Surface Antibody Research Articles

Study on the Retreatment, Outcome, and Potential Predictors of Recurrence in Patients With Recurrence of Hepatitis B After Functional Cure.

BackgroundStudies about the retreatment and predictors for patients with hepatitis B recurrence after functional cure are rare. This study aimed to evaluate the effect of retreatment, outcome, and potential predictors of recurrence in patients with recurrence after functional cure.MethodsA long-term follow-up was conducted with 32 cumulatively obtained patients who relapsed after cessation of pegylated interferon (Peg-IFN)-based antiviral treatment. The decision of whether to treatment or which therapeutic method to use [Peg-IFN or nucleos(t)ide analogs (NAs)] was based on the patient’s preferences and wishes. The rate of achieving functional cure and the clinical outcomes of different therapeutic methods were analyzed. Hepatitis B surface antibody (anti-HBs) and hepatitis B core antibody (anti-HBc) levels were detected in patients with blood samples during follow-up to evaluate the predictive ability of recurrence.ResultsThe follow-up time of 32 recurrence cases was 42–532 weeks after recurrence (median 226 weeks). In the 20 patients who received retreatment (15 received Peg-IFN and 5 received NAs only), the rate of functional cure was 65.0% (13/20); it was 86.7% (13/15) in the patients retreated with Peg-IFN. Three cases experienced recurrence again. Five patients received NA treatment, and no functional cure was achieved. No drug intervention was administered for 12 patients, 2 of them with hepatitis B virus (HBV) DNA spontaneous clearance, and one patient achieved spontaneous hepatitis B surface antigen (HBsAg) clearance during follow-up. Patients who relapsed after functional cure with Peg-IFN treatment did not have liver cirrhosis or hepatocellular carcinoma during the follow-up, regardless of whether they received retreatment. Anti-HBs and anti-HBc levels at the end of therapy were predictors of recurrence (p < 0.001, p = 0.023). The value of combining the above two indicators in predicting recurrence was further improved, the areas under the receiver operating characteristic curves were 0.833, at combining predictors >-0.386, the predictive sensitivity and specificity for recurrence were 86.67% and 90.62%.ConclusionThe functional cure rate was above 80% for patients with recurrence treated by Peg-IFN. During the follow-up, liver cirrhosis and hepatocellular carcinoma were not observed in all recurrence cases. High levels of anti-HBs and anti-HBc at the time of drug discontinuation are less likely to relapse.

Improving Hepatitis B Vaccination Rates among At-risk Children and Adolescents with Inflammatory Bowel Disease.

Introduction:Patients with inflammatory bowel disease (IBD) receiving tumor necrosis factor alpha inhibitors (TNFai) may be at higher risk for hepatitis B virus (HBV) infection. We conducted a quality improvement (QI) initiative to improve HBV vaccination rates in seronegative children with IBD.Methods:This QI initiative implemented an HBV vaccination strategy from September 2018 to March 2020 in patients with newly diagnosed IBD with hepatitis B surface antibody (HBsAb) <10 mIU/mL. The project aimed to (1) increase HBV vaccination rates in seronegative patients and (2) document immunogenicity after completing a three-dose vaccine series. Outcome measures included the percentage of seronegative patients who received HBV vaccines (dose 1 and three-dose series). Interventions included applying a standardized vaccination protocol, and creating a vaccine workflow in two clinical areas, previsit planning and stakeholder engagement.Results:One hundred seventy-four children and adolescents with IBD were evaluated during the study period, and 132 (76%) were HBsAb negative. After plan-do-study-act (PDSA) 1, the proportion of eligible patients who received HBV vaccine dose 1 increased from a baseline of 7% to 100% and was sustained for over 12 months. During PDSA 2, the proportion of patients completing the three-dose vaccine series improved from a baseline of 0% to 82% (n = 100); among 93 children in this subgroup who had repeat serology performed, 86 (92%) demonstrated serologic evidence of HBV protection.Conclusions:A multidisciplinary approach applying QI methodology allowed for improved and sustained HBV vaccination rates in at-risk seronegative children and adolescents with IBD. A three-dose HBV vaccine series proved immunogenic in 92% of eligible patients.

