[Establishment and consistency verification of the standard operation procedure for laboratory detection of immunoglobulin G oligoclonal bands in cerebrospinal fluid].

Objective: To establish the standard operation procedure (SOP) for detection of oligoclonal band (OCB) in cerebrospinal fluid (CSF) and verify consistency by using this SOP in different laboratories. Methods: The SOP for detection of CSF-OCB fluid was successfully established by an expert feedback approach. Neuroimmunology laboratories in 3 representative Chinese three-tier research hospitals were selected for this study, and commercially available protein electrophoresis automation systems and detection SOP were set up. The quality control product was provided by the College of American Pathologists (CAP) and Sebiacompany were used for interior quality and compared to each other, respectively. Seventeen serum and CSF paired samples were tested and compared using the same SOP. Kappa test or Kendall W test were adopted to evaluate the inter-laboratory consistency among different hospitals. Results: The results of repeated testings in a single hospital suggested that the 2-and 4-fold dilution for CSF-OCB were reported as positive, while 64-and 128-fold dilution were reported as negative. Positive or negative inconsistencies were reported in 8-, 16-, and 32-fold dilution. After increasing the number of repetitions, the results showed that both 16-and 32-fold dilution were reported as negative, and 8-fold dilution exhibited negative results (2 positive results for 40 repetitions, coincidence rate=95%).The results of multi-center inter-laboratory quality assessment showed that the detection consistency rate among 3 hospitals was 100% (Kappa value =1). Conclusions: The SOP to detect CSF-OCB established in this study demonstrates a good repeatability and stability. Therefore,such SOP would be a good reference for diagnostic laboratories to detect CSF-OCB in China.

[Correlation study on anti-Ro52 antibodies frequently co-occur with other myositis-specific and myositis-associated autoantibodies].

OBJECTIVE: Anti-Ro52 antibodies are frequently co-occur with other myositis-specific and myositis-associated autoantibodies, we here to study this phenomenon in Chinese patients suspected with inflammatory myopathies. METHODS: In the study, 1 509 patients clinically suspected with inflammatory myopathies were tested for 11 kinds of myositis-specific and myositis-associated autoantibodies (including: anti-Jo-1, PL-7, PL-12, EJ, OJ, Mi-2, SRP, Ku, PM-Scl 75, PM-Scl 100, and Ro52 antibo-dies) by line-blot immunoassay from 2010 to 2016 in Peking University First Hospital. This retrospective study was to analyze these results to reveal the characteristics of anti-Ro52 antibodies co-occuring with other myositis autoantibodies. The data were analyzed using SPSS 17.0 and Graph Pad PRISM for Chi-square test, independent t-test, Pearson's correlation analysis, and drawing statistical graphs. Significance level was set at P < 0.05. RESULTS: The positive rate of anti-Ro52 antibodies was 18.3% (276/1 509 cases), which was the most frequently detected myositis antibodies in our center. 51.8% (143/276) of the patients with anti-Ro52 antibodies were combined with the other myositis antibodies, and the most common co-occurred antibodies were anti-SRP antibodies (18.8%, 52/276), and the second common co-occurred antibodies were anti-Jo-1 antibodies (13.0%, 36/276). Anti-Ro52 antibodies were the most common antibodies that co-occurred in other myositis antibodies positive patients except in anti-OJ antibodies positive group. The co-positive rate with anti-Ro52 antibodies was the lowest in anti-PM-Scl 75 positive group (30.4%, 31/102), and the highest in anti-EJ positive group (80.0%, 12/15). The positive rate of anti-Ro52 antibodies in anti-synthase antibodies (including anti-Jo-1, EJ, OJ, PL-7, and PL-12 antibodies) positive group was 57.3% (75/131), which was significantly higher than that in the other antibodies (including: anti-Mi-2, SRP, Ku, PM-Scl 75, and PM-Scl 100 antibodies) positive group with 35.2% (119/338) (χ2=18.916, P < 0.001). The intensity of anti-Jo-1, EJ, and SRP antibodies in the group of the patients that co-occurred with anti-Ro52 antibodies was significantly higher than that in the other group without anti-Ro52 antibodies respectively (P < 0.05). The intensity of anti-SRP antibodies was significantly correlated with that of anti-Ro52 antibodies (r=0.44, P=0.001). CONCLUSION: Anti-Ro52 antibodies were commonly associated with other myositis-specific and myositis-associated autoantibodies, especially with anti-synthase antibodies, and the co-presence of anti-Ro52 antibodies may be correlated with the myositis antibody intensity.

