Autoimmune encephalitis: differences of clinical features between antibody-positive and antibody-negative conditions

Autoimmune encephalitis is a kind of immune disease with a higher incidence while classication and analysis based upon different states of antibodies are scarcely reported. We intend to compare the clinical features of autoimmune encephalitis between antibody-positive and antibody-negative, with a goal of contributing to early diagnosis and facilitate clinical treatment decisions, especially concerning antibody negative cases. A total of 38 patients with autoimmune encephalitis were enrolled and patients with infections, hereditary, metabolic or toxic neurological diseases were excluded. Based on the serum and cerebrospinal uid (CSF) antibody results, all patients were divided into antibody-positive group and antibody-negative group. The clinical manifestations of the two groups were compared in detail. The number of antibody-positive group with immuno-related injury (p=0.012), number of combined symptoms (p=0.027) and abnormal secretion of exocrine glands (p=0.023) were higher than antibody-negative group; patients with psychiatry department treatment history exceeded antibody-negative group (p=0.014) and patients with epileptic of antibody-positive group was less than that of antibody-negative group (p=0.028). Majority of patients in antibody-positive group displayed cerebrospinal uid (CSF) pressure and white blood cell (WBC) count increased while that in a small number of antibody-negative group (p = 0.043, p=0.045). Baseline mRS scores of antibody-positive group were higher than negative group at admission (p=0.049), and mRS in both groups were signicantly lower at discharge than admission (p=0.010, p =0.011). Compared with antibody-negative group, the combined therapy in antibody-positive group was more common (p=0.025).Although antibody-positive patients may have more serious conditions,but the data suggest that in spite of some differences between antibody positive and negative patients, they are quite similar and perhaps should be subjected to the same rigorous treatment or management protocols. Immunotherapy might play an important role in relieving symptoms.

group; patients with psychiatry department treatment history exceeded antibody-negative group (p=0.014) and patients with epileptic of antibody-positive group was less than that of antibody-negative group (p=0.028). Majority of patients in antibody-positive group displayed cerebrospinal uid (CSF) pressure and white blood cell (WBC) count increased while that in a small number of antibody-negative group (p = 0.043, p=0.045). Baseline mRS scores of antibody-positive group were higher than negative group at admission (p=0.049), and mRS in both groups were signi cantly lower at discharge than admission (p=0.010, p =0.011). Compared with antibody-negative group, the combined therapy in antibody-positive group was more common (p=0.025).Although antibody-positive patients may have more serious conditions,but the data suggest that in spite of some differences between antibody positive and negative patients, they are quite similar and perhaps should be subjected to the same rigorous treatment or management protocols. Immunotherapy might play an important role in relieving symptoms.

Background
Autoimmune encephalitis (AE), a group of autoantibody-mediated neuronal cell destruction disorders, has a diverse manifestations with acutely or subacute onset, accompanied by neuropsychiatric behavior, memory loss, seizures, sometimes decreased levels of consciousness, abnormal exercise and autonomic dysfunction [1]. In addition, some patients have potential risk for mergers neoplasm [2]. It can be happened in all age and accounts for a high proportion in non-infectious encephalitis [3]. The researchers have identi ed that AE has a high disability rate, and may have a fatal outcome without early immunotherapy [4].
We found in clinical practice that antibody-negative patients are common and its pathogenesis is unclear [14]. Probably because antibody-negative AE have only been identi ed in the last few years. As far as we know, there is currently few comparative study of AE with the different states of antibodies. Further understanding the different antibody states of AE on the clinical features with differences and similarities may be bene cial to early clinical decisions. To have a better understanding of AE, we will preliminarily compare the clinical features among antibody-positive and antibody-negative patients.

