mice is significantly more responsive to cisplatin therapy than control ACHN cell tumors. Our observations validate PAX2 as a potential therapeutic gene target in renal cancer and suggest that adjunctive PAX2 knockdown may enhance the efficacy of other chemotherapeutic agents.
- Published
- 2008
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60. Role of chloride channels in the regulation of corpus cavernosum tone: a potential therapeutic target for erectile dysfunction.
- Author
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Chu LL and Adaikan PG
- Subjects
- Animals, Chloride Channels metabolism, Dose-Response Relationship, Drug, Male, Muscle Contraction physiology, Muscle Relaxation physiology, Muscle, Smooth physiology, Penile Erection drug effects, Rabbits, Anthracenes pharmacology, Chloride Channels antagonists & inhibitors, Muscle Contraction drug effects, Muscle Relaxation drug effects, Muscle, Smooth drug effects, Niflumic Acid pharmacology
- Abstract
Introduction: Recent electrophysiological recordings have identified the existence of outward, excitatory chloride (Cl(-)) currents in rat, rabbit, and human corpus cavernosum (CC) muscle cells., Aim: The aim of this study was to investigate the physiological role of Cl(-) currents in the maintenance of cavernosal muscle tone in isolated rabbit CC tissues., Methods: CC strips (1 x 1 x 5 mm) were suspended in tissue bath chambers for isometric tension experiments. Spontaneous cavernosal tone and contractions elicited by field stimulation or administration of established smooth muscle constrictors were examined in the presence of chloride channel (ClC) blockers, niflumic acid (NFA), and anthracene-9-carboxylic acid (A9C)., Main Outcome Measure: Both spontaneous myogenic activity and contractile responses to field stimulation, norepinephrine, histamine, and endothelin-1 were reduced by ClC blockers. Results. In CC strips exhibiting intrinsic myogenic tone, NFA (30 and 100 microM) and A9C (1 mM) caused a relaxation of the tone. In addition, spontaneous contractile activity in CC was abolished in the presence of either ClC blocker. In CC strips precontracted with norepinephrine, histamine, and endothelin-1, both ClC blockers significantly reversed the tone. The ability of NFA and A9C to reverse norepinephrine-induced tone was unaffected by N(omega)-nitro-L-arginine, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, and cis-N-[2-phenylcyclopentyl]-azacyclotridec-1-en-2-amine. In addition, neither indomethacin nor tetraethylammonium affected the relaxant response of NFA. NFA and A9C inhibited the neurogenic and norepinephrine-induced contractions in a concentration-dependent manner. While NFA exerted persistent inhibition on neurogenic contraction, inhibition of neurogenic contractions by A9C was readily reversible. On K(+)-depolarized CC, NFA induced a concentration-related relaxation, whereas A9C was inert, suggesting an additional mechanism of NFA on voltage-gated calcium channels., Conclusions: These results underline the importance of Cl(-) currents as a mechanism in the maintenance of cavernosal tone produced by adrenergic and various endogenous constrictors. Thus, the modulation of Cl(-) current, as an attractive and effective approach to regulate penile erection, and specific ClC blockers, as potential erectogenic agents, merits further research.
- Published
- 2008
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61. [Effect of induced occlusal disorders and removed occlusal disorders on the expression of bone morphogenetic protein-2 of condylar cartilage in rats].
- Author
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Li XF, Wang MQ, Chu LL, and Yu SB
- Subjects
- Animals, Cartilage, Cartilage, Articular, Dental Occlusion, Female, Rats, Rats, Sprague-Dawley, Temporomandibular Joint, Bone Morphogenetic Protein 2, Mandibular Condyle
- Abstract
Objective: This article was to study the effect of induced occlusal disorders and removed occlusal disorders on the expression of bone morphogenetic protein-2 (BMP-2) of condylar cartilage., Methods: Young and adult female Sprague-Dawley rats were divided respectively into induced occlusal disorders group, removed occlusal disorders group and control group, 3 rats every group. For induced occlusal disorders rats, the elastic rubbers were inserted between the first and second molar in the left upper side and right lower side to form the disordered occlusion. They were killed under anaesthesia 8 weeks after the treatment. For removed occlusal disorders rats, the first molars that caused disordered occlusion were extracted 6 weeks after forming disordered occlusion. 2 weeks later, they were killed under anaesthesia. For normal rats, they were killed at the same time with experimental rats. Hibateral temporomandibular joints of each rat were removed and stained with HE and monoclone antibody of BMP-2. The thickness of condylar cartilage was measured. The expression of BMP-2 in condylar cartilage was detected by half-quantity immunohistochemical analysis., Results: For adult induced occlusal disorders group, the thickness of cartilage in intermediate part of condyle decreased. However, it increased in the posterior part. After removing occlusal disorders, the thickness of posterior condylar cartilage returned to normal level. But it was still thinner than control group in the intermediate part. The expression of BMP-2 in anterior, intermediate, posterior part of condylar cartilage of young induced occlusal disorders group was higher than that of young removed occlusal disorders group expression of BMP-2 showed induced occlusal disorders group was higher than removed occlusal disorders group, which was higher than control group. and control group. No difference of the expression of BMP-2 was found in young removed occlusal disorders group and control group. For the expression of BMP-2 in intermediate part of condylar cartilage, both adult induced and removed occlusal disorders groups were higher than adult control group. For the posterior part of adult condyle cartilage, the expression of BMP-2 showed induced occlusal disorders group was higher than removed occlusal disorders group, which was higher than control group., Conclusion: Induced occlusal disorders can lead higher expression of BMP-2 in condylar cartilage of young and adult rats. Adaptability of condylar cartilage of adult rats is weaker than young rats, especially the intermediate part.
- Published
- 2008
62. Patient-specific time to peak abdominal organ enhancement varies with time to peak aortic enhancement at MR imaging.
- Author
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Chu LL, Joe BN, Westphalen AC, Webb EM, Coakley FV, and Yeh BM
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- Adolescent, Adult, Aged, Aged, 80 and over, Child, Female, Humans, Male, Middle Aged, Retrospective Studies, Time Factors, Aorta, Abdominal pathology, Jejunum pathology, Liver pathology, Magnetic Resonance Imaging, Pancreas pathology
- Abstract
Purpose: To retrospectively evaluate the relationship between the times to peak enhancement of the liver, pancreas, and jejunum with respect to the time to peak aortic enhancement at magnetic resonance (MR) imaging., Materials and Methods: The committee on human research approved this study and waived written informed consent. This study was HIPAA compliant. The study retrospectively identified 141 patients (63 men, 78 women; mean age, 57 years) who underwent abdominal MR imaging by using a test bolus that was monitored approximately every second for 2 minutes with a spoiled gradient-echo T1 transverse section through the upper abdomen. The times to peak enhancement of the aorta, liver, pancreas, and jejunum were recorded and correlated with the time to peak aortic enhancement, age, and sex by means of univariate and multivariate linear regression analyses., Results: The mean time to peak aortic enhancement was 21.1 seconds (range, 8.7-41.8 seconds). The times to peak enhancement of the liver, pancreas, and jejunum were positively and linearly correlated with the time to peak aortic enhancement (r = 0.69, 0.86, and 0.80, respectively, all P < .001) and were 3.39, 1.64, and 2.04 times longer than the time to peak aortic enhancement, respectively. Age, sex, and history of heart disease did not give additional predictive information for determining the time to peak visceral enhancement., Conclusion: The times to peak enhancement of the liver, pancreas, and jejunum are linearly related to that of the aorta. These results could potentially allow tailored patient- and organ-specific scan delay optimization at contrast material-enhanced MR image evaluation.
- Published
- 2007
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63. [Effect of the immune strategy based on BCG priming and Ag85A/GM-CSF DNA vaccine boosting in mice].
