High-risk groups of Neonatal Lupus Erythematosus in term infants: a birth cohort study

Purpose: This study aims to analyse the clinical characteristics and risk factors of high-risk groups of neonatal lupus erythematosus (NLE) in term infants. Methods: The high-risk groups of NLE whose mothers were positive of anti-SSA, SSB or U1RNP antibodies during pregnancy were enrolled. They were born from February 2013 to February 2020, with a gestational age not less than 37 weeks. We analyzed their clinical data from birth to 24 months after birth. Results: A total number of 105 cases of NLE high-risk groups were included. Among them, 30 cases were diagnosed with NLE (NLE group) and 75 cases were not (non-NLE group). The affected systems of the NLE group included dermal (13.3%), hepatic (76.0%), and hematological system (43.3%). Hepatic involvement, anemia and thrombocytopenia could not emerge until 60 days, 41 days and 22 days after birth in some cases. The systems involvement could be cured within 3 to 12 months after birth. The clearance time of speci�c autoantibodies was 12 months after birth. There was no signi�cant difference in clinical characteristics of babies and their mothers between the two groups, neither of the positive rate or clearance time of speci�c autoantibodies. Conclusion: After standardized prenatal health care, there is still a high risk of dermal, hepatic, or hematological system involvement for high-risk groups of NLE. There are no speci�c indicators to predict whether the babies will develop to NLE or not. All of them need to be followed up closely within one year after birth.


Introduction
Neonatal Lupus Erythematosus (NLE) is a relatively rare acquired autoimmune disease.It is related to the mother's positive autoantibodies to Sjögren's syndrome types A or B autoantigens (anti-SSA, or SSB), sometimes positive anti-U1 ribonucleoprotein (RNP) antibody, which are transmitted to the fetus through the placenta [1].Most of the mothers suffer from systemic lupus erythematosus (SLE), Sjogren's Syndrome (SS), rheumatoid arthritis, mixed connective tissue disease, or undifferentiated connective tissue disease, while 25-60% of the mothers are asymptomatic and unaware of their diseases [2].In addition, 0.1-1.5% of the healthy pregnant women also have positive autoantibodies mentioned above [3,4].Although NLE is relatively rare, it has a great impact on the health of the fetus and the newborn.
High-risk groups of NLE refer to newborns whose mother has positive NLE-speci c autoantibodies (anti-SSA and/or anti-SSB and/or RNP), including those diagnosed with NLE and those who do not meet the diagnostic criteria.Mothers with autoimmune diseases may have premature delivery due to their diseases.In view of the complex system involvement of premature infants, which are confounding factors of the system involvement of NLE, we only include full-term infants in this study.
More and more high-risk infants of NLE will be born as their mothers' good disease control.This study aims to evaluate the clinical characteristics of high-risk groups of NLE under current medical interventions, and risk factors of developing NLE, so as to guide the diagnosis and treatment of high-risk infants of NLE.