[Distribution characteristics and correlation analysis of antibody detection value in myasthenia gravis].

Objective: To determine the factors affecting distribution and magnitude of antibody detection value in myasthenia gravis (MG). Methods: A total of 406 MG patients diagnosed at Department of Neurology, Peking University First Hospital from May 2015 to November 2017 were included.All of them exhibited muscle fatigue with decreased response in repetitive nerve stimulation test. There were 200 males and 206 females whose ages ranged from 2 to 85 years old. According to clinical classification of MG recommended by Myasthenia Gravis Foundation of America (MGFA), patients assigned to class I to class V included 200,140, 46, 15 and 5 cases, respectively. There were 33 cases of thymic hyperplasia and 63 cases of thymoma confirmed by radiological or pathological findings. Quantile plots and quantile regression model were used to determine the effects of age, gender and MGFA classification, thymus disease on acetylcholine receptors (AChR)antibody, acetylcholinesterase (AChE) antibody, Titin antibody, ryanodine receptor (RyR) antibody and muscle-specific tyrosine kinase (MuSK) antibody detection values detected by enzyme-linked immunosorbent assay (ELISA). Results: MGFA classification had effects on distribution of AChR antibody level. There was a positive correlation between age and AChR antibody level(P<0.05). Negative correlation was found between age and AChE, Titin and RyR antibody level (P<0.05). No significant correlation was shown between any factors and MuSK antibody level(P≥0.05). MGFA classification had a positive correlation with AChR antibody level (P<0.05) and no correlation with other antibody levels (P>0.05). Gender and thymus disease had no correlation with any tested antibody levels (P>0.05). Conclusion: MGFA classification has significant effects on distribution of AChR antibody level. Age and MGFA classification have positive correlation with AChR antibody level.

Multiple antibody detection in 'seronegative' myasthenia gravis patients.

BACKGROUND AND PURPOSE: Myasthenia gravis (MG) is an autoimmune disease caused by antibody mediated impairment in the neuromuscular junction. Seronegative MG (SNMG) without antibodies against acetylcholine receptor (AChR) and muscle-specific kinase (MuSK) by routine assays accounts for about 20% of all MG patients. METHODS: Plasma from 81 Chinese MG patients previously found to be seronegative was tested by routine assays for AChR and MuSK antibodies. These samples were screened by (i) a novel, highly sensitive radioimmunoassay for AChR antibodies; (ii) cell-based assays for clustered AChR, MuSK and lipoprotein receptor-related protein 4 (LRP4) antibodies; (iii) a radioimmunoassay for titin antibodies. RESULTS: Antibodies to AChR, MuSK, LRP4 and titin were found in 25% (20/81), 4% (3/81), 7% (6/81) and 6% (5/78) of SNMG patients, respectively. In total, 37% of SNMG patients were found to be positive for at least one of the tested antibodies. AChR antibody positive patients had more severe disease (P = 0.008) and a trend towards fewer remissions/minimal manifestations than AChR antibody negative patients. The four patients with coexistence of antibodies had more severe disease, whilst the seronegative patients had milder MG (P = 0.015). CONCLUSIONS: Detection of multiple muscle antibodies by more sensitive assays provides additional information in diagnosing and subgrouping of MG and may guide MG treatment.

[An analysis and literature review of two cases of autoimmune encephalitis with GABAB receptor antibodies].