Evaluation of Hepatitis B viral Serological Patterns and HBV DNA levels in Hepatitis B subjects at Nauth Nnewi, Southeastern Nigeria

Nigeria is one of the countries with the highest incidence of Hepatitis B Virus (HBV) infection worldwide. The inaccessibility and unaffordability of HBV DNA quantification (viral load) assay which is the key laboratory test for therapy initiation, and monitoring is a challenge to HBV management. This study aimed at evaluating the HBV DNA viral load differences across the serological markers of HBV in order to develop a more cost-effective diagnostic algorithm for Hepatitis B management. Cross sectional study design was used with a total of 264 subjects comprising of 88 HBsAg seropositive treatment naïve subjects, 88 HBsAg seropositive subjects on antiviral therapy as case subjects and 88 age-matched apparently healthy HBsAg seronegative individuals were recruited as control subjects. Hepatitis B Virus DNA assay was performed using real time PCR technique, Hepatitis B core Antibody Immunoglobulin M and Hepatitis D Virus Immunoglobulin G assay. Immunochromatography was used for HBV Panel, Hepatitis C Virus assay and Human Immunodeficiency Virus testing. The non-treatment group has higher viral load (M=805.50 IU/ml) compared with treatment group (M = 65.50 IU/ml) (p&lt;0.001). There was a significant difference in the HBV DNA levels of the four serological patterns observed in the study (P&lt; 0.001). Among the four serological patterns observed, the second pattern with positive surface antibody, positive envelope antigen and negative envelope antibody showed highest viral load (M= 46850189.50 IU/ml) followed by the third pattern with negative surface antibody, negative envelope antigen, and negative envelope antibody (M=46555 IU/ml). The first pattern which has negative surface antibody, negative envelope antigen and positive envelope antibody has the lowest viral load (M=21.00 IU/ml) followed by the fourth pattern with positive surface antibody, negative envelope antigen and positive envelope antibody (M=493.00 IU/ml). This study showed that HBV serological markers can predict viral load and should be used as its alternative in resource poor settings.

Optimized flow cytometry protocol for dihydrorhodamine 123-based detection of reactive oxygen species in leukocyte subpopulations in whole blood

Reactive oxygen species (ROS) and the ability of immune cells to mount an oxidative burst represent an important defense during microbial invasion, but is also recognized for playing a significant role in the progression of inflammatory disorders and disease. Although neutrophils produce the strongest ROS-response, other leukocytes and their cell subsets could play a significant role. Isolation of specific cells for determining their ROS-response can affect their functionality and is laborious or hard to replicate in different settings. We have therefore established a whole blood assay, that only requires 100 μL heparinized blood and utilizes the dihydrorhodamine (DHR) 123 ROS-probe combined with cell surface antibody staining for the specific detection of ROS in several subsets of cells simultaneously using flow cytometry. Although the flow markers chosen are interchangeable with other direct conjugated and cell specific antibodies depending on the research question, we focused on neutrophils (SSChighCD16brightHLA-DRneg/low), eosinophils (SSChighCD16lowHLA-DRlow/negCD193positiveCD125positive) and monocyte subsets (SSCintermediateHLA-DRhighCD14low-positiveCD16negative-positive). As a RBC-lysis reagent we compared BD FACS Lysis Solution to the in-house prepared ammonium-chloride‑potassium based ACK Lysis Buffer, that does not fix or permeabilize the immune cells. We find that ACK-lysis of stimulated and stained samples results in superior surface staining, decreased loss of cell subsets, and enhanced resolution of the DHR-signal. Compared to the other cells analyzed in healthy blood donors, neutrophils responded with the highest ROS-response to all tested stimuli (fMLP (low stimuli), E. coli, and PMA (high stimuli)), where eosinophils and the three monocyte subsets also showed an extensive ROS-response when stimulated with E. coli or PMA. Our assay provides the possibility for researchers to examine the ROS-response of specific cell subsets in specific patient groups ex vivo and could also allow the analysis of pharmacological intervention studies targeting ROS, which ultimately can advance the field of immunological research.

P982: MYELOID-DERIVED SUPPRESSOR CELLS: CHARACTERIZATION IN PERIPHERAL BLOOD AND SPLEEN TISSUE OF PATIENTS WITH MYELOFIBROSIS