- Author
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Tang Q, Dou J, Zhao FS, Chu LL, Pan M, and Wang YF
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- Acyltransferases genetics, Animals, Antigens, Bacterial genetics, BCG Vaccine administration & dosage, Cell Line, Tumor, Enzyme-Linked Immunosorbent Assay, Granulocyte-Macrophage Colony-Stimulating Factor genetics, Killer Cells, Natural immunology, Mice, Mice, Inbred BALB C, Mycobacterium bovis genetics, T-Lymphocytes, Cytotoxic immunology, Vaccines, DNA administration & dosage, Vaccines, DNA genetics, Acyltransferases immunology, Antigens, Bacterial immunology, BCG Vaccine immunology, Granulocyte-Macrophage Colony-Stimulating Factor immunology, Mycobacterium bovis immunology, Vaccines, DNA immunology
- Abstract
Aim: To investigate immune effect in mice on the basis of BCG priming and DNA vaccine boosting, and to provide a new strategy for development of new type of anti-tuberculosis DNA vaccine further., Methods: After mice were inoculated with BCG of 1x10(6) clone formation unit each three weeks, 100 microg of DNA vaccine was injected intramuscularly in mice two times at 3 week intervals. The proliferative responses of murine cytotoxic T lymphocytes (CTL), natural killer (NK) and spleen cell to antigen 85A(Ag85A) were measured by MTT method respectively. The antibody titers and IFN-gamma level from the immunized mice were detected by ELISA., Results: The proliferative responses of CTL and spleen cell to Ag85A, as well as IFN-gamma level in mice immunized with prime-boost strategy were significantly increased respectively compared with the control mice immunized with blank plasmid or BCG only. Although NK activity was a little higher in mice immunized with prime-boost strategy than that of immunized with blank plasmid mice, it was still lower than that of mice immunized with BCG alone. The titer of the specific antibody against Ag85A in mice immunized with prime-boost strategy was also higher than that of mice immunized with DNA vaccine alone., Conclusion: The immune strategy of BCG-prime and Ag85A/GM-CSF DNA vaccine boost improve immune effect, especially the Th1 cellular immune response increase obviously. This study provides the possibility of further research for investigating protective function in immunized mice challenged by Mycobacterium tuberculosis.
- Published
- 2007
64. [Anti-tumor mechanisms of Sp2/0 tumor vaccine transfected with mIL-21 gene in mice].
- Author
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Chu LL, Dou J, Zhao FS, Tang Q, Zhang AF, Wang YF, and Gu N
- Subjects
- Animals, B7-1 Antigen immunology, B7-1 Antigen metabolism, Cell Line, Tumor, Cell Proliferation, Chemokine CXCL11 genetics, Female, Flow Cytometry, Histocompatibility Antigens Class I immunology, Histocompatibility Antigens Class I metabolism, Immunity, Cellular immunology, Killer Cells, Natural immunology, Killer Cells, Natural metabolism, Male, Mice, Mice, Inbred BALB C, Random Allocation, Reverse Transcriptase Polymerase Chain Reaction, Spleen cytology, T-Lymphocytes, Cytotoxic immunology, T-Lymphocytes, Cytotoxic metabolism, Antineoplastic Agents immunology, Cancer Vaccines immunology, Interleukins genetics
- Abstract
Aim: To explore the anti-tumor mechanism of Sp2/0-mIL-21 tumor vaccine in mice., Methods: The molecules of MHC-I and CD80 on the surface of Sp2/0-mIL-21 tumor vaccine were detected by flow cytometry (FCM) respectively. A flow cytometric CFSE-7-AAD cytotoxicity assay was used to detect the cytotoxic activities of NK cells and CTLs. The expression of I-TAC in the tumor tissue was tested by RT-PCR., Results: The expression of MHC-I molecule on the surface of tumor vaccine was up-regulated obviously. The cytotoxic activities of NK cells and CTLs were significantly enhanced in the mice inoculated with Sp2/0-mIL-21 tumor vaccine compared with the mice inoculated with Sp2/0 tumor cell in control group. The expression of I-TAC in the tumor tissue was up-regulated. The histopathologic section analysis showed more lymphocytes were infiltrated in the tumor tissue., Conclusion: Sp2/0-mIL-21 tumor vaccine can induce strong cell-mediated immune response to tumor cells after it was inoculated s.c into mice. The anti-tumor mechanisms induced by Sp2/0 tumor cell vaccine are associated with the proliferation and activation of T lymphocyte, the differentiation and maturity of NK cell, the infiltration of lymphocytes in tumor tissue, and the enharuement of cytotoxic activities of NK cells and CTLs.
- Published
- 2007
65. [Effect of gradually induced occlusal disorders on the expression of FGFR1 of the condylar cartilage in rats].
- Author
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Chu LL, Wang MQ, Li XF, and Yu SB
- Subjects
- Animals, Cartilage, Cartilage, Articular, Chondrocytes, Immunohistochemistry, Rats, Malocclusion, Mandibular Condyle metabolism, Receptor, Fibroblast Growth Factor, Type 1 biosynthesis
- Abstract
Purpose: To investigate the effect of gradually induced occlusal disorders on the expression of FGFR1 in rat condylar cartilage., Methods: A model of gradually induced occlusal disorders in rat was established.The expression of FGFR1 was detected by SABC immunocytochemistry and analyzed by density of positive cells in condylar cartilage. The data were analyzed using SPSS 11.0 software package., Results: FGFR1-positive chondrocytes were abundant in the maturative layer and hypertrophic layer, but only few FGFR1-positive chondrocytes were found in the proliferative layer. In the control group, the expression of FGFR1 increased from 6-week-old to 10-week-old rats, and then decreased and went stable. In both young and adult group, the expression of FGFR1 in the experimental group was significantly lower at 4, 6 weeks and higher at 8 weeks than that in the control group, especially in the young group(P<0.05). But no difference was found between the experimental group and the control group at 2 weeks in both groups., Conclusion: FGFR1 may play an important role in the remodling of condylar cartilage induced by occlusal disorders.
- Published
- 2007
66. [Effect of gradually induced occlusal disorders on the expression of basic fibroblast growth factor of condylar cartilage in rat].
- Author
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Chu LL, Wang MQ, Li XF, and Yu S
- Subjects
- Animals, Cartilage, Articular, Mandibular Condyle, Rats, Cartilage, Fibroblast Growth Factor 2
- Abstract
Objective: To study the effect of gradually induced occlusal disorders on the expression of basic fibroblast growth factor (bFGF) of condylar cartilage in rat., Methods: The model of gradually induced occlusal disorders was established in rat. The expression of bFGF was examined by SABC technique of immunohistochemistry. The expression of bFGF was analyzed by amount of positive cells., Results: bFGF was expressed positively in the proliferative cell layer, maturative layer and hypertrophical cell layer in the rat mandibular condyle cartilage. In control group, expression of bFGF increased from 2-week-old to 6-week-old, then it had a decrease during experiment. Compared with the control group, bFGF of experiment group was increasing at 2 week, 6 week and 8 week during experiment. But there was decreaseing at 4 week. There was no difference between young experiment group and the adult experiment group., Conclusion: The gradually induced occlusal disorders may lead to significant increase of expressiong of bFGF in condyle cartilage, which suggests that the bFGF may be involve in the procedure of repairing process of articular cartilage.
- Published
- 2007
67. PAX2 activates WNT4 expression during mammalian kidney development.
- Author
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Torban E, Dziarmaga A, Iglesias D, Chu LL, Vassilieva T, Little M, Eccles M, Discenza M, Pelletier J, and Goodyer P
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- Animals, Base Sequence, Glycoproteins chemistry, Heterozygote, In Situ Hybridization, Kidney Tubules metabolism, Mesoderm metabolism, Mice, Mice, Transgenic, Molecular Sequence Data, Wnt4 Protein, Gene Expression Regulation, Developmental, Kidney embryology, PAX2 Transcription Factor physiology, Proto-Oncogene Proteins biosynthesis, Wnt Proteins biosynthesis
- Abstract
The transcription factor PAX2 is expressed during normal kidney development and is thought to influence outgrowth and branching of the ureteric bud. Mice with homozygous null Pax2 mutations have developmental defects of the midbrain-hindbrain region, optic nerve, and ear and are anephric. During nephrogenesis, PAX2 is also expressed by mesenchymal cells as they cluster and reorganize to form proximal elements of each nephron, but the function of PAX2 in these cells is unknown. In this study we hypothesized that PAX2 activates expression of WNT4, a secreted glycoprotein known to be critical for successful nephrogenesis. PAX2 protein was identified in distal portions of the "S-shaped" body, and the protein persists in the emerging proximal tubules of murine fetal kidney. PAX2 activated WNT4 promoter activity 5-fold in co-transfection assays with JTC12 cells derived from the proximal tubule. Inspection of the 5'-flanking sequence of the human WNT4 gene identified three novel PAX2 recognition motifs; each exhibited specific PAX2 protein binding in electromobility shift assays. Two motifs were contained within a completely duplicated 0.66-kb cassette. Transfection of JTC12 cells with a PAX2 expression vector was associated with a 7-fold increase in endogenous WNT4 mRNA. In contrast, Wnt4 mRNA was decreased by 60% in mesenchymal cell condensates of fetal kidney from mice with a heterozygous Pax2 mutation. We speculated that a key function of PAX2 is to activate WNT4 gene expression in metanephric mesenchymal cells as they differentiate to form elements of the renal tubules.