Research objects
Inclusion criteria: Term babies born in our hospital from February 2013 to February 2020 whose mothers had positive NLE-speci c autoantibodies (anti-SSA and/or anti-SSB and/or RNP) were included.The birth gestational age was no less than 37 weeks.
Exclusion criteria: a. Birth gestational age was less than 37 weeks.B. Babies were suffered from other serious chronic diseases.
Diagnostic criteria of the NLE group: The diagnosis of NLE was made when the mother had positive anti-SSA and/or anti-SSB and/or RNP, and fetus or newborn developed atrioventricular heart block, or the newborn had the typical rash (circular erythema or oval erythema) or hepatic (elevated transaminase, cholestasis, liver or spleen enlargement) or hematologic manifestations (anemia, neutrophilia or thrombocytopenia) in the absence of another explanation.
Criteria of non-NLE group: The pregnant mothers were positive for anti-SSA and/or anti-SSB and/or RNP antibodies, but the offspring did not meet the diagnostic criteria of NLE.
Their mothers were 31 ± 4 (22-43) years old, and cesarean sections accounted for 51.4%.The abnormal child-bearing history included: 1 case of labor induction for fetal bradycardia, 2 cases of labor inductions for mothers' poor disease control, 4 cases of embryo damage, 4 cases of spontaneous abortion (excluding biochemical pregnancy), and 1 case of ectopic pregnancy in the NLE group.And 1 case of labor induction for fetal III° atrioventricular block (twice in this case), 1 case of labor induction for fetal heart deformation and atrioventricular block, 4 cases of labor inductions for mothers' poor disease control, 3 cases of fetal death, 11 cases of embryo damage, 1 case of labor induction for fetal abdominal cyst, 1 case of trisomy 21 syndrome delivery, 5 cases of spontaneous abortions (excluding biochemical pregnancy), and 1 case of hydatidiformatitus in the non-NLE group.
There was no signi cant difference in gestational age, birth weight, length, head circumference, mothers' age, delivery mode, abnormal childbearing history, or the usage of prednisone or hydroxychloroquine during pregnancy between the two groups, which was shown in Table 1.The affected systems of the NLE group included dermal (4/30 = 13.3%),hepatic (19/25 = 76.0%), and hematological system (13/30 = 43.3%).The emerging time of typical rash was 16.5 ± 10.9 days after birth.Hepatic involvement, anemia and thrombocytopenia could not emerge until at most 60 days, 41 days and 22 days after birth in some cases.Most of the system involvement could return to normal after observation, but some of them required active intervention, such as the application of topical hormones, liver-protecting drugs, iron supplements, and human immunoglobulin.All of the skin involvement could recover.The hematological system involvement could be cured within 3 months after birth, and hepatic involvement within 12 months.
More details were shown in Table 2.In the NLE group, 29 cases (29/30 = 96.7%)were followed up to a median time of 18 months after birth (3.5/24 months, 1.5-24 months), of which 14 cases were followed up to 24 months after birth.In the non-NLE group, 66 cases (66/75 = 88.0%) were followed up to a median time of 12 months after birth (3.5/24 months, 0.5-24 months), of which 21 cases were followed up to 24 months after birth.There was no signi cant difference in the follow-up time between the two groups (P = 0.347).
In the NLE group, 11 cases monitored their autoantibodies during follow up.Among them, the autoantibodies turned negative in 8 cases within a median time of 10.3 months (9.0/13.0months, 6-15 months) after birth, while in the other 3 cases, the autoantibodies were still positive in 4 months (RNP), 6 months (SSA) and 12 months (SSA) after birth respectively, and they didn't test the autoantibodies anymore within 24 months after birth.In the non-NLE group, 26 cases monitored the autoantibodies.In 22 cases of them, the autoantibodies turned negative within a median time of 12.0 months (9.0/12.3months, 6-23 months) after birth, while 4 cases still had positive autoantibodies in 2-6 months (SSA and RNP) after birth, and they did not re-examine the autoantibodies within 24 months after birth.There was no signi cant difference in the disappearing time of autoantibodies between the two groups (P = 0.615).The disappearing time of autoantibodies were shown in Table 4.