Background: Myeloid-derived suppressor cells (MDSCs) are immature myeloid cells that accumulate in patients with malignancies, sepsis or chronic inflammation. Features of MDSCs are: myeloid origin, immature state, strong ability to reduce cytotoxic functions of T/NK cells, potential to differentiate into endothelial cells favoring neoangiogenesis. MDSCs are conventionally divided into polymorphonuclear (PMN)- and monocytic (M)-MDSCs. PMN-MDSCs are characterized by the production of reactive oxygen species and arginase-1 (Arg-1), that directly inhibits T-cell proliferation and activation, while M-MDSCs predominantly express inducible nitric oxide synthase and produce nitric oxide. Myelofibrosis (MF) is characterized by clonal expansion of a hematopoietic stem cell, extramedullary hematopoiesis, bone marrow (BM) fibrosis, splenomegaly, BM and splenic neoangiogenesis and an acquired mutation of JAK2, CALR or MPL gene. Inflammation plays a relevant role in MF pathogenesis, as proven by high levels of inflammatory cytokines with prognostic significance and by a state of chronic oxidative stress. Aims: To characterize circulating MDSCs and assess their localization and quantification in spleen samples from MF patients. Methods: Peripheral blood mononuclear cells (PBMCs) from MF patients (n=15) and healthy subjects (HDs; n=10) were stained with surface antibodies (CD11b, CD14, HLA-DR, CD15, CD33) and intracellular Arg-1 and analysed by flow cytometry. Formalin-fixed paraffin-embedded samples of spleen specimens (MF n=41; HDs n=20) were immunostained with anti-Arg-1 antibody using the automated platform Dako Omnis Envision Flex with the EnVision FLEX, High pH, HRP Rabbit/Mouse High pH [GV800] revelation kit. Results: In PBMCs, the percentage of PMN-MDSCs (identified as CD11b+HLA-DRlow/-CD33dimCD14-CD15+) was higher (p=0) in patients (median 9.2, range 1-59) than HDs (median 1.7, range 0.1-4.5) and directly correlated with the allelic burden (R=0.86, p=0.006) in JAK2-mutated patients. Patients in pre-fibrotic phase (n=7) had levels of PMN-MDSCs (median 9.2, range 1-31.4) comparable to HDs; on the contrary, the percentage of PMN-MDSCs in patients with BM fibrosis (n=8; median 13.3, range 3.5-59) was higher (p=0) than in HDs. All PMN-MDSCs were Arg-1+, both in MF and in HDs. Gene expression studies are ongoing. The expression of the CXCR4 receptor on PMN-MDSCs was lower, although not statistically significant, in MF (median 1.1, range 0-9.3) than in HDs (median 2.5, range 0-11.9). PMN-MDSCs in MF inversely correlated (R=-0.89 p=0.007) with CXCR4 expressed on CD34+ cells. The percentage of M-MDSCs (identified as CD11b+HLA-DRlow/-CD33hiCD14+CD15-) was higher (p=0.015) in HDs (median 0.49, range 0.25-1.5) than in MF (median 0.29, range 0.02-0.76). Immunohistochemical staining of spleen sections showed a higher (p=0) percentage of Arg-1+ cells (calculated on total spleen cells) in MF (median 20, range 5-60) than in HDs (median 7, range 2-20); Arg-1+ cells were present in all patients and predominantly located in the red pulp in MF patients, while in HDs were detected in the white pulp. Summary/Conclusion: The identification of MDSCs and their relation with the mutational status suggest a new mechanism of MF pathogenesis, either related to the chronic inflammatory status of patients or as players of the neoangiogenesis that characterizes the disease. Moreover, the low expression of CXCR4 on PMN-MDSCs could be, on one hand, related to the low expression of this receptor on CD34+ cells or suggest a MF-specific recruitment mechanism, other than that observed in solid tumors.

Longitudinal mapping of hepatitis B vaccine-induced B-cell linear epitopes in healthy individuals.

The elimination of hepatitis B virus (HBV) infection is partially facilitated by the prophylactic HB vaccine. As the loss of seroprotection over time remains a conundrum for long-lasting protection, a comprehensive dynamic analysis of immunogenic targets of the HB vaccine will provide novel insights into the improvement and design of potential targets. In this study, 36 healthy subjects without prior history of hepatitis B infection and negative for hepatitis B surface antibody (anti-HBs) were enrolled. Participants were given a series of three doses of HB vaccine on a 0-, 1-, and 6-month schedule and longitudinally followed up. We systematically mapped 55 overlapping 15-mer peptides covering the small S protein of hepatitis B virus (SHBs) of vaccinees' serum samples at seven time points by performing an ELISA assay. Additionally, the frequencies and function dynamics of adaptive immune response were assessed by flow cytometry. We found that the SHBs peptide coverage presented an overall upward trend along with the vaccination progress, and the individual subpartition recognition was strongly correlated with the anti-HBs titers. Moreover, we identified one dominant epitope (S29) located on "a determinant region" associated with effective vaccine response. Besides, significant correlations between the proportion of plasmablasts and proliferating B cells and levels of anti-HBs were ascertained. Taken together, our data characterized the dynamics of HB vaccine-induced neutralizing antibodies against B-cell linear epitopes on SHBs and adaptive immune response, which will be constructive to develop the next-generation vaccine.