- Published
- 2006
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68. [Effect of a Ruditapes philippinarum diet on the development of experimental fatty liver in rabbits].
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Sun Y, Xin YN, Zhang M, Chu LL, Zhou RR, Lü WH, and Zhang J
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- Animals, Dietary Fats, Fatty Liver etiology, Fatty Liver pathology, Female, Male, Rabbits, Random Allocation, Fatty Liver diet therapy, Mollusca
- Published
- 2006
69. Novel constructs of tuberculosis gene vaccine and its immune effect on mice.
- Author
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Dou J, Chen JS, Wang J, Chen GB, Zhao FS, Tang Q, Fang XS, Chu LL, and Pan M
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- Acyltransferases genetics, Acyltransferases immunology, Acyltransferases metabolism, Animals, Antibodies, Bacterial blood, Antibodies, Bacterial immunology, Antigens, Bacterial genetics, Antigens, Bacterial immunology, Antigens, Bacterial metabolism, Blotting, Western, Cell Line, Tumor, Cell Proliferation, DNA, Complementary genetics, Gene Expression, Granulocyte-Macrophage Colony-Stimulating Factor genetics, Granulocyte-Macrophage Colony-Stimulating Factor immunology, Granulocyte-Macrophage Colony-Stimulating Factor metabolism, Humans, Killer Cells, Natural metabolism, Mice, Mice, Inbred BALB C, Recombinant Proteins genetics, Recombinant Proteins immunology, Recombinant Proteins metabolism, Reverse Transcriptase Polymerase Chain Reaction, Spleen cytology, Spleen immunology, T-Lymphocytes, Cytotoxic immunology, T-Lymphocytes, Cytotoxic metabolism, Tuberculosis Vaccines genetics, Tuberculosis Vaccines immunology
- Abstract
A novel tuberculosis (TB) gene vaccine containing mouse granulocyte macrophage-colony stimulating factor (mGM-CSF) and a TB antigen (Ag85A) was developed in this study. The genes encoding Ag85A and mGM-CSF were amplified by PCR respectively from the Ag85A-containing pBSby5 and pC-mGM-CSF. The genes were then cloned into two different polylinker sites of plasmid pIRES, forming a novel TB gene vaccine construct pI85AGM. Following transfection of pI85AGM plasmid into 7721 cell line by Lipofectamine(TM), the expression of Ag85A and GM-CSF proteins was identified by Western blotting or RT-PCR. Then Balb/c mice were inoculated with the recombinant pI85AGM, pI85A, pIGM or plasmid alone, respectively. The activities of CTL, NK cells and the Ag85A-stimulated proliferation of spleen cells were measured by MTT method. The serum antibody against Ag85A was detected by ELISA. The results showed that the Ag85A and GM-CSF proteins could be expressed in 7721 cell line and the activity of CTLs and the proliferation of spleen cells were significantly increased in the pI85AGM-immunized mice, indicating that the pI85AGM-immunized mice could generate specific immune responses to Ag85A. This study might provide possibility for developing novel anti-TB gene vaccine.
- Published
- 2005
70. Amplification and assembly of chip-eluted DNA (AACED): a method for high-throughput gene synthesis.
- Author
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Richmond KE, Li MH, Rodesch MJ, Patel M, Lowe AM, Kim C, Chu LL, Venkataramaian N, Flickinger SF, Kaysen J, Belshaw PJ, Sussman MR, and Cerrina F
- Subjects
- Oligodeoxyribonucleotides chemistry, Oligodeoxyribonucleotides isolation & purification, Genes, Oligodeoxyribonucleotides biosynthesis, Oligonucleotide Array Sequence Analysis methods, Polymerase Chain Reaction
- Abstract
A basic problem in gene synthesis is the acquisition of many short oligonucleotide sequences needed for the assembly of genes. Photolithographic methods for the massively parallel synthesis of high-density oligonucleotide arrays provides a potential source, once appropriate methods have been devised for their elution in forms suitable for enzyme-catalyzed assembly. Here, we describe a method based on the photolithographic synthesis of long (>60mers) single-stranded oligonucleotides, using a modified maskless array synthesizer. Once the covalent bond between the DNA and the glass surface is cleaved, the full-length oligonucleotides are selected and amplified using PCR. After cleavage of flanking primer sites, a population of unique, internal 40mer dsDNA sequences are released and are ready for use in biological applications. Subsequent gene assembly experiments using this DNA pool were performed and were successful in creating longer DNA fragments. This is the first report demonstrating the use of eluted chip oligonucleotides in biological applications such as PCR and assembly PCR.
- Published
- 2004
- Full Text
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71. WT1 is a modifier of the Pax2 mutant phenotype: cooperation and interaction between WT1 and Pax2.
- Author
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Discenza MT, He S, Lee TH, Chu LL, Bolon B, Goodyer P, Eccles M, and Pelletier J
- Subjects
- Animals, DNA-Binding Proteins metabolism, Kidney embryology, Mice, Mice, Inbred C57BL, PAX2 Transcription Factor, Transcription Factors metabolism, DNA-Binding Proteins genetics, Mutation, Transcription Factors genetics, WT1 Proteins metabolism
- Abstract
Metanephric kidney development requires an inductive interaction between the ureteric bud and progenitor mesenchyme, where the early expression of two genes, Wilms' tumour 1 (WT1) and paired box 2 (Pax2), establishes critical but unknown developmental pathways. Indeed, transgenic mice with deregulated overexpression of Pax2 exhibit structural kidney defects and impaired renal function, as do mice harboring targeted disruptions and/or spontaneous mutations of either the Pax2 or WT1 genes. WT1 and Pax2 are thought to regulate each other's expression during renal development. To better define the relationship between WT1 and Pax2, we generated mouse embryos containing heterozygous mutations in both genes. WT1(+/-)/Pax2(1Neu/+) kidneys were 50% smaller than wild-type kidneys. They were characterized by severe attenuation of the renal medulla, and reduced development of calyces and the renal pelvis. Renal cortex development in compound heterozygotes culminated in fewer nephrons than in WT1(+/-), Pax2(1Neu/+) or wild-type mice. Only minor variations in the mesenchymal expression pattern of Pax2 protein, and the mRNA expression levels of Pax2 and WT1, were noted in mutant kidneys. We show that WT1 and Pax2 proteins interact in vitro and in vivo, demonstrating that WT1 and Pax2 can form a molecular complex. Our data suggest that WT1 is a modifier of the Pax2 mutant phenotype.
- Published
- 2003
- Full Text
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72. Full-length cDNAs: more than just reaching the ends.
- Author
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Das M, Harvey I, Chu LL, Sinha M, and Pelletier J
- Subjects
- Automation, Chromatography, Affinity, Cloning, Molecular, DNA Primers, DNA, Complementary biosynthesis, DNA, Complementary isolation & purification, Genetic Vectors, Genome, Human, Humans, Open Reading Frames, RNA, Messenger isolation & purification, RNA-Directed DNA Polymerase chemistry, Untranslated Regions, Gene Library
- Abstract
The development of functional genomic resources is essential to understand and utilize information generated from genome sequencing projects. Central to the development of this technology is the creation of high-quality cDNA resources and improved technologies for analyzing coding and noncoding mRNA sequences. The isolation and mapping of cDNAs is an entrée to characterizing the information that is of significant biological relevance in the genome of an organism. However, a bottleneck is often encountered when attempting to bring to full-length (or at least full-coding) a number of incomplete cDNAs in parallel, since this involves the nonsystematic, time consuming, and labor-intensive iterative screening of a number of cDNA libraries of variable quality and/or directed strategies to process individual clones (e.g., 5' rapid amplification of cDNA ends). Here, we review the current state of the art in cDNA library generation, as well as present an analysis of the different steps involved in cDNA library generation.