Autoimmune encephalitis with GABAB receptor antibodies has been rarely reported. Two cases of GABAB receptor antibodies encephalitis were presented here.Epilepsy was the onset symptom, followed by declined consciousness and frequent seizures. Fever was presented in the whole course of the disease. Myorhythmia of the two hands and pilomotor seizures were shown in the later course of the disease. No specificity was demonstrated in electroencephalograms and magnetic resonance imaging. Sensitive response was shown to the first-line immunotherapy.

[Clinical and myopathological features of Jo-1 syndrome].

OBJECTIVE: To report the clinical and myopathological features of 16 patients with Jo-1 syndrome. METHODS: Sixteen patients were recruited in this study, who were diagnosed as Jo-1 syndrome in Department of Neurology of Peking University First Hospital from January, 2011 to July, 2015. The clinical data and myopathological data were analyzed. RESULTS: The mean onset age was 41±14 (21-68) years old. 87.5% was female. The median duration was 9.5 months (1-192 months). The main clinical manifestations were weakness in 13 cases (81.2%), arthritis in 10 cases (62.5%), interstitial lung diseases in 8 cases (50%), dermatomyositis-like skin lesions in 5 cases (31.2%), fever in 3 cases (18.8%), Raynaud's phenomenon in 2 cases (12.5%) and mechanic's hands in 2 cases (12.5%). There were 3 cases with other connective tissue diseases and 1 case with non-Hodgkin's lymphoma. Mean serum CK was 3 054±2 058(470-5 222) U/L. All patients had anti-Jo-1antibody, combined with anti- Mi-2 antibody in 1 case, anti-Ro-52 antibody in 5 cases, and anti-nuclear antibody in 5 cases. 4/5 cases showed myopathic changes for electromyography (EMG) tests. Myopathological changes included edema, fragmentation and inflammatory infiltration in perimysium in 14 cases (87.5%), muscle atrophy in 13 cases including 7 cases(43.8%) predominantly in perifascicular field. Muscle fiber necrosis appeared in 8 cases with predominantly in perifascicular area in 4 cases (25%). Muscle fiber regeneration occurred in 11 cases with predominantly in perifascicular field in 5 cases (31.2%). CD8 positive T-lymphocytes, CD20 positive B-lymphocytes and CD68 positive macrophages infiltrated in various degrees, most of which were located in perimysium. MHC-Ⅰ were expressed on muscle fiber membranes in different degrees, including 7 cases (43.8%) predominantly in the cytoplasm of perifascicular muscle fibers. C5b-9 deposited in perifascicular muscle fiber membranes in 7 cases (43.8%) and perifascicular capillaries in 2 cases (12.5%). CONCLUSIONS: The main manifestations of this group of Jo-1 syndrome are weakness, arthritis and interstitial lung diseases, and dermatomyositis-like skin lesions, fever, Raynaud's phenomenon, and mechanic's hands can also be seen. Edema, fragmentation and inflammatory infiltration in perimysium are common. Pathological changes in perifascicular fields appear in some cases.

Level of soluble CD30 after kidney transplantation correlates with acute rejection episodes.

Measurement of soluble CD30 (sCD30) levels may predict acute rejection episodes (ARE). To explore the value of sCD30 after transplantation, we tested serum sCD30 levels in 58 kidney transplant cases at 1 day before and 7 and 28 days after transplantation by enzyme-linked immunosorbent assay (ELISA). The incidences of ARE after kidney transplantation were recorded simultaneously. Meanwhile, 31 healthy individuals were selected as a control group. The results showed a relationship between sCD30 level in serum before kidney transplantation and the incidence of ARE. However, the relationship was more significant between serum sCD30 levels at day 7 after kidney transplantation and the incidence of ARE. There was no obvious relationship between serum sCD30 levels at day 28 after kidney transplantation and the incidence of ARE. These results suggested that the level of sCD30 at day 7 posttransplantation provides valuable data to predict ARE.

Guillain-Barre syndrome and Campylobacter jejuni infection. A study on the etiological characteristics of Guillain-Barre syndrome in China.