Abstract 1977: Extending the capabilities of a high-parameter immunophenotyping assay with cytoplasmic staining applications for mass cytometry

Abstract Mass cytometry, empowered by CyTOF® technology, utilizes monoisotopic metal-tagged antibodies and a high-sensitivity mass cytometer to allow high-dimensional single-cell analysis in complex biological samples. The 30-marker panel Maxpar® Direct™ Immune Profiling Assay™ (Cat. No. 201325) for suspension mass cytometry provides an unprecedented sample-to-answer solution for detecting and analyzing 30 surface markers in a single experiment. With 18 open mass channels for additional biological markers, the Maxpar Direct Assay facilitates panel expansion and enables flexibility for higher multiplexity and applications. Among the potential complementary applications with the Maxpar Direct Assay, intracellular cytokine staining (ICS) is of particular interest as it may be used to assess infiltrating immune cell phenotypes in the tumor microenvironment. However, for the purpose of assessing cell viability in this workflow, the effectiveness of the Cell-ID™ Intercalator-Rh (Cat. No. 201103) that is included in the Maxpar Direct Assay is in question, since cell permeabilization during ICS can potentially damage the DNA-intercalator bondIn this study, we investigated the compatibility of the Cell-ID Intercalator-Rh (103Rh) with intracellular staining. To do this, we stained either human peripheral blood mononuclear cell (PBMC) or whole blood samples (FLDM-400287) with the Maxpar Direct Assay followed by intracellular staining for the detection of expressed cytokines. The intercalator was evaluated for its ability to discriminate live and dead cells when the sample undergoes surface antibody staining. The monoisotopic Cell-ID Cisplatin-194Pt (Cat. No. 201194) was used as the control to provide benchmark measurement of cell viability of the samples. For both sample types, known percentages of heat-killed PBMC were spiked into the samples in order to evaluate the influence of dead cells. We demonstrate that 103Rh provides equivalent functionality as a cell viability indicator during intracellular staining for cytoplasmic proteins compared to Cell-ID Cisplatin-194Pt. This work was designed to support the use of the Maxpar Direct Immune Profiling Assay in combination with additional intracellular markers. Overall, these findings expand the applicability of Cell-ID Intercalator-Rh (103Rh) to processes that involve cytoplasmic staining. For Research Use Only. Not for use in diagnostic procedures. Citation Format: Noah Saederup, Huihui Yao, Michael Cohen, Leslie Fung. Extending the capabilities of a high-parameter immunophenotyping assay with cytoplasmic staining applications for mass cytometry [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1977.

Association of Coexistent Hepatitis B Surface Antigen and Antibody With Severe Liver Fibrosis and Cirrhosis in Treatment-Naive Patients With Chronic Hepatitis B