- Published
- 2001
- Full Text
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73. Genomic organization of the canine p53 gene and its mutational status in canine mammary neoplasia.
- Author
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Chu LL, Rutteman GR, Kong JM, Ghahremani M, Schmeing M, Misdorp W, van Garderen E, and Pelletier J
- Subjects
- Alleles, Amino Acid Sequence, Animals, Base Sequence, DNA Mutational Analysis, DNA Primers, Dogs, Female, Molecular Sequence Data, Polymerase Chain Reaction, Polymorphism, Single-Stranded Conformational, Reverse Transcriptase Polymerase Chain Reaction, Genes, p53 genetics, Mammary Neoplasms, Animal genetics
- Abstract
To determine whether canine malignancies share common genetic lesions with their human counterparts, and are thus potentially interesting model systems in which to pose questions regarding tumor etiology and progression, we have elucidated the entire exon/intron structure of the canine p53 gene. A search for p53 gene abnormalities in mammary tumor tissue was undertaken utilizing single strand conformation polymorphism analysis. Mutations were detected in exons 4, 5, 6, and 7 of the p53 gene and consisted of nonsense, splicing, and frameshift mutations. None of 11 benign tumors and 6 of 40 primary carcinomas (15%) were found to harbor subtle p53 mutations. In 14 carcinomas examined the results in primary tumors and metastases were the same. These findings implicate involvement of this gene in the genesis of some malignant canine tumors, in a fashion similar to their human counterparts.
- Published
- 1998
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74. Characterization of an abundant short interspersed nuclear element (SINE) present in Canis familiaris.
- Author
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Das M, Chu LL, Ghahremani M, Abrams-Ogg T, Roy MS, Housman D, and Pelletier J
- Subjects
- Animals, Base Sequence, DNA Primers, DNA, Complementary, Molecular Sequence Data, Polymerase Chain Reaction, Polymorphism, Genetic, Sequence Homology, Nucleic Acid, Dogs genetics, Repetitive Sequences, Nucleic Acid
- Abstract
A short interspersed nuclear element (Can SINE) of approximately 130-150 bp was cloned and characterized from Canis familiaris. We demonstrate that this element is interspersed, present approximately every 5-8.3 kbp, and many are sufficiently close to allow IRS (interspersed repetitive DNA sequences) PCR. Sequence analysis of > 20 Can SINEs from the dog has identified a conserved region that was used to design oligonucleotides for IRS PCR. Since Can SINEs are not present in human or rodent genomes, IRS PCR using oligonucleotides directed to the conserved region of Can SINEs can be used to simplify analysis of canid DNA in somatic cell hybrids, as well as in large insert cloning vectors. We demonstrate that the canid IRS products are polymorphic and could be developed as genetic markers for filter-based genotyping in this organism.
- Published
- 1998
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75. Dexamethasone suppresses apoptosis in a human gastric cancer cell line through modulation of bcl-x gene expression.
- Author
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Chang TC, Hung MW, Jiang SY, Chu JT, Chu LL, and Tsai LC
- Subjects
- Cycloheximide pharmacology, DNA Fragmentation drug effects, Dactinomycin pharmacology, Dichlororibofuranosylbenzimidazole pharmacology, Flow Cytometry, Humans, Immunoblotting, Mifepristone pharmacology, Nucleic Acid Synthesis Inhibitors pharmacology, Protein Synthesis Inhibitors pharmacology, Proto-Oncogene Proteins c-bcl-2 metabolism, RNA, Messenger analysis, RNA, Messenger metabolism, Receptors, Glucocorticoid antagonists & inhibitors, Receptors, Glucocorticoid metabolism, Stomach Neoplasms, Tumor Cells, Cultured, Up-Regulation drug effects, bcl-X Protein, Apoptosis drug effects, Dexamethasone pharmacology, Gene Expression Regulation, Neoplastic drug effects, Proto-Oncogene Proteins c-bcl-2 genetics
- Abstract
Treatment of human gastric cancer TMK-1 cells with transcription and translation inhibitors rapidly triggered cell apoptosis. Along with cell apoptosis, the Bcl-xS level was markedly upregulated suggesting a crucial role of this protein in promoting the apoptotic process. In the presence of dexamethasone, however, cell apoptosis was greatly attenuated as demonstrated by DNA histogram shift and DNA fragmentation. Studies using the glucocorticoid receptor antagonist RU486 indicated that attenuation of apoptosis was mediated through glucocorticoid receptors. Dexamethasone not only suppressed the apoptosis-associated upregulation of Bcl-xS but also enhanced the basal level of Bcl-xL in the cells. In addition, bcl-x mRNA stability was significantly extended in the presence of dexamethasone. These results indicate that dexamethasone exerted a protective effect and delayed apoptosis of TMK-1 cells by modulating bcl-x gene expression.
- Published
- 1997
- Full Text
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76. Functional characterization of human nucleosome assembly protein-2 (NAP1L4) suggests a role as a histone chaperone.
- Author
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Rodriguez P, Munroe D, Prawitt D, Chu LL, Bric E, Kim J, Reid LH, Davies C, Nakagama H, Loebbert R, Winterpacht A, Petruzzi MJ, Higgins MJ, Nowak N, Evans G, Shows T, Weissman BE, Zabel B, Housman DE, and Pelletier J
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Cloning, Molecular, DNA, Complementary isolation & purification, DNA-Binding Proteins, Gene Transfer Techniques, Histones genetics, Humans, Mice, Mice, Nude, Molecular Chaperones genetics, Molecular Sequence Data, Nuclear Proteins chemistry, Nuclear Proteins genetics, Nucleosomes chemistry, Nucleosomes genetics, Protein Binding genetics, Recombinant Proteins chemistry, Subcellular Fractions chemistry, Wilms Tumor genetics, Histones physiology, Molecular Chaperones physiology, Nuclear Proteins physiology, Nucleosomes physiology
- Abstract
Histones are thought to play a key role in regulating gene expression at the level of DNA packaging. Recent evidence suggests that transcriptional activation requires competition of transcription factors with histones for binding to regulatory regions and that there may be several mechanisms by which this is achieved. We have characterized a human nucleosome assembly protein, NAP-2, previously identified by positional cloning at 11p15.5, a region implicated in several disease processes including Wilms tumor (WT) etiology. The deduced amino acid sequence of NAP-2 indicates that it encodes a protein with a potential nuclear localization motif and two clusters of highly acidic residues. Functional analysis of recombinant NAP-2 protein purified from Escherichia coli demonstrates that this protein can interact with both core and linker histones. We demonstrate that recombinant NAP-2 can transfer histones onto naked DNA templates. Deletion mutagenesis of NAP-2 demonstrates that both NH3- and COOH-terminal domains are required for histone transfer activity. Subcellular localization studies of NAP-2 indicate that it can shuttle between the cytoplasm and the nucleus, suggesting a role as a histone chaperone. Given the potential role of the human NAP-2 gene (HGMW-approved symbol NAP1L4) in WT etiology, we have elucidated the exon/intron structure of this gene and have analyzed the mutational status of NAP-2 in sporadic WTs. Our results, coupled with tumor suppression assays in G401 WT cells, do not support a role for NAP-2 in the etiology of WT. A putative role for NAP-2 in regulating cellular differentiation is discussed.
- Published
- 1997
- Full Text
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77. Effects of transcription and translation inhibitors on a human gastric carcinoma cell line. Potential role of Bcl-X(S) in apoptosis triggered by these inhibitors.