Discussion
Studies have shown that when the mother's NLE-speci c autoantibodies are positive, the risk of giving birth to a child with NLE is 1%-2%.However, the risk will increase to 17%-20% if the previous babies are suffered from NLE [5,10,11,12], but the data are not consistent according to different research, ranging from 25-33% [13].In our study, infants diagnosed with NLE account to 28.6% (30/105) in high risk populations whose mothers are positive for NLE-speci c autoantibodies, which is higher than in previous studies.The rst reason is probably that the diagnostic criteria of NLE are different in different studies.There is no de nite diagnostic criteria for NLE.In order to facilitate registration of NLE, the Research Registry for Neonatal Lupus in the United States only took children with congenital heart block (CHB) or typical skin rash into consideration [14].The lower risks they mentioned mainly refer to the risk of developing CHB, not the other systems involvement.While other researchers took hematological and hepatic systems involvement into account as a part of the diagnostic criteria [5].After standardized prenatal health care, the risk of CHB may decrease dramatically, but there is still a high risk of dermal, hepatic, or hematological system involvement, some of which require active intervention and may increase the risk of NLE of the next child.So we also adopt the diagnostic criteria which includes hematological and hepatic systems involvement in addition to CHB and dermal manifestations, for the purpose of identifying potential NLE babies.The second reason of higher risk of NLE in our study is possibly that mothers have more abnormal child-bearing histories.Up to 40.0% (12/30) of the mothers in the NLE group had abnormal child-bearing histories, such as labor induction for fetal bradycardia, with the possibility of carrying a NLE fetus before and resulting in an increased risk of NLE for the next pregnancy.
NLE can affect the cardiac system which mostly manifest cardiac block, and the other systems such as dermal, hepatic, hematological and sometimes central nervous system [15].Poor prognosis is seen in cardiac block patients [5].No cardiac block babies were born in our study, which might be related to the standardized prenatal health care and intervention in our hospital by good multi-department cooperation.
Besides cardiac system, the most common manifestations are glutamate transpeptidase (GGT) elevation (68%), aspartate aminotransferase (AST) elevation (40%), anemia (30%), and skin rash (13.3%) in the other affected systems, while thrombocytopenia (10%), alanine aminotransferase (ALT) elevation (8%), total bile acid (TBA) elevation (8%) and neutropenia (3.3%) are relatively rare.Previous studies have indicated that dermal system involvement is seen in 4%-16% of anti-Ro (SSA) antibodies and/or anti-La (SSB) antibodies exposed infants [16,17,18,19], which is consistent with our study.In some cases, there is a skin rash right after birth, but the rash may also occur several days later after exposure to ultraviolet light, which may cause or the rash.In a cohort study of 57 infants, rashes can be recognized at an average time of 6 weeks after birth [21].In our study, the appearance time of rash was 16.5 days after birth in average.None appeared immediately after birth, with the earliest appearance time of 5 days, and latest of 30 days after birth.Other systems involvement can occur with no obvious symptoms, including thrombocytopenia in 1-22 days after birth, and anemia, neutropenia and liver enzymes elevation within 2-3 months after birth, which need to pay more attention.Therefore we recommend to monitor closely the hepatic and hematological system within 2-3 months after birth, and treat the patients if necessary.Similar to our data, a follow-up study of infants who had positive anti-SSA antibodies found that some patients developed NLE during follow-up, whose hematological system was involed as anemia, neutropenia, or thrombocytopenia without symptoms.About 56% of cases showed mild and temporary transaminase elevation in 3 months after birth.So researchers recommend clinical and laboratory examinations at the age of 3 months after birth, because hepatic and hematological systems are most probably affected at this time point [22].As children grow up, the speci c autoantibodies taken from their mothers gradually turn negative, and the NLE symptoms and signs also gradually disappear except for cardiac block [22].The prognosis is good for children with dermal, hepatic or haematological systems involvement.A follow-up study of infants whose mothers had positive anti-SSA antibodies found that a majority of hematological system involvement recovered completely within 9 months after birth [22].In our study, the hematological system involvement could recover within 3 months after birth.The hepatic system involvement could all recover within 12 months, and mostly 6 months after birth.But some of the conditions were serious and required timely and active intervention, such as thrombocytopenia.In a severe case, platelet (PLT) could be reduced to 35*109/L due to NLE.
Researchers have found that the anti-SSA of 10% of the infants whose mothers have positive anti-SSA are still positive at the age of 9 months [22].We have found that the median time for anti-SSA, SSB and RNP antibodies to turn negative is 12 months after birth.What's more, the involved systems of NLE patients, especially hepatic system, may not recover until 12 months after birth for some babies.So we recommend that the high-risk babies of NLE to be followed up until at least 12 months after birth.
Studies have shown that young mother, rst child and lack of glucocorticoid therapy may be risk factors to increase the risk of NLE of the offspring [23].Other studies believe that the interaction of genetic susceptibility, intrauterine environment and maternal antibodies is more complex than currently known [24].We have found that there is no signi cant difference between the NLE group and the non-NLE group in mothers' age, delivery mode, abnormal child-bearing history, or the usage of prednisone or hydroxychloroquine during pregnancy, neither of the kinds and amounts of positive autoantibodies, indicating that no single factor can affect the systems involvement of NLE except for cardiac block.
There are some limitations in this study.First of all, our study is a single-center study.Due to the standardized prenatal health care in our center, no neonates have cardiac block in this study.Secondly, this study only includes full-term infants.Thirdly, babies are followed up to 24 months after birth in our study, and more follow-up studies are still needed for the long-term prognosis of high-risk groups of NLE.
In conclusion, the dermal, hepatic, and hematological systems involvement of NLE can gradually appear during follow-up.The affected systems can recover with a good prognosis, but some of the conditions are serious and require timely and active intervention.No single factor has been found to predict whether the high-risk offspring will suffer from NLE or not.Therefore, for all the high-risk populations of NLE, it is necessary to follow up closely within one year after birth, paying attention to the dermal, hepatic, and hematological systems involvement and monitoring their autoantibodies.
Continuous variables were expressed as mean ± standard deviation which correspond to normal distribution, and median (25 percentile/75 percentile) which correspond to non-normal distribution.Continuous data were analyzed using Mann-Whitney U test; categorical variables were examined using Fisher's exact test.And P value < 0.05 was considered signi cant.SPSS for Windows version 23 software (SPSS Inc., Chicago, IL, USA) was used.

Table 1
Comparison of basic information between NLE group and non-NLE group
For infants who did not test their own antibodies after birth (1 case in the NLE group and 19 cases in the non-NLE group) or infants whose NLE-speci c antibodies were all negative (7 cases in the NLE group and 3 cases in the non-NLE group, probably caused by detection errors by different test methods), we treated their mothers' antibodies types as the infants'.There was no signi cant difference in positive rates of SSA antibody (