Coexistence of hepatitis B surface antigen (HBsAg) and antibody against HBsAg (anti-HBs) constitutes an atypical serological profile in chronic hepatitis B virus infection, and the association between coexistent HBsAg and anti-HBs with severe liver fibrosis and cirrhosis in patients with chronic hepatitis B (CHB) remains unclear. To investigate the association of coexistent HBsAg and anti-HBs with severe liver fibrosis and cirrhosis in patients with CHB. Consecutive treatment-naive patients with CHB from 2 medical institutions in China were enrolled between January 10, 2015, and March 31, 2021. Severe liver fibrosis and cirrhosis were identified using the aspartate transaminase (AST) to platelet ratio index (APRI), the fibrosis index based on 4 factors (FIB-4; factors comprise age, AST level, alanine aminotransferase [ALT] level, and platelet count), transient elastography, or ultrasonography. Data were analyzed from August 1, 2021, to April 15, 2022. The primary outcomes were rates of severe liver fibrosis and cirrhosis among patients with vs patients without coexistant HBsAg and anti-HBs. Severe liver fibrosis was defined as an APRI score of 1.5 or higher, a FIB-4 score of 3.25 or higher, or a liver stiffness measurement of 8 kPa or higher; cirrhosis was defined as an APRI score of 2.0 or higher, a FIB-4 score of 6.5 or higher, a liver stiffness measurement of 11 kPa or higher, or ultrasonographic findings suggestive of cirrhosis. Of 6534 enrolled patients, 4033 patients (61.7%) were male, and the median (IQR) age was 41.0 (33.0-52.0) years. A total of 277 patients (4.2%) had coexistent HBsAg and anti-HBs. Patients with vs without anti-HBs were older (median [IQR], 46.0 [33.0-55.5] years vs 41.0 [33.0-52.0] years) and had a higher proportion of hepatitis B e antigen (HBeAg) positivity (123 of 277 patients [44.4%] vs 2115 of 6257 patients [33.8%]; P < .001), higher ALT levels (median [IQR], 45.1 [24.6-119.0] U/L vs 36.7 [22.0-77.0] U/L; P = .001) and AST levels (median [IQR], 35.0 [23.5-68.4] U/L vs 28.3 [21.6-51.0] U/L; P < .001), and lower platelet counts (median [IQR], 173.0 × 103/μL [129.0-212.5 × 103/μL] vs 185.0 × 103/μL [141.0-224.0 × 103/μL]; P = .004), albumin levels (median [IQR], 4.37 [4.11-4.56] g/dL vs 4.43 [4.17-4.61] g/dL; P = .02), and HBsAg levels (median [IQR], 2.8 log10 [1.6-3.4 log10] IU/mL vs 3.3 log10 [2.6-3.9 log10] IU/mL; P < .001). Compared with patients without anti-HBs, those with anti-HBs had higher APRI scores (median [IQR], 0.5 [0.3-1.4] vs 0.4 [0.3-0.9]; P < .001), FIB-4 scores (median [IQR], 1.4 [0.9-2.6] vs 1.1 [0.7-2.1]; P < .001), and liver stiffness values (median [IQR], 7.5 [6.2-9.8] kPa vs 6.3 [5.2-8.1] kPa; P = .003). Patients with anti-HBs also had higher proportions of severe liver fibrosis (102 of 277 patients [36.8%] vs 1397 of 6207 patients [22.5%]; P < .001) and cirrhosis (87 of 277 patients [31.4%] vs 1194 of 6213 patients [19.2%]; P < .001) compared with patients without anti-HBs. The coexistence of HBsAg and anti-HBs was independently associated with severe liver fibrosis (odds ratio [OR], 2.29; 95% CI, 1.56-3.38; P < .001) and cirrhosis (OR, 1.73; 95% CI, 1.12-2.68; P = .01) in the multivariate analysis. However, the association of coexistent HBsAg and anti-HBs with cirrhosis was only observed in patients with HBeAg negativity (OR, 1.66; 95% CI, 1.05-2.62; P = .03) and not in patients with HBeAg positivity (OR, 1.45; 95% CI, 0.87-2.43; P = .16). In this cross-sectional study, the coexistence of HBsAg and anti-HBs was unusual in hepatitis B virus infection and was associated with more advanced liver diseases, such as severe liver fibrosis and cirrhosis, especially among patients with HBeAg negativity. These results suggest that close monitoring for liver fibrosis and cirrhosis is warranted in patients with CHB who have this serological profile.

Horizontal transmission might be a common route of hepatitis B virus exposure in highly endemic areas.

Hepatitis B virus (HBV) is a common viral pathogen that infects more than a third of the world's population; however, the transmission route remains to be further defined. The 18-year implementation of the free HBV vaccine for children has greatly changed the prevalence of HBV infection in China, which presents a unique real-world model for assessing the pattern of HBV transmission. Cross-sectional data of HBV seromarkers between July 2019 and April 2020 were collected from 53 371 individuals aged 1-60 years in four areas of North to South in Eastern China. Longitudinal data of HBV seromarkers between 2007 and 2020 were collected from 177 adults in an area of South China. The regional- and age-specific changes in HBV seromarkers were analyzed. Overall, positive rates of HBV surface antigen (HBsAg; from 3.44% to 15.1%) and antibody against HBV core antigen (anti-HBc; from 7.6% to 44.0%) significantly increased from North to South. Among persons aged ≤18 years, the positive rates of antibody against HBsAg (anti-HBs) and anti-HBc (+) remained at low levels in the North, while they were increasing among persons aged >12 years in the South, despite higher positive rates of anti-HBs (+). Among persons aged >18 years, the anti-HBs (+) rates remained relatively stable (~60%), while anti-HBc (+) rates increased significantly with age. Up to ~80% of the anti-HBs (+) adults in the South was anti-HBc (+) while it was 13.6% in the North. In the longitudinal cohort, the anti-HBc (+) rate among adults in the South increased by 14.2% during 10 years of follow-up. Horizontal transmission might be a common route in highly endemic areas, and may help to explain the high HBV exposure worldwide. The risk of horizontal transmission among children without seroprotective anti-HBs should be notified in highly endemic areas.