- Author
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Chang TC, Tsai LC, Hung MW, Chu LL, Chu JT, and Chen YC
- Subjects
- Apoptosis genetics, Cycloheximide pharmacology, DNA Fragmentation, Dactinomycin pharmacology, Gene Expression drug effects, Humans, Microscopy, Fluorescence, RNA, Messenger analysis, Stomach Neoplasms ultrastructure, Tumor Cells, Cultured drug effects, Tumor Cells, Cultured ultrastructure, Apoptosis drug effects, Nucleic Acid Synthesis Inhibitors pharmacology, Protein Biosynthesis drug effects, Protein Synthesis Inhibitors pharmacology, Stomach Neoplasms genetics, Transcription, Genetic drug effects
- Abstract
The effects of the macromolecular synthesis inhibitors 5,6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB), actinomycin D, and cycloheximide on the human gastric cancer TMK-1 cell line were studied. These agents inhibited DNA, RNA, or protein synthesis efficiently and induced cell death rapidly in a wide range of concentrations. After 8 hr of exposure to these agents, the cells exhibited morphological features of apoptosis, including cell shrinkage, nuclear condensation, DNA fragmentation, and formation of apoptotic bodies. Western blot analysis revealed that these inhibitors altered the protein levels of apoptosis-related gene products such as c-Myc, Bcl-X(S), and the mutant p53 (mp53) in TMK-1 cells markedly. The c-myc mRNA and protein levels were decreased initially and were then induced markedly to a new level after 4 hr of exposure to DRB, a RNA polymerase II inhibitor. The Bcl-X(S) levels were increased rapidly after treatment with all of these agents, whereas the levels of Bcl-X(L) and Bax remained largely unchanged. Northern blot analysis indicated that the c-myc overexpression is concomitant to DRB-induced DNA fragmentation and that the increased mp53 protein level was mainly a posttranscriptional event. Our observations suggest that the up-regulation of Bcl-X(S) may serve as an important mechanism for the apoptosis triggered by these inhibitors. This study also provides evidence for the notion that interference with the cellular survival pathway may lead to apoptosis.
- Published
- 1997
- Full Text
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78. The human steroidogenic acute regulatory (StAR) gene is expressed in the urogenital system and encodes a mitochondrial polypeptide.
- Author
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Gradi A, Tang-Wai R, McBride HM, Chu LL, Shore GC, and Pelletier J
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Biological Transport, Carbonyl Cyanide m-Chlorophenyl Hydrazone pharmacology, Cell Line, Cholesterol metabolism, Cloning, Molecular, Female, Gene Expression genetics, Humans, Male, Mitochondria metabolism, Molecular Sequence Data, Neoplasms chemistry, Neoplasms genetics, Peptides chemistry, Phosphoproteins biosynthesis, Phosphoproteins metabolism, Rats, Sequence Alignment, Trypsin metabolism, Tumor Cells, Cultured, Mitochondria chemistry, Phosphoproteins genetics, Urogenital System metabolism
- Abstract
The first enzymatic step in the biosynthesis of steroid hormones occurs in the mitochondrial inner membrane and is dependent on the mobilization of cholesterol from cellular stores. We report on the isolation of a human cDNA which encodes a mitochondrial protein called steroidogenic acute regulatory (StAR) protein, implicated in transport of cholesterol into mitochondria. Nucleotide and predicted amino acid sequence analyses indicate that the human and murine polypeptides are highly conserved, sharing 87% identity with an overall homology of 92%. Analysis of the distribution of StAR mRNA transcripts in human tissues by Northern blotting reveals several mRNA species, the most abundant of which is a 1.8 kb mRNA transcript present in testes, ovaries and kidneys. Using in vitro translated protein, we demonstrate that the StAR gene product can be efficiently imported into exogenously added mitochondria.
- Published
- 1995
- Full Text
- View/download PDF
79. An efficient strategy to isolate full-length cDNAs based on an mRNA cap retention procedure (CAPture).
- Author
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Edery I, Chu LL, Sonenberg N, and Pelletier J
- Subjects
- Amino Acid Sequence, Animals, Base Sequence, Eukaryotic Initiation Factor-4E, Humans, Mice, Molecular Sequence Data, Peptide Initiation Factors isolation & purification, RNA, Messenger analysis, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins isolation & purification, Sequence Alignment, Cloning, Molecular methods, DNA, Complementary isolation & purification, Peptide Initiation Factors genetics
- Abstract
The ability to generate cDNA libraries is one of the most fundamental procedures in contemporary molecular biology. One of the major drawbacks of current methods is that most cDNAs present in any given library are incomplete, rendering the characterization of genes an inefficient and time-consuming task. We have developed an affinity selection procedure using a fusion protein containing the murine cap-binding protein (eukaryotic initiation factor 4E), coupled to a solid support matrix, that allows for the purification of mRNAs via the 5' cap structure. When combined with a single-strand-specific RNase digestion step, specific retention of complete cDNA-RNA duplexes following first-strand synthesis is achieved. This method can be used to generate cDNA libraries in which polyadenylated and nonpolyadenylated mRNAs are equally represented and to enrich for full-length or 5'-end clones, thus facilitating cDNA cloning and promoter mapping.
- Published
- 1995
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80. Energy-dependent intracellular translocation of proparathormone.
- Author
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Chu LL, MacGregor RR, and Cohn DV
- Subjects
- Anaerobiosis, Animals, Antimycin A pharmacology, Cattle, Chloroquine pharmacology, Colchicine pharmacology, Cycloheximide pharmacology, Cytochalasin B pharmacology, Deoxyglucose pharmacology, Deuterium pharmacology, Dinitrophenols pharmacology, In Vitro Techniques, Kinetics, Oligomycins pharmacology, Parathyroid Hormone biosynthesis, Rotenone pharmacology, Temperature, Tromethamine pharmacology, Vinblastine pharmacology, Parathyroid Glands metabolism, Parathyroid Hormone metabolism
- Abstract
We previously suggested that after synthesis, proparathormone is transferred from rough endoplasmic reticulum to the Golgi region where its conversion to parathormone occurs. We have attempted to define more closely this transfer process. In the first type of study, bovine parathyroid slices were incubated with [3H]leucine for 10 min and then radioisotope labeling was restricted by addition of a large excess of nonradioactive leucine. Under these conditions, more than 90% of the initially labeled proparathormone was converted to parathormone in 40 min. Lowered temperature in the chase period markedly inhibited the conversion. Several chemical agents were employed individually in the chase period to examine their effect on the conversion process. Antimycin A, dinitrophenol, oligomycin, and anaerobiosis (N2) inhibited the conversion, whereas sodium flouride and cycloheximide had no effect. In the second type of study, parathyroid slices were incubated with [3H]leucine for the entire incubation period. Lowered temperature and inhibitors of energy metabolism and microtubular function all lengthened the interval (lag) between the initial synthesis of [3H]parathormone. Cycloheximide, Tris, and chloroquine decreased the rates of protein synthesis and conversion, respectively, but none had any effect on the lag. We interpret the lag to represent the time of transit for proparathormone from rough endoplasmic reticulum to the Golgi region. We conclude that this transfer process is independent of the synthesis of the prohormone and its conversion to the hormone. Moreover, this translocation requires metabolic energy and appears to be mediated by microtubules.
- Published
- 1977
- Full Text
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81. Isolation and partial characterization of secretory protein I from bovine parathyroid glands.
- Author
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Cohn DV, Morrissey JJ, Hamilton JW, Shofstall RE, Smardo FL, and Chu LL
- Subjects
- Amino Acid Sequence, Animals, Carbohydrates, Cattle, Chromogranin A, Chromogranins, Peptide Fragments analysis, Radioimmunoassay, Trypsin, Calcium-Binding Proteins isolation & purification, Parathyroid Glands analysis, Parathyroid Hormone isolation & purification
- Abstract
Secretory protein I, a protein that is cosecreted with parathormone, has been isolated from bovine parathyroid tissue. The purification procedure was aided by the inclusion in the starting material of fresh tissue that had been incubated with radioactive amino acids to label the newly formed secretory protein I. The isolation of the secretory protein I was then followed by locating the radioactive species. Later, purification was also followed by radioimmunoassay. The procedures included salt fractionation, gel filtration, and two steps of ion-exchange chromatography, yielding a 96-fold purification of secretory protein I. The final product contained two species that were shown to be related by comparison of their tryptic peptides and the release of only a single major residue at each step of the Edman degradation. On the basis of amino acid analysis, secretory protein I contains about 30% acidic amino acid residues, contributing to an isoelectric point of 4.5, and has a minimum molecular weight of about 70 000. It contains 2.6% carbohydrate. A radioimmunoassay was established for secretory protein I. A partial amino acid sequence spanning the first 32 residues of the amino-terminal region was obtained. This portion of the structure appeared to be unrelated to those of the known parathyroid hormonal peptides.
