Guillain-Barré Syndrome: Retrospective Analysis Of Patients In Southern China & Mechanism Research Of Wnt/?-catentin Signaling Pathway In Its Rat Model

BrackgroudGuillain-Barrésyndrome?GBS?is an immune-mediated peripheral neuropathy that is an important cause of acute neuromuscular paralysis.GBS in northern China has been widely concerned,"AMAN is the main subtype of GBS in China"is the consensus of foreign scholars,the disorder was initially called Chinese paralytic syndrome.However,there has been controversy about the main stubybpe of GBS in China.For China has a vast territory,are there any differences in clinical characteristics between the southern and northern GBS patients?In addition,intravenous immunoglobulin?IVIG?and plasma exchange are the main treatments for GBS.Most of the patients have good prognosis.However,despite receiving immunotherapy,up to 25%of the patients still suffer from respiratory insufficiency,20%of the patients are severely disabled and nearly 5%of the patients die.Therefore,accurate diagnosis in the early stage of the disease is helpful for timely treatment and prevention of complications;in-depth study of its pathogenesis,open up new ways to improve the clinical outcomes of GBS patients is of great significance.Objective?1?To analyze the clinical features and validation of Brighton criteria in GBS patients from southern China.?2?This study was designed to exmine the dynamic expression of Wnt/?-catentin signaling pathway in experimental autoimmune neuritis rats?EAN?,in order to explore its possible mechanism and to provide a new therapeutic targets for the treatment of GBS.Methods?1?The clinical data of hospitalized GBS patients from 69 hospitals of 14 provinces/cities in southern China,i.e.the area south of the Huaihe River, between 1 Janurary 2013 and 30 September 2016,were collected and analyzed retrospectively,and patients were classified according to the Brighton criteria of case definition,ranging from a highest?defined as level one?to a lowest?level four?level of diagnostic certainty.?2?28 male Lewis rats weighing 170±10 g were randomly divided into control group?n=7?and EAN group?n=21?.EAN group included the early group?n=7?,the peak group?n=7?and the recovery group?n=7?.EAN rats was immunized with P2_57-81 peptide;weighed daily and scored nervous system function for 30 days;the sciatic nerve was taken on the days 10,17,and 30 p.i.for H&E staining,transmission electron microscopy and immunohistochemical staining;blood samples were collected weekly from caudal vein to detect IFN-?,IL-4,TGF-?1;and the sciatic nerve was taken to examinate the dynamics expression of Wnt/?-catenin pathway molecules.Results?1?a)A total of 1 358 GBS patients were collected,including 51 cases with cranial nerve variants,157 Miler-Fisher syndrome and 1 150 classic GBS characterized by flaccid weakness of limbs.b)Among 1 150 cases of classic GBS,49.57% ?570/1 150?patients had antecedent events,with respiratory infection predominated?71.23%,406/570?;83.74%?963/1 150?presented limb weakness at onset,99.21%?1 124/1 133?reached the nadir within four weeks,with a score of 3.15±1.16 for Hughes Disability Scale;99.56%?1 128/1 133?developed bilateral weakness and 95.39%?1 097/1 150?manifested flexia or hyporeflexia at the nadir; the cerebrospinal fluid showed albuminocytologic dissociation in 80.58% ?772/958?patients whose lumbar puncture was performed;demyelinating GBS accounted for 48.14%?401/833?and axonal subtype 18.01%?150/833? respectively in patients with findings of nerve conduction studies available.c) According to Brighton criteria,the patients were stratified as level 1 in 44.09%?507/1 150?,level 2 in 45.74%?526/1 150?,level 3 in 7.57%?87/1 150?and level 4 in 2.61%?30/1 150?of all the patients,and 69.55%?507/729?,28.67% ?209/729?,0%?0/729?and 1.78%?13/729?,respectively in the patients with complete data?n=729?.?2?a)The body weight of EAN rats increased steadily in the first week,then decreased on the on day 10 p.i.and reached the nadir on days 16-18 p.i.;In contrast,rats in the control group showed continuous weight gain during the period.All rats in the EAN group developed typical neurologic signs on day 7 p.i., and reached peak disease on day 17 p.i.;Rats in the control group did not develop any of these symptoms.b)During the early phase,infiltration of inflammatory cells was seen in the sciatic nerve by H&E staining;endoneurium edema,myelin swelling and myelin separation were already present by TEM;CD4⁺T cells were 0.44±0.10 cells per mm².During the peak phase,there was a large amount of inflammatory cell infiltration between the sciatic nerve bundles,endoneurium edema,and myelin swelling;macrophages occasionally seemed to invade the Schwann cell membranes and strip the myelin sheaths and endoneurium edema, myelin swelling and demyelination became more widespread;CD4⁺T cells increased dramatically to a statistically significant and maximal level?3.59±0.33 cells per mm²?.During the recovery phase,inflammatory cell infiltrates reduced; thinly myelinated axons were prominent which infers that remyelination is occurring;CD4⁺T cells were 2.40±0.21 cells per mm²?F=203.9,P<0.0001?. No inflammatory lymphocytes,macrophages endoneurium edema,myelin swelling or demyelination were found in the sciatic nerve sections from the control group.c)The proinflammatory cytokine?IFN-??reached a maximum in the early phase in the sciatic nerves?P<0.0001?;but reached a maximum on day 21 p.i.?P<0.0001?.IL-4 was nearly undetectable during the acute phases and sharply increased during the recovery phase?P<0.0001?;in the serum,IL-4 gradually increased and reached a maximum during the late recovery phase?P<0.0001?.The level of TGF-?1 was similar to that of IL-4.d)GSK-3?mRNA levels were upregulated in the early phase of EAN?early group vs.control group,P<0.05?,with peak levels observed in the recovery phase?P<0.0001?.?-catenin ?P<0.0001?mRNA and LEF-1?P<0.0001?mRNA expression patterns were similar to that of GSK-3?.e)The protein level of GSK-3?was significantly increased in the early phase?early group vs.control group,P<0.05?,then gradually increased in the peak phase,and its high expression persisted through day 30 p.i.?P<0.01?.The total?-catenin level was continuously elevated compared with levels in control rats?P<0.01?,we also noted a continuous increase in nuclear translocation?P<0.01?.The level of LEF-1 was increased in the early phase and its expression remained elevated until the recovery phase?P<0.01?.Conclusion?1?In southern China,demyelinating subtype of GBS is predominated,whereas the proportion of axonal subtype is remarkably lower than that in northern China.The Brighton criteria have a high sensitivity for the diagnosis of GBS in southern China,and examination of cerebrospinal fluid and electrodiagnostic studies are necessary for stratified diagnosis.?2?The expression of IFN-?reached a maximum in the acute phase of EAN,and the level of IL-4 and TGF-?1 were the highest in the EAN recovery phase; Wnt/?-catentin singalling pathway related molecules GSK-3?,?-catenin and LEF-1 were upregulated in EAN rats,it probably achieved through the TGF-? pathway.