Patient selection
The term AE or possible AE patients were searched in the digital archive of discharge reports of Qianfoshan hospital between 2016 and 2018. The clinical features are consistent with the latest version of the diagnostic criteria for AE published by Lancet Neurol in 2016 [2]. If anti-neuronal surface or synaptic antibody positive, and exclude other possible causes, antibody-positive AE can be directly diagnosed.
Antibody-negative AE can be made when the following conditions are met: 1.Fast progression within 3 months of recent memory de cits, altered state of consciousness, or psychiatric symptoms. had an incomplete clinical data were also excluded. All patients CSF and serum samples were sent to Jinan Golden Medical Testing Center to detect the antibodies of central nervous system AE(detection types include NMDA-R-Ab, AMPA1-R-Ab, AMPA2-R-Ab, GABAB -R-Ab, LGI1-Ab, CASPR2-Ab, GAD65-Ab) and para-neoplastic syndrome autoantibodies were excluded from paraneoplastic diseases or sent to Tianjin Huada Genetic Testing Institute for high-throughput genetic testing of infectious agents in order to rule out infectious diseases. All patients, according to the serum and CSF antibody results, were divided into antibody-positive group (22 cases) and antibody-negative group (16 cases). And the antibody-positive group was included 4 subgroups according to the type of antibodies including NMDAR (15cases), LGI1 (4cases), GABAB (2cases), AMPA1 (1cases). The patients' medical information were obtained from the inpatient history, as well as the descriptions of his or her attending physicians and family members. All patients received immunotherapy after completing laboratory tests during hospitalization, and the immunotherapy effect was evaluated according to Modi ed Rankin Scale (mRS) score which were evaluated on admission and discharge respectively. Ethical approval for this study was from the Qianfoshan Hospital Ethical Committee. Informed consent comes from the patients or their guardians.
Statistical analysis: Statistical software using SPSS20.0. The Student's T test and the Fisher's exact test were used for analysis the differences between variables, then, the results of test are reported accordingly.
Statistical differences were considered meaningful when P < 0.05.

Comparison of clinical manifestations
There are no differences of the gender and age in two groups. And no differences were seen for the number of patients in intensive care unit (mainly due to unsatisfactory seizure control or persistent hypoventilation disorder, p=0.675) or the days of hospitalization between the two groups (P=0.102).
Patients with immune-related injury history that including hypothyroidism, hyperthyroidism, rheumatoid arthritis and type I diabetes, and psychiatry department history of treatment in antibody-positive group signi cantly exceeded antibody-negative group (p=0.012, p=0.014).The basic symptom types of the two groups were basically same, but the number of multiple types of symptoms were higher in antibodypositive group(P=0.027). Furthermore, patients with seizure in antibody-negative group signi cantly exceeded antibody-positive group (p =0.028) and abnormal secretion of exocrine glands, including hypersecretion of salivary and sweat glands, was signi cantly less than antibody-positive group (p=0.023). Demographics and clinical data for both groups are shown in Table 1.

CSF analysis, EEG and MRI studies
The pressure and white blood cell (WBC) count of CSF were found to be increased in both groups, which was more common in antibody-positive group. The abnormal value of CSF pressure is from 200 mmH2O to 250 mmH2O, and of WBC count is 10 to 50/mm3. The CSF-High-throughput seuencing were no abnormalities found in both groups, which could exclude the presence of infection. The incidence rates of Electroencephalography (EEG) abnormal were equivalent (p=0.36) in two groups, mainly expressed as low-amplitude background rhythm. Abnormal hyperintensity signal of T2/Flair was found in head MRI which was consistent with the manifestation of encephalitis. No differences were seen of multiple MRI abnormalities between the two groups, with temporal lobe, insular lobe and hippocampus being the most common. Table2 shows the general analysis.

Comparison of neurological status and immunotherapy
The two groups were treated with different immunotherapy methods , the positive group would be more likely received combined therapy (corticosteroids and intravenous immunoglobulins(IVIG) )and the negative group more likely received monotherapy(corticosteroids or IVIG). Both groups have a dramatically decline of mRS after immunotherapy (Table4, 5). Baseline mRS scores of antibody-positive group were not signi cantly different from that of the antibody-negative group at discharge, but at admission were signi cantly upper than those in antibody-negative group (p=0.220, p=0.049, respectively) (Table3, 4).

Discussion
AE is divided into two categories of antibody-positive AE and antibody-negative AE by antibody testing that has become an increasingly recognized [15]. However, only a few studies have compared the clinical features or outcomes in AE patients with the different states of antibodies. Our study showed that the symptoms of antibody-positive patients are relatively severer and more complicated than antibodynegative patients, but both can bene t from immunotherapy.