- Published
- 1981
- Full Text
- View/download PDF
82. Biosynthesis of proparathyroid hormone and parathyroid hormone by human parathyroid glands.
- Author
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Chu LL, MacGregor RR, Liu PI, Hamilton JW, and Cohn DV
- Subjects
- Amino Acid Sequence, Amino Acids metabolism, Chromatography, Humans, Kinetics, Male, Parathyroid Glands analysis, Parathyroid Hormone analysis, Protein Precursors analysis, Protein Precursors biosynthesis, Radioimmunoassay, Tritium, Parathyroid Glands metabolism, Parathyroid Hormone biosynthesis
- Abstract
Human parathyroid glands obtained at autopsy were incubated with [(3)H]leucine and [(3)H]lysine. After incubation, nonradioactive parathyroid tissue of either human or bovine origin was added. Radioactive parathyroid hormone and proparathyroid hormone were isolated from the gland and medium by organic solvent and salt fractionation, trichloroacetic acid precipitation, Sephadex G-100 gel filtration, and carboxymethyl cellulose column chromatography. The human hormonal peptides were identified in the ion-exchange column eluates by their relatively high levels of radioactivity, their elution positions, and their immunoreactivity to anti-PTH antiserum. The time-course of radioactive amino acid incorporation into these peptides and a brief incubation of the gland with radioactive amino acids, followed by various lengths of incubation with nonradioactive amino acids, indicated that a precursor-product relationship exists for the two peptides. An alternate method for isolation of the hormone and prohormone, which involves separation of peptides by urea-polyacrylamide gel electrophoresis, confirmed the identities of the human parathyroid hormone and proparathyroid hormone.
- Published
- 1973
- Full Text
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83. Similarity of secretory protein I from parathyroid gland to chromogranin A from adrenal medulla.
- Author
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Cohn DV, Zangerle R, Fischer-Colbrie R, Chu LL, Elting JJ, Hamilton JW, and Winkler H
- Subjects
- Amino Acid Sequence, Amino Acids analysis, Animals, Cattle, Chromogranin A, Cross Reactions, Immunodiffusion, Molecular Weight, Structure-Activity Relationship, Adrenal Medulla analysis, Calcium-Binding Proteins isolation & purification, Chromaffin Granules analysis, Chromaffin System analysis, Chromogranins isolation & purification, Nerve Tissue Proteins isolation & purification, Parathyroid Glands metabolism
- Abstract
We have compared the amino acid and carbohydrate compositions, partial amino acid sequences, immunological crossreactivity, and physical properties of secretory protein I of the parathyroid gland and chromogranin A of adrenal gland. This comparison indicates that these proteins are similar molecules. Because secretory protein I is present in secretory granules containing parathormone and is cosecreted with the hormone, and because chromogranin A is contained within chromaffin granules and, likewise, is secreted with the catecholamines, the present observations raise the possibility that this class of protein plays a general role in hormone secretion or storage mechanisms.
- Published
- 1982
- Full Text
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84. The NH2-terminal amino acid sequence of human proparathyroid hormone by radioisotope microanalysis.
- Author
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Huang WY, Chu LL, Hamilton JW, McGregor DH, and Cohn DV
- Subjects
- Amino Acid Sequence, Amino Acids analysis, Animals, Cattle, Chromatography, Thin Layer, Humans, Hydrolysis, Methods, Microchemistry, Protein Precursors, Species Specificity, Tritium, Parathyroid Hormone
- Published
- 1975
- Full Text
- View/download PDF
85. Structural characterization of adrenal chromogranin A and parathyroid secretory protein-I as homologs.
- Author
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Hamilton JW, Chu LL, Rouse JB, Reddig K, and MacGregor RR
- Subjects
- Adrenal Glands analysis, Amino Acids analysis, Animals, Cattle, Chemical Phenomena, Chemistry, Chromatography, Gel, Chromatography, High Pressure Liquid, Chromatography, Ion Exchange, Chromogranin A, Calcium-Binding Proteins isolation & purification, Chromogranins isolation & purification, Nerve Tissue Proteins isolation & purification
- Abstract
We have isolated and purified adrenal chromogranin A (Ch A) for the purpose of making structural comparisons to parathyroid secretory protein-I (SP-I), because our earlier data indicated these two molecules may be the same protein. An improved purification step, using high-performance liquid chromatography (HPLC), has enabled us to demonstrate that both SP-I and Ch A consists of two species, one of approximately 72,000 Da and one of approximately 66,000 Da. The amino acid composition is the same for all four species. The difference in molecular mass is assumed to be due to carbohydrate content. Cyanogen bromide digestion of each of the four samples, followed by HPLC separation of the generated peptides, resulted in a chromatographic profile that was the same for each digest. Amino acid analysis of the eight peptide fragments obtained from each digest indicates that both species of Ch A and both species of SP-I yielded the same peptide mixtures following this cleavage reaction. One large (approximately 50,000 Da) CNBr peptide was obtained and seven smaller ones, one of which contains cysteine. The large fragment behaved similarly to the intact molecule in a radioimmunoassay. HPLC separation of tryptic digests of Ch A (72,000 Da) and SP-I (72,000 Da) also resulted in elution profiles that were very similar to each other. Amino acid analysis revealed 23 peptides common to each digest. Ch A contained four peptides ranging in size from 4 to 30 residues that were not observed in the SP-I digest. SP-I contained two peptides, each with about 30 residues, that were not found in the Ch A digest. Nothing unusual was noted in any of the uncommon peptides. Thus, both a chemical and an enzymatic digestion of these molecules followed by analysis of the peptides generated, indicates that SP-I and Ch A are nearly identical homologs.
- Published
- 1986
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86. Conversion of proparathyroid hormone to parathyroid hormone: the use of amines as specific inhibitors.
- Author
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Chu LL, Macgregor RR, Hamilton JW, and Cohn DV
- Subjects
- Amides pharmacology, Animals, Buffers, Cattle, Chromatography, Diethylamines pharmacology, Dose-Response Relationship, Drug, Glycine pharmacology, Golgi Apparatus ultrastructure, In Vitro Techniques, Leucine metabolism, Parathyroid Glands ultrastructure, Time Factors, Tritium, Parathyroid Hormone biosynthesis, Tromethamine pharmacology
- Published
- 1974
- Full Text
- View/download PDF
87. Studies on the subcellular localization of proparathyroid hormone and parathyroid hormone in the bovine parathyroid gland: separation of newly synthesized from mature forms.
- Author
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MacGregor RR, Chu LL, Hamilton JW, and Cohn DV
- Subjects
- Animals, Biological Assay, Carbon Radioisotopes, Cattle, Centrifugation, Density Gradient, Chromatography, Gel, Chromatography, Ion Exchange, Cytosol analysis, Deoxycholic Acid, Immunoassay, Leucine metabolism, Lysine metabolism, Microscopy, Electron, Parathyroid Hormone biosynthesis, Rats, Subcellular Fractions analysis, Tritium, Parathyroid Glands cytology, Parathyroid Hormone analysis
- Published
- 1973
- Full Text
- View/download PDF
88. Disruption of the Golgi zone and inhibition of the conversion of proparathyroid hormone to parathyroid hormone in human parathyroid tissue by tris(hydroxymethyl)aminomethane.
- Author
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McGregor DH, Chu LL, MacGregor RR, and Cohn DV
- Subjects
- Adenoma ultrastructure, Cell Nucleus drug effects, Humans, Hyperplasia pathology, Mitochondrial Swelling drug effects, Parathyroid Diseases pathology, Parathyroid Glands metabolism, Parathyroid Glands ultrastructure, Parathyroid Hormone biosynthesis, Parathyroid Neoplasms ultrastructure, Protein Precursors biosynthesis, Golgi Apparatus drug effects, Parathyroid Glands drug effects, Parathyroid Hormone antagonists & inhibitors, Protein Precursors antagonists & inhibitors, Tromethamine pharmacology
- Abstract
Tris(hydroxymethyl)aminomethane (Tris, Tromethamine, THAM) and other non-amphoteric amines were previously reported to inhibit the conversion of proparathyroid hormone to parathyroid hormone in bovine parathyroid glands incubated in vitro. This inhibition correlated with a striking dilation of the Golgi complex. This work has now been extended to normal, hyperplastic, and adenomatous parathyroid glands from human subjects. The tissues were incubated for up to 3 hours with 3H-leucine in physiologic solutions (control) or in the same solutions containing 50 mM Tris. In one case, diethylamine also was tested. Electron microscopy revealed that the amines produced a dilation of the Golgi complex and swelling of vesicles, predominantly in the region of the Golgi zone. Other organelles were normal in appearance. During the same period, Tris reduced by sixfold the ratio of the parathyroid hormone to proparathyroid hormone, from a control value of 2:1 to 1:3. It was apparent that Tris exerted the same biochemical and morphologic actions in human parathyroid tissues as it was previously shown to do in bovine glands. These studies support the concept that the Golgi zone is that region in the parathyroid gland in which proparathyroid hormone to parathyroid hormone conversion is initiated and that Tris inhibits this conversion through disruption of the converting site.