Fecal culture for Campylobacter jejuni with the method of Skirrow and serum class specific antibodies (IgG, IgM, IgA) against Campylobacter jejuni using a solid phase enzyme linked immunosorbent assay were performed in 17 cases of Guillain-Barre Syndrome, 17 other neurological disease controls, and 33 normal controls. The results revealed: 1. the incidence of Campylobacter jejuni infection, especially the recent one in Guillain-Barre Syndrome was much higher than in the other two groups; 2. the recent incidence of infection in those below 30 years old and those within the period of summer and autumn was higher than that above 30 years old and that in the other seasons. The results suggested that Campylobacter jejuni infection might be one of the important precipitating factors of Guillain-Barre syndrome and might play an important role in the epidemiological pattern of Guillain-Barre Syndrome in China.

[Five cases of Campylobacter jejuni enteritis accompanied with Guillain-Barré syndrome].

17 cases of Guillain-Barré syndrome (GBS) were admitted to The First Hospital of Beijing Medical University and Baoding Hospital from July, 1991 to October, 1992. Five patients had diarrhea before the onset of GBS and showed positive reaction for serum IgM antibodies to Campylobacter jejuni. They were considered to have Campylobacter jejuni enteritis as well. This incidence of Campylobacter jejuni enteritis in GBS patients was higher than in those with other neurological diseases and in normal controls. These five patients were all from countryside and had their onset in the summer and autumn. Moreover, four of them were below twenty-four years old. The epidemiological pattern was quite similar to that of Campylobacter jejuni enteritis in China. Our data showed the close association between GBS and Campylobacter jejuni enteritis and the possible mechanism was discussed.

Mutations in the leucine zipper-like heptad repeat sequence of human immunodeficiency virus type 1 gp41 dominantly interfere with wild-type virus infectivity.

It has been previously shown that a proline substitution for any of the conserved leucine or isoleucine residues located in the leucine zipper-like heptad repeat sequence of human immunodeficiency virus type 1 (HIV-1) gp41 renders viruses noninfectious and envelope (Env) protein unable to mediate membrane fusion (S. S.-L. Chen, C.-N. Lee, W.-R. Lee, K. McIntosh, and T.-M. Lee, J. Virol. 67:3615-3619, 1993; S. S.-L. Chen, J. Virol. 68:2002-2010, 1994). To understand whether these variants could act as trans-dominant inhibitory mutants, the ability of these mutants to inhibit wild-type (wt) virus infectivity was examined. Comparable amounts of cell- and virion-associated gag gene products as well as virion-associated gp41 were found in transfection with wt or mutant HIV-1 provirus. Viruses obtained from coexpression of wt provirus with mutant 566 or 580 provirus inhibited more potently the production of infectious virus than did viruses generated from cotransfection of wt provirus with other mutant proviruses. Nevertheless, all viruses produced from mixed transfection showed decreased infectivity compared with that of the wt virus when a multinuclear-activation beta-galactosidase induction assay was performed. The ability of wt Env to induce cytopathic effects was inhibited by coexpression with mutant Env. Coexpression of mutants inhibited the ability of the wt protein to mediate virus-to-cell transmission, as demonstrated by an env trans-complementation assay with a defective HIV-1 proviral vector. These observations indicated that mutant Env, per se, interferes with wt Env function. Moreover, cotransfection of wt and mutant proviruses produced amounts of cell- and virion-associated gag gene products comparable to those produced by transfection of wt provirus. Similar amounts of gp41 were also found in virions generated from wt-mutant cotransfection as well as from wt transfection alone. These results indicated that the inhibitory effect conferred by mutants on the wt virus infectivity does not involve the late steps of Gag protein assembly and budding, but they suggest that the wt and mutant Env proteins form a dysfunctional hetero-oligomer which is impaired in an early step of the virus replication cycle. Our study demonstrates that mutations in the HIV-1 gp41 leucine zipper-like heptad repeat sequence dominantly inhibit infectious virus production.