Patients with different states of antibodies have very similar clinical characteristics [16]
, and also showed similar in auxiliary inspection ndings [17]. However, clinicians sometimes pay insu cient attention to antibody-negative and consider that it uncorrelated to AE, leading to misdiagnosis or delay diagnosis. Clinicians often found both types AE may have a signi cant improvement in the early stages of after immunotherapy and showed similar short-term prognosis and neurological dysfunction, which also be con rmed by Yael Hacohen [14]. Because in our study that the patients with positive antibody had severer neurological symptoms at admission and more likely received combined therapy , which is superior to monotherapy previous literature reports [18]. Therefore, it does not indicate which types of patients are more sensitive to immunotherapy, but can be explained that the immune response is closely related to both types of AE. Our study also con rmed that AE is an immune-mediated disease.
Interestingly, exacting comparison of clinical features still reveal differences. Our study showed that antibody-positive AE are more likely to have medical history of immune-related injuries, and studies on other types of immune diseases have also been found a similar phenomenon that the presence of antibody in the body often accompanied with other immunity disease [19][20]. Since antibody-positive patients more likely to have medical history of immune-related injury, multiple immune injuries coexist will inevitably lead to the poorer of their immunologic function, therefore the distinction between antibody-positive and antibody-negative has important clinical signi cance. On the other hand, as showed in our study, compared with antibody-negative patients, antibody-positive patients were often accompanied by abnormal secretion of exocrine glands. Immune injury is one of the main causes of abnormal secretion of exocrine glands, which is widely recognized in present study [21]. Considering the potential relationship between antibody status and immune injury, we speculate that the body suffer immune injury might easier to induce the occurrence of other immune diseases, those conditions are existed in antibody-positive AE which was reported by Ohkawa T [22][23], but not in antibody-negative AE, although further investigations are needed. Furthermore, the state of antibodies may re ect the severity of immune damage, as found by others that the antibody level could be reduced as the improvement of symptoms [10]. As expected, it can also be fully validated in our study that patients with antibody-positive patients have more clinical symptoms, more abnormalities in pressure and WBC count of CSF. Further prospective studies are necessary to con rm this hypothesis.
In addition, it is not clear why antibody negative patients are prone to suffer epileptic while the mechanism of immune damage cannot be completely explained reasonably. However, other researchers found that epileptic which is caused by AE can have better responses to immunotherapy than anti-seizure medication [24]. Still some unexplored causes are existed, and the further investigations of patients with antibody negative may lead to discover of new pathogenic mechanisms. And more fundamental experimental researches are necessary in the future.

Conclusions
Antibody-negative patients are not rare and its clinical features are similar to those of antibody-positive patients. A few differences may be related to the degree of immune damage. Immunotherapy might play an important role in relieving symptoms. The pathogen and mechanism of the disease should be further investigated in the future. University. All patients or the patient's next-of-kin provided written informed consent to participate if a patient could not sign due to disability. And this was also approved by the ethics committee. A copy of the written consent is available for review by the Editor of this journal.

Consent for publication
Written informed consent was obtained from the patients for publication of this research and any accompanying images or from the patient's next-of-kin if a patient could not sign informed consents due to disability.
Availability of data and materials The datasets used and/or analysed during the current study are available from the corresponding author on reasonable request.

Con icts of interest
There are no con icts of interest. Funding: Data collection, analysis, and interpretation of the study was supported by grants from the Natural Science Foundation of Shandong Province (No. ZR2016HP23) and the Science and Technology Develop Project of Jinan (No. 201704113).

Authors' contributions
Tingting Li collected and analyzed data, and drafted the manuscript. Zhihua Si and Lu Lu participated in the acquisition and interpretation of data, and were involved in revising the manuscript. Aihua Wang conceived of the study, and helped to draft and revise the manuscript. All authors read and approved the nal manuscript, and agreed to be accountable for all aspects of the work.  Table4. Comparison of immunotherapy method and change of mRS mean