- Published
- 1977
89. Effects of isoproterenol and cycloheximide on parathyroid secretion.
- Author
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Chu LL, MacGregor RR, and Hamilton JW
- Subjects
- Animals, Calcium metabolism, Calcium-Binding Proteins metabolism, Cattle, Chromogranin A, Chromogranins, Mathematics, Parathyroid Glands drug effects, Parathyroid Hormone metabolism, Cycloheximide pharmacology, Isoproterenol pharmacology, Parathyroid Glands metabolism
- Abstract
Tissue slices or dispersed cells of bovine parathyroid gland were incubated with [3H]leucine to label the intracellular proteins and then tested for their secretory response to isoproterenol and cycloheximide at different calcium concentrations. Secretion of the newly synthesized as well as the older PTH and SP-I was stimulated by isoproterenol at all calcium levels tested, even when it was maximally enhanced by low calcium. Cycloheximide interfered with neither the secretory process nor the secretory response to different stimuli, but decreased the amount of PTH and SP-I secreted. We conclude that the inhibitor decreased the secretion by reducing the supply of PTH and SP-I. Calculations derived from the data reveal that, under most secretory conditions, newly synthesized PTH contributed a major portion of the total hormone secretion in bovine parathyroid cells.
- Published
- 1983
- Full Text
- View/download PDF
90. Functioning oxyphil adenoma of parathyroid gland. An ultrastructural and biochemical study.
- Author
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McGregor DH, Lotuaco LG, Rao MS, and Chu LL
- Subjects
- Adenoma ultrastructure, Adult, Aged, Endoplasmic Reticulum ultrastructure, Female, Golgi Apparatus ultrastructure, Humans, Male, Middle Aged, Mitochondria ultrastructure, Parathyroid Glands metabolism, Parathyroid Glands ultrastructure, Parathyroid Neoplasms ultrastructure, Adenoma metabolism, Parathyroid Hormone metabolism, Parathyroid Neoplasms metabolism
- Abstract
Oxyphil cells and oxyphil cell adenomas of parathyroid glands are, in most instances, regarded to be nonfunctioning. Although 21 cases of hyperparathyroidism associated with parathyroid oxyphil cell adenoma have been reported, secretion of hormone by these tumors has not been conclusively demonstrated. A parathyroid adenoma, diagnosed by light microscopy as oxyphil type, together with the results from ultrastructural and biochemical studies of the patient's adenomatous tissue, are reported here. The patient, a 64-year-old male, was found to have elevated serum calcium, low serum phosphorus, and elevated serum immunoreactive parathormone: findings consistent with hyperparathyroidism. After excision of two small normal-appearing glands and one greatly enlarged (1.9 g) parathyroid gland, those laboratory values returned to normal. Light microscopy of the enlarged parathyroid indicated that it consisted almost entirely of an oxyphil adenoma. Electron microscopy revealed that the adenoma was composed mainly of mitochondria-rich oxyphil cells but also of interspersed transitional oxyphil cells and rare scattered chief cells. Golgi zones, rough endoplasmic reticulum, and prosecretory and secretory-like granules were observed in some oxyphil cells, in most transitional oxyphil cells, and in the infrequent chief cells. Thus, many of these cells appear to contribute to the production and secretion of parathormone. Biochemical studies performed directly on the adenomatous tissue demonstrated that it was able to synthesize proparathormone and parathormone, although the proportion of hormonal peptide synthesis relative to that of the total protein synthesis in this tissue was much smaller (0.9%) than that found in normal parathyroid tissue (5.7%). There was a small increase in immunoreactive parathormone when the adenoma tissue was incubated in a low-calcium medium. These findings indicate that this oxyphil adenoma of the parathyroid gland synthesized and secreted parathormone, apparently to some extent autonomously, but suggest that its capacity to do so was largely dependent on its component of cells other than fully developed oxyphil cells, such as transitional oxyphil cells.
- Published
- 1978
91. Cosecretion of secretory protein-I and parathormone by dispersed bovine parathyroid cells.
- Author
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Cohn DV, Morrissey JJ, Shofstall RE, and Chu LL
- Subjects
- Animals, Calcium pharmacology, Cattle, Chromogranin A, Chromogranins, In Vitro Techniques, Parathyroid Glands cytology, Calcium-Binding Proteins metabolism, Parathyroid Glands metabolism, Parathyroid Hormone metabolism
- Abstract
A RIA with a minimal sensitivity of 0.5 ng protein was developed for the measurement of bovine secretory protein-I (SP-I). With this assay and a previously established RIA for parathormone, the secretion and cell content of SP-I and parathormone were determined in dispersed bovine parathyroid cells. SP-I and parathormone were secreted in linear fashion over a 2-h period. The net secretion of both of these proteins diminished progressively as the concentration of calcium was raised from 0.25 mM to 3.0 mM. The molar ratio for the secreted proteins and those remaining in the cell varied from experiment to experiment but on average was 0.70 +/- 0.07 for the secreted proteins and 0.47 +/- 0.03 for the cellular proteins. Possible explanations for the difference in the ratio of SP-I to parathormone between cellular and secreted proteins include 1) a preferential secretion of SP-I; 2) a preferential intracellular degradation of SP-I; 3) a preferential postsecretory degradation of parathormone, or 4) differential affinities of potential fragments of either or both proteins for their antisera. These results suggest that SP-I and parathormone bear close but not identical metabolic and secretory fates.
- Published
- 1982
- Full Text
- View/download PDF
92. Conversion of proparathyroid hormone to parathyroid hormone by a particulate enzyme of the parathyroid gland.
- Author
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MacGregor RR, Chu LL, and Cohn DV
- Subjects
- Animals, Cattle, Hydrogen-Ion Concentration, Kinetics, Peptide Hydrolases isolation & purification, Protein Precursors, Subcellular Fractions enzymology, Trypsin metabolism, Parathyroid Glands enzymology, Parathyroid Hormone biosynthesis, Peptide Hydrolases metabolism
- Abstract
The conversion of proparathyroid hormone (proparathormone) to parathyroid hormone (parathormone) by subcellular fractions of the bovine parathyroid has been investigated. The identification of the conversion product as parathormone was established by its elution postion during ion exchange chromatography and gel filtration, and by partial amino acid sequence analysis of its NH2-terminal region. Total homogenates and derived subcellular fractions (600 X g pellet, 5,000 X g pellet, 20,000 X g pellet, 190,000 X g pellet, and 190,000 X g supernatant) all catalyzed the conversion of exogenous [3H]- or [14C]prohormone. Over 60% of the converting activity was in the particulate fractions; the 190,000 X g particulate fraction contained the highest specific converting activity. The converting activity appeared to be an integral component of the membranes since it could only be partially removed by extraction with Triton X-100. The production of parathormone by the particulate converting enzyme increased with time and the concentration of enzyme protein. The optimum pH range was between 7 and 9, and the enzyme was inactive below pH 6. Conversion by the particulate enzyme was inhibited by benzamidine or chloroquine, but not by pancreatic trypsin inhibitor, indicating its dissimilarity to trypsin. When a mixture of [14C]proparathormone and [3H]parathormone was used as substrate, the particulate enzyme did not metabolize the hormone despite over 70% conversion of the prohormone to hormone and other peptides. There was a close correlation between the subcellular distribution of converting activity and that of newly formed parathormone found in the membrane fraction. These data suggest that the particulate converting activity is that concerned with the formation of parathormone in vivo.
- Published
- 1976
93. Interaction of parathyroid hormone with membranes of kidney cortex: degradation of the hormone and activation of adenylate cyclase.
