Designed a pair of primers through modifying N-terminal bases (5bps) of gene after ATG but not changing amino acid, and amplified a smaller mutated gene sequnce (468bp) containing two protective antigenic determinants from pBlue-BoNTaHc, N-terminal codon of mutated gene fragment is changed from low to high frenqence in E. coli. Mutated gene was ligated into pGEM-T vector and sequenced, then, cloned into a expression plasmid pBV220. As a result, cloned gene was expressed in insoluble form by temperature inducing (from 30 degrees C to 42 degrees C) in E. coli. Expression product is 40% of total proteins and is of specific binding activity to antibody in ELISA. The successful modification and high level expression of protective fragment of botulinum neurotoxin serotype A(BoNTaHc468) gene is conducive to further study on antitoxin and vaccine.
Bacterial Vaccines ; immunology ; Botulinum Toxins, Type A ; genetics ; immunology ; Clostridium botulinum type A ; immunology ; Enzyme-Linked Immunosorbent Assay ; Escherichia coli ; genetics ; Plasmids ; Polymerase Chain Reaction ; Recombinant Proteins ; biosynthesis

OBJECTIVES: Oxidized LDL is thought to play a key role in atherogenesis. Among their wide variety of biological properties, oxidized LDL injures the endothelium as an early event of atherogenesis. However, the mechanisms by which oxidized LDL injures endothelial cells are not definitely known. In order to evaluate the involvement of GTP-binding protein on the mechanism of apoptosis, we studied the effects of pertussis and cholera toxin on oxidized LDL-induced apoptosis in bovine aortic endothelial cells(BAECs). METHODS: Oxidized LDL elicited apoptosis in cultured BAECs as shown by characteristic morphological and biochemical changes. Chromatin condensation and nucleus fragmentation were visualized by using fluorescence microscopy of intact cells staining by acridine orange/ ethidium bromide. DNA fragmentation was quantified by an ELISA with specific antibody for bromodeoxyuridine- labelled DNA fragments and confirmed with DNA ladder formation. RESULTS: Studies using a combination of bacterial toxins which change the function of GTP-binding protein suggest that oxidized LDL-induced apoptosis was regulated by GTP-binding protein. Oxidized LDL-induced apoptosis was not changed by pretreatment of BAECs with pertussis toxin. In contrast, pretreatment with cholera toxin completely prevented the oxidized LDL- induced apoptosis. CONCLUSION: These results show that oxidized LDL induces apoptosis of BAECs and suggest that cholera toxin-sensitive G-proteins are involved in signal transduction of oxidized LDL-induced apoptosis of BAEC.
Apoptosis* ; Atherosclerosis ; Bacterial Toxins ; Cholera Toxin* ; Cholera* ; Chromatin ; DNA ; DNA Fragmentation ; Endothelial Cells* ; Endothelium ; Enzyme-Linked Immunosorbent Assay ; Ethidium ; GTP-Binding Proteins ; Lipoproteins* ; Microscopy, Fluorescence ; Pertussis Toxin ; Signal Transduction ; Whooping Cough*

A completely synthetic gene encoding the He domain of Clostridium botulinum neurotoxin serotype A (AHc, 1287 bp, 429 aa, -50 kD) was constructed with oligonucleotides. After expressed in Escherichia coli, soluble product AHc was gained and verified by SDS-PAGE and Western blot analysis. The expressive level of recombinant AHc in E. coli was very high (36%-53% of soluble total proteins) and the purified yield was more than 30 mg/L by one-step purification. Then, the purified AHc was used to vaccinate Balb/c mice, which developed a strong and specific immune response as expected following administration of AHe protein via the subcutaneous route. Results from BoNT/A neutralization assay showed that the serum from mice vaccinated with AHc contained high titer protective antibody. These results showed that the soluble, stable and high-levelly expressive AHc not only could be produced by the prokaryotic expression system built in our lab, but also owned strong immunogenicity to prepare antitoxin for treatment and as sub-unit candidate vaccine for prophylaxis against botulinum toxin serotype A.
Animals ; Antibodies, Bacterial ; blood ; Bacterial Vaccines ; genetics ; immunology ; Botulinum Toxins, Type A ; biosynthesis ; genetics ; immunology ; Botulism ; immunology ; prevention & control ; Clostridium botulinum type A ; genetics ; immunology ; Escherichia coli ; genetics ; metabolism ; Female ; Lymphocyte Activation ; Mice ; Mice, Inbred BALB C ; Recombinant Proteins ; biosynthesis ; genetics ; immunology ; T-Lymphocytes ; immunology ; Vaccines, DNA ; genetics ; immunology

After direct superior rectus muscle injection of BtA in rabbit eyes, we examined the ultrasructural changes of the muscles from 1 day to 8 weeks after injection. The most profound changes seen at electron microxcopic levels after BtA injedtion were early vacuolization of the sarcoplasmic structure, extensive damage of myofibril, degeneration of the postjunctional fold and widening of the synaptic cleft. Myofiber changes were reversible with no apparent long-term consequence. However, most of the degenerations of myoneuronal junction were still present in 8 weeks post-injedtion. Comparing myotoxic effects according to rabbit age, the botulinum toxin seems to make more severe histologic damage in the fibers of the two-month old than six-month old or older. AchE activity of injection group is mildly decrease in number of positive fibers rather than control group, which was not statistically significant in the quantitative analysis. In conclusion the early vacuolization and degeneration of the sarcoplasmic structure, and degeneration of the postjunctional folds after toxin injection in the muscles are most likely due to a direct myotoxic effect of BtA.
Botulinum Toxins* ; Botulinum Toxins, Type A ; Muscles ; Myofibrils

No abstract available.
Botulinum Toxins* ; Botulinum Toxins, Type A* ; Hyperhidrosis*

Background: Resting tremor is a prominent cardinal motor symptom of Parkinson’s disease (PD). In some cases, the tremor may be refractory to dopaminergic and anticholinergic treatment. Multiple studies were previously done to evaluate the effectiveness of Botulinum Neurotoxin A (BoNT/A) with essential tremors and dystonia, but data regarding its use on tremors of PD is still lacking. Objective: This meta-analytic study aims to determine the effectiveness of BoNT/A in treating tremors of patients with PD. Data Sources: Data Sources: Researches were searched at PubMed, ScienceDirect and EBSCO Host. Review Methods: Articles on the effect of BoNT/A on PD hand tremors were searched. Studies and data pertaining to non-PD tremors like essential tremors excluded in the analysis due to difference in pathophysiology. Standardized mean difference was used as the effect measure and was computed with Review Manager version 5.4 software. Results: Three open label studies were used for final analysis in this study. Studies included are those pertaining to tremors due to PD. Pooled estimates showed a significant change in decreasing tremor score after BoNT/A injection. Conclusion Botulinum Toxin A injections can be used to manage PD tremors effectively.
Botulinum Toxins, Type A ; Tremor

PURPOSE: We conducted the study to evaluate the immunogenicity and safety of three component DTaP vaccine (Infanrix(R)) in a group of Korean healthy infants on a three-dose primary vaccination. And we compared the immunogenicity of this DTaP vaccine with two component DTaP vaccine which has been widely used in Korea. METHODS: We enrolled one hundred fifty one healthy infants aged 8-9 weeks. These infants were vaccinated at age 2, 4 and 6 months of age with three component DTaP vaccine. Solicited adverse events were actively monitored for 72 hours following each vaccination, and all adverse events after each vaccination were observed for three weeks. Anti-diphtheria toxoid Ab., anti-tetanus toxoid Ab., anti-pertussis toxin Ab., anti-filamentous hemagglutinin Ab., and anti-pertactin Ab. were measured using ELISA for assessing immunogenicity of study vaccine in 60 infants. Immunogenicity analysis of two component DTaP vaccine was performed with same methods in 14 infants as control. RESULTS: The seroconversion rates of anti-diphtheria toxoid Ab, anti-tetanus toxoid Ab. anti- filamentous hemagglutinin Ab. were 100% in both group. Seroconversion rate of anti-pertactin Ab in study group was 100%, but the rate in control group was 50%. However, geometric mean concentration of anti-pertussis toxin Ab. was higher in control group. Mild local and systemic reactions were observed within three days after vaccination, and no serious adverse events related study vaccine were happened during study period. CONCLUSIONS: Our study results suggest that three component DTaP vaccine (Infanrix(R)) is a well- tolerable and high immunogenic vaccine, especially anti-Pertactin Ab. of the study vaccine is very immunogenic. It can be available as routine DTaP vaccination in our infants.
Diphtheria-Tetanus-acellular Pertussis Vaccines* ; Enzyme-Linked Immunosorbent Assay ; Hemagglutinins ; Humans ; Infant* ; Korea ; Pertussis Toxin ; Vaccination