Demonstration of durable hepatitis B immune memory in children vaccinated with a DTaP5-IPV-HepB-Hib infant-toddler series 7 to 8 years previously

ABSTRACT Vaccination against hepatitis B (HepB) provides long-term protection against infection. This is despite a reduction in HepB surface antibody (anti-HBs) concentrations over time to levels below the well-accepted correlate of protection of ≥10 mIU/mL. Continued evidence of immune memory and protection despite declined anti-HBs concentrations can be demonstrated by HepB virus surface antigen challenge studies. Long-term immune memory and protection against HepB infection has not been demonstrated previously for the pediatric hexavalent vaccine DTaP5-IPV-HepB-Hib. This phase 3, multicenter, single-group, open-label challenge study (NCT04490499; EudraCT: 2020–000126–26) evaluated immune memory against HepB infection in children who had received DTaP5-IPV-HepB-Hib at 2, 4, and 11–12 months of age, or at 2, 3, 4, and 12 months of age. At age 8–9 years, they were each challenged with 5 μg of monovalent HepB vaccine. Anti-HBs levels were measured on pre-challenge day 1 and post-challenge day 30. At baseline, 45.4% (93 of 205) had anti-HBs levels ≥10 mIU/mL. On post-challenge day 30, 99.5% (201 of 202) had anti-HBs levels ≥10 mIU/mL, regardless of initial vaccination schedule. Post-challenge, geometric mean concentrations increased 71-fold over baseline and 96.0% of children had a ≥4-fold rise in anti-HBs concentrations with similar results across both dosing schedules. The challenge dose was well tolerated. The robust anti-HBs responses after a single 5-μg dose of HepB vaccine confirm the persistence of a HepB immune memory and demonstrate that DTaP5-IPV-HepB-Hib provides long-term protection against HepB.

Hepatitis B Related Liver Cirrhosis in Oman.

ObjectivesAn estimated 887 000 deaths were due to chronic hepatitis B (CHB) related complications in 2015 worldwide. Most of these deaths were related to decompensated liver cirrhosis and hepatocellular carcinoma (HCC). Oman is a country with an intermediate prevalence of CHB. The Hepatitis B vaccine was introduced in Oman in 1990, with a vaccine coverage rate of > 95% reported in 2005. Despite the association between CHB and liver cirrhosis and HCC, no available data from Oman demonstrates CHB-related liver cirrhosis. We sought to estimate the prevalence of CHB among patients with liver cirrhosis from Oman.MethodsWe conducted a retrospective chart review of patients diagnosed with liver cirrhosis at Sultan Qaboos University Hospital and Armed Forces Hospital between January 2006 and April 2013. All pediatric and adult patients with liver cirrhosis were included. We collected demographic data and liver cirrhosis investigations.ResultsA total of 419 patients were included. Two-thirds of the patients were males. The median age was 59 years. Omani patients represented the majority (97.1%) of patients with cirrhosis. Diabetes mellitus was present in almost half of the patients, and 22.2% indicated alcohol consumption. Evidence of previous or current hepatitis B virus (HBV) infection was found in about half of the cohort (51.3%). Only 3.3% of CHB patients were positive for hepatitis B e-Antigen. HBV DNA was detected in 47 patients (21.9%), of which 20 patients had a high viral load > 2000 IU/ml. More than a third (36.7%) had positive hepatitis B surface antibody (anti-HBs), indicating immunity to HBV, and 27.1% was due to previous HBV infection, 5.2% was immune due to vaccination, and 3.7% had positive anti-HBs and unknown anti-HBc status. Negative anti-HBs was found in 34.1% of the cohort and 29.9% had unknown immunity status. HBV coinfection with HCV was found in 24.7% of HBV patients with cirrhosis.ConclusionsSerological markers of CHB are common among liver cirrhosis patients in Oman. CHB related cirrhosis was more common in old age males than females (70.7% vs. 29.3%, respectively; p < 0.010). Evidence of past or present HBV infection was found in > 50% of the patients.

Hepatitis Virus Reactivation in