- Author
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Chu LL, Forte LR, Anast CS, and Cohn DV
- Subjects
- Animals, Cell Membrane enzymology, Cell Membrane metabolism, Enzyme Activation, In Vitro Techniques, Kidney Cortex enzymology, Rats, Adenylyl Cyclases metabolism, Kidney Cortex metabolism, Parathyroid Hormone metabolism
- Abstract
Plasma membranes prepared from rat renal cortex contain both a parathyroid hormone-sensitive adenylate cyclase and a potent proteolytic activity which degrades the hormone into peptide fragments. The degree and pattern of degradation was determined by subjecting incubation mixtures to gel filtration and ion exchange chromatography. Estimation of the degree of degradation by acid precipitation of the intact hormone was inadequate since metabolism of the hormone apparently generated acid-insoluble fragments. When parathyroid hormone was incubated with membrane fraction, the capacity of its stimulatory effect on adenylate cyclase decreased steadily. This decrease of PTH activitiy could be closely related to the degradation of intact hormone by the same membrane preparation. The adenylate cyclase and degradative activity appeared to exist in similar membrane structures since they could not be separated by centrifugation through sucrose density gradients. The degradation of the hormone could not be inhibited by Trasylol and pancreatic or soybean trypsin inhibitors and was only slightly inhibited by ribonuclease and benzamidine. Histone (1 mg per ml), on the other hand, was able to decrease the degradation of the hormone and prevent the loss of its activity. Radioimmunoassay of the incubation mixtures showed that the rapid degradation of both amino- and carboxy-terminal regions of the hormone was prevented by histone. The oxidized, inactive hormone was also degraded to the same extent by the renal cortical membrane. Furthermore, the degradative activity was also found in plasma membrane preparations of renal medulla and liver. This lack of hormone and tissue specificity suggests that similar degradative activity exists in all tissues and that caution should be exercised in estimating hormonal potency based on activation of adenylate cyclase.
- Published
- 1975
- Full Text
- View/download PDF
94. Studies on the biosynthesis of rat parathyroid hormone and proparathyroid hormone: adaptation of the parathyroid gland to dietary restriction of calcium.
- Author
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Chu LL, MacGregor RR, Anast CS, Hamilton JW, and Cohn DV
- Subjects
- Adaptation, Physiological, Animals, Calcium blood, Cattle, Chromatography, Gel, Chromatography, Ion Exchange, Culture Techniques, Female, Iodine Isotopes, Leucine metabolism, Lysine metabolism, Molecular Weight, Parathyroid Hormone metabolism, Peptides metabolism, Radioimmunoassay, Rats, Time Factors, Tritium, Calcium, Dietary, Parathyroid Glands metabolism, Parathyroid Hormone biosynthesis
- Published
- 1973
- Full Text
- View/download PDF
95. Partial purification of parathyroid hormone from chicken parathyroid glands.
- Author
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MacGregor RR, Chu LL, Hamilton JW, and Cohn DV
- Subjects
- Animals, Calcium metabolism, Carbon Isotopes, Cattle, Chemical Precipitation, Chickens, Chromatography, Gel, Chromatography, Ion Exchange, Citrates metabolism, Cross Reactions, Electrophoresis, Polyacrylamide Gel, Immune Sera, Mice, Tritium, Vitamin D Deficiency, Parathyroid Hormone isolation & purification
- Published
- 1973
- Full Text
- View/download PDF
96. Calcemic fraction-A: biosynthetic peptide precursor of parathyroid hormone.
- Author
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Cohn DV, Macgregor RR, Chu LL, Kimmel JR, and Hamilton JW
- Subjects
- Acrylamides, Amino Acids analysis, Animals, Antibody Specificity, Bone Resorption, Calcium biosynthesis, Cattle, Chemical Phenomena, Chemistry, Chromatography, Chromatography, Gel, Electrophoresis, Immune Sera, Isotope Labeling, Kinetics, Leucine metabolism, Lysine metabolism, Molecular Weight, Parathyroid Glands metabolism, Parathyroid Hormone biosynthesis, Peptide Biosynthesis, Sodium Dodecyl Sulfate, Time Factors, Tritium, Calcium analysis, Parathyroid Hormone analysis, Peptides analysis
- Abstract
Calcemic fraction-A (CF-A) is a biologically active, hypercalcemic and bone resorptive peptide, which was detected in, and isolated from, bovine parathyroid glands [Hamilton et al. (1971) Endocrinology 89, 1440-1447]. It has been further purified, and its relationship to parathyroid hormone clarified. The peptide is present in fresh glands at a concentration of about 3 mug/g (parathyroid hormone, 100 mug/g). It contains 109 amino acids (hormone, 84), each of which is present in equal or greater molar ratio than in the hormone. Its molecular weight, calculated from amino-acid composition, is 12,144; determined by dodecyl sulfate-polyacrylamide gel electrophoresis, it is 12,500 (hormone, 9563). Per mole, it reacts with antiserum to parathyroid hormone to an extent of 7-10% that of the hormone, and is about 50% as active in its hypercalcemic and bone resorptive properties in the appropriate assays. Time course and pulse-chase experiments with parathyroid gland slices, in which the incorporation of amino acid into isolated peptide and hormone were measured, indicate that the hormone is made from a protein precursor; the patterns of incorporation of radioactivity are those that would be predicted from a precursor-product relationship. When the large peptide was incubated with parathyroid gland extracts it was partially converted to a molecule that appeared to be the hormone, as based upon its coelution with marker hormone from ion-exchange columns. Finally, tryptic digestion of the peptide increased the immunoreactivity of the sample in accord with the known greater immunoreactivity of the hormone than the peptide. On the basis of these results, it is proposed that the peptide is a biosynthetic precursor of the hormone in bovine parathyroid gland.
- Published
- 1972
- Full Text
- View/download PDF
97. Cordycepin and alpha-amanitin: inhibitors of transcription as probes of aldosterone action.
- Author
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Chu LL and Edelman IS
- Subjects
- Adrenal Glands physiology, Adrenalectomy, Animals, Basidiomycota, Biological Transport drug effects, Bufo marinus, Cell Nucleolus enzymology, Cell Nucleus enzymology, Cytoplasm enzymology, DNA-Directed RNA Polymerases antagonists & inhibitors, DNA-Directed RNA Polymerases metabolism, Epithelial Cells, Epithelium metabolism, Kidney cytology, Kidney drug effects, Kidney enzymology, Male, Peptides, Cyclic pharmacology, Rats, Sodium metabolism, Tritium, Urinary Bladder cytology, Urinary Bladder drug effects, Aldosterone pharmacology, Deoxyadenosines pharmacology, Kidney metabolism, Mycotoxins pharmacology, Oligopeptides pharmacology, Transcription, Genetic drug effects, Urinary Bladder metabolism
- Published
- 1972
- Full Text
- View/download PDF
98. A bioassay for parathyroid hormone based on hormonal inhibition of CO2 production from citrate in mouse calvarium.
- Author
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Chu LL, Macgregor RR, Hamilton JW, and Cohn DV
- Subjects
- Animals, Bone Resorption metabolism, Calcitonin, Carbon Isotopes, Cattle, In Vitro Techniques, Methods, Rats, Biological Assay, Carbon Dioxide metabolism, Citrates metabolism, Parathyroid Hormone, Skull metabolism
- Published
- 1971
- Full Text
- View/download PDF
99. On the mechanism of iron-induced synthesis of apoferritin in HeLa cells.
- Author
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Chu LL and Fineberg RA
- Subjects
- Antibodies, Carbon Isotopes, Cycloheximide pharmacology, Dactinomycin pharmacology, Deferoxamine pharmacology, Ferritins metabolism, HeLa Cells drug effects, Humans, Iron Isotopes, Leucine metabolism, Phenanthrolines pharmacology, Time Factors, Ferritins biosynthesis, HeLa Cells metabolism, Iron
- Published
- 1969
100. The isolation and partial purification of a non-parathyroid hormone calcemic fraction from bovine parathyroid glands.
- Author
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Hamilton JW, Macgregor RR, Chu LL, and Cohn DV
- Subjects
- Animals, Biological Assay, Carbon Isotopes, Cattle, Chromatography, Gel, Chromatography, Ion Exchange, Citrates metabolism, Mercaptoethanol, Parathyroid Hormone antagonists & inhibitors, Parathyroid Hormone biosynthesis, Radioimmunoassay, Rats, Trichloroacetic Acid, Tritium, Calcium metabolism, Parathyroid Glands analysis, Parathyroid Hormone isolation & purification
- Published
- 1971
- Full Text
- View/download PDF
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