myasthenia gravis acetylcholine
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Key words receptor
Key words: receptors, cholinergic; antibody; myasthenia gravis
Abstract: Objective severe muscle (myasthenia gravis, MG) in patients with acetylcholine receptor antibody (AChRab) and monkey central nervous system nicotinic acetylcholine receptor (nerve-nAChR) binding between the immune responses. Approach from AChRab positive MG extraction and purification of the blood of patients AChRab, then immunohistochemistry of AChRab and monkey CNS nerve-nAChR binding between the immune response. The results for the first time that AChRab with monkey CNS immune cross between nerve-nAChR binding, with the positive immune reaction is widely distributed in many parts of the monkey CNS, such as the cerebral cortex ( , , layer), hippocampus cortex, basal ganglia, hypothalamus, brainstem motor nuclei, brainstem reticular formation, spinal cord anterior horn motor neurons, cerebellar deep nuclei and cortex cells and other feeding wild willing or strong positive reaction was strong. Conclusion AChRab MG patients with CNS nerve-nAChR cross-monkey combination.
Immunoreactivity of acetylcholine receptor antibody from myasthenia gravis with monkey neuronal nico tinic acetylcholine receptorDepartment of Neurology, Tangdu Hospital, Fourth Military Medical University, Xi''an710038, China
Keywords: receptors, cholinergic; antibodies; myasthenia gravis
Abstract: AIM To explore the immunoreactivity of acetyl-cholinereceptor antibody from myasthenia gravis and monkey central neuronal nicotinic acetylcholine receptor.METHODS Immunoreactivity of acetylcholine receptor antibody puri-fied from myasthenia gravis serum and monkey central neu-ronal nicotinic acetylcholine receptor was investigated by immunocytochemical method . RESULTS Acetylcholine re-ceptor antibody crossreacted with monkey central neuronal nicotinic acetylcholin receptor.The strong positive im-munoreaction was widely distributed among monkey central nervous system, for instance, cerebral cortex (layer , , ), hippocampus, basal ganglia, hypothalamus , brain stem motor nucleus, reticular formation of brain stem, motor neu-ron in anterior horn of spinal cord, deep nucleus in cerebel-lum and Purkinje cell in cerebellum cortex.CONCLUSION Acetylcholine receptor antibody from myasthenia gravis can crossreact with monkey central neuronal nicotinic acetyl-choline receptor.Myasthenia gravis on the current international (myasthenia gravis, MG) in the presence of serum and cerebrospinal fluid of AChRab can cause central nervous system (CNS) injury is controversial In 1973 with the muscle - acetylcholine receptor (muscle-nAChR) immunized animals MG made of experimental animal models, and from the blood of MG patients detected AChRab, that the MG is limited at the neuromuscular junction autoimmune disease. However, studies have shown that MG in addition to the current weakness, but also associated with CNS injury performance and neurophysiological abnormalities [1,2]. we found that the IgG MG can cause central nervous system dysfunction in rats [3-6]. The study by immunoaffinity chromatography from blood of patients with MG extraction and purification of IgG, and then use it to discuss with the monkey CNS nerve-nAChR binding between the immune response.1.1 MG material according to the typical clinical symptoms, positive anti-cholinesterase drugs test and the results of serum titration AChRab diagnostic MG. Were taken of the MG and healthy human blood 10mL, centrifugation in serum frozen for use.1.2.1 ammonium sulfate precipitation and immunoaffinity chromatography, the MG patients and healthy human serum diluted 1 10mL times with PBS, home ice bath, magnetic stirrer, add an equal volume of cold saturated ammonium sulfate, 4 under stirring 1.5 ~ 2h. centrifuged 15min, 5000r min-1, to collect sediment, discard the supernatant. add PBS to the original times the serum volume, red soluble precipitate, centrifuged 15min, centrifugal force of 1200g, the supernatant discarded. with 330g L-1 of the original times the volume of saturated ammonium sulfate precipitate washed solution, 4 C and centrifuged 15min, centrifugal force of 1200g, Shen fluid collection. Repeat 2 to 3 times. will collect the dialysis bag into the immunoglobulin solution, placed in PBS (1 1000) of dialysis, 4 overnight, collected subpackage. Protein G affinity purified IgG: column installed according to the instructions, the use of 0.02mol L-1, pH7.3 the PBS80mL analysis column. would have been crude The MG and healthy human immunoglobulin diluted with PBS1 10, joined the column, the non-IgG components from the column bottom out, IgG component (AChRab) and Protein G chromatography column with left. tomography washed first with PBS column 3, after the use of 1mol L-1, pH2.7 glycine dissolved part of the night IgG elution column and collected immediately after adjustment of pH to 7.3, 4 with under the PBS (1 1000) dialysis, overnight. collected and frozen memory IgG, or directly used for experiments.