PURPOSE: To investigate the efficacy of botulinum toxin type A chemodenervation in various types of esotropia(ET). METHODS: Enrolled eleven esotropic patients treated with botulinum toxin type A and examined the amount of esotropic correction and success rate of less than 10 PD (prism diopter) of postinjection deviation at 5 months after injection. RESULTS: Among the eleven esotropic patients, there were 7 cases of infantile ET, 2 cases of partially accommodative ET and 2 cases of basic ET. The mean preinjection deviation was 30.5 +/- 7.3 PD. The amount of correction was 17.7 +/- 4.3 PD and correction rate of deviation was 57.5%. In 6 of 11 cases, the postinjection deviation was within 10 PD and therefore success rate was 54.5%. In 5 patients (45.5%), their ET was undercorrected. In patients with under 20 PD of preinjection deviation, success rate was 75% (3/4) and those with over 20PD, the rate was 42.9% (3/7). In partially accommodative ET, 2 (100%) of 2 cases were aligned within 10PD. CONCLUSIONS: In comitant ET, botulinum toxin chemodenervation shows high undercorrection rate. The effects of botulinum toxin type A chemodenervation were better for relatively small angle and in partially accommodative ET than nonaccommodative one. But further study with more cases is needed.
Botulinum Toxins* ; Botulinum Toxins, Type A* ; Esotropia* ; Humans ; Nerve Block

We studied the safety and complication associated with treatment with botulinum A toxin chemodenervation. 70 eyes was entrolled and treated with botulinum A toxin chemodenervation. They were examed at 1 day, 1 week, 2 weeks, 1 month, 3 months and 6 months after the injection. Ptosis developed in 19 of 70 eyes(27.1%), but all of them recovered fully at 6 months after injection. A vertical deviation was noted in 19 eyes(27.1%), but all except one recovered fully at 6 months. Ptosis and vertical deviation developed more frequently in the group with the medial rectus muscle injection than with lateral rectus muscle injection. Subconjunctival he.morrhage and headache were also noted in 1 eye, respectively, but they disappeared at 4 weeks after the injection. Aggrevation of diplopia after injection was noted in 1 eye, which disappeared at 3 weeks after injection. Botulinum A toxin chemodenervation can be considered to be a safe method for the treatment of strabismus without the long-lasting serious complications.
Botulinum Toxins* ; Botulinum Toxins, Type A* ; Diplopia ; Headache ; Nerve Block ; Strabismus*

BACKGROUND:This study was designed to evaluate the efficacy of subcutaneous Botulinum toxin type A (BoNT-A) injection for treating chronic medial knee pain with osteoarthritis. METHODS:A randomized, double-blind, placebo-controlled clinical trial was conducted at a university hospital in Korea.The subjects suffering from chronic medial knee pain with osteoarthritis were randomly allocated to either the BoNT-A (treatment, n = 23) group or the normal saline (placebo, n = 27) group.Injections were given to 10 points per unilateral knee along the course of the saphenous nerve.A total of 90 units of BoNT-A (60 units at baseline and 30 units after 1 week) was given for treating unilateral knee pain and a total of 180 units of BoNT-A was given for treating bilateral knee pain.The placebo group received the same volume of normal saline. RESULTS:The Visual Analog Scale (VAS) pain score was significantly decreased in both the BoNT-A and normal saline groups 1, 4 and 12 weeks after injection.After adjusting for covariates, BoNT-A had a 0.788 times higher effect to decrease the VAS score than did normal saline, but the effect was marginally significant (P = 0.050). CONCLUSIONS:Subcutaneous injection along the course of the saphenous nerve significantly reduces chronic medial knee pain. The pain reduction effect of BoNT-A is higher than that of placebo, but the effect is marginally significant
Botulinum Toxins ; Botulinum Toxins, Type A ; Knee ; Osteoarthritis ; Stress, Psychological