1.2.2 Immunohistochemical selection of adult monkeys 2 and body mass were 6.9,7.1 kg.1g L-1 sodium pentobarbital 70mL (25mg kg-1) after anesthesia, the left heart infusion 2000mL saline wash, 40g L-1 and then cold (4 ) paraformaldehyde solution 5000mL perfusion fixation 20 ~ 30min, quickly the brain, into 200g L-1 sucrose 1d. frozen sections, thickness 40 m, into the 3g L-1 TritonX-100, 30min, with KPBS (10mmol L-1) wash 15min 3 times, and then at 4 respectively AChRab (KPBS1 100 dilution) and IgG healthy slice incubated 48h, KPBS washing 15min 3 times; then biotinylated sheep anti-human-IgG (Wuhan Boster Company, 1 100 dilution) slices were incubated at room temperature 4h, KPBS washing 15min 3 times; Finally, ABC (Sig-ma, 1 500 dilution) were incubated at room temperature slices 3h, KPBS washing 15min 3 times. with dimethyl benzidine - nickel ammonium sulfate Enhancement Act coloring, timely terminate the reaction.Immunohistochemistry showed that, AChRab and nerve-nAChR binding reaction between the diffuse positive immunoreactivity distributed in many parts of the monkey CNS, such as the cerebral cortex ( , , layer), hippocampus cortex, basal ganglia, hypothalamus, brain stem Sports nuclei, brainstem reticular formation, spinal cord anterior horn motor neurons, deep cerebellar nuclei and cortex, such as feeding cells were willing to field a strong positive reaction; and other parts of the hypothalamus were moderately positive (Tab1). nerve-nAChR-like positive immune response concentrated on the membrane of neurons and their dendrites connected by light microscopy can clearly see the outline of the membrane cortex pyramidal cells and long dendrites extending toward the top of the cortical surface (Fig1).3 Discussion
Muscle and nerve-nAChR structure and function comparison. ACh and other agonists with nerve-nAChR -subunit activated Ca2 channels open; receptor antagonist -BuTx with the nerve-AChR 9 form the nerve -nAChR binding, blocking ion channels. Studies show that muscle-nAChR, and both nerve-nAChR are ligand - gated - ion channel gene family, homologous to the cell membrane polypeptide 80% to 90% , and human and rat muscle and nerve-nAChR and -subunit homology between the peptide chain of up to 90% . nerve-nAChR by , subunit composition of the two polymer [9 ], sub-units are based on their genetic code, divided into several subtypes. by immunoaffinity chromatography, from a variety of extraction and purification of vertebrate CNS, the nerve-nAChR-like protein . of nerve - nAChR and subunit subtypes found that the nerve-nAChR subtype
9 and 4 subtypes in many parts of the vertebrate CNS , such as the spinal cord, brain stem, hypothalamus, basal ganglia, hypothalamus, cortex, cerebellum other. AChRab and CNS nerve-nAChR immune cross combination. study found that some of MG associated with CNS damage, such as some MG patients have abnormal nerve electrophysiological testing [1,2]; the MG patients with IgG injected into the caudate putamen, a seizure like discharge. Our previous studies have shown , injected into the rat ventricular system AChRab can cause CNS dysfunction in SD rats. AChRab this study show that CNS nerve with monkey immune cross-nAChR binding, suggesting that patients with MG with CNS dysfunction and neurophysiological mechanisms and AChRab test abnormalities and neural-nAChR binding.
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 Li ZY, Ju G, Qiu XF, You GX.Effect of acetylcholine receptor antibodies derived from myasthenia gravis on the rat neuronal nicotinic acetylcholine receptor [J]. Zhonghua Neike Zazhi (Chin Intern Med J), 1997; 36 (1 ) :15-1Tang Du Hospital, Fourth Military Medical University, Department of Neurology, Xi''an 710038, ChinaLi Zhu A (1962 -), male (Korean), Antu County, Jilin Province. PhD, Associate Professor. Tel. (029) 3577443 Email.lizhuyi @ fmmu.edu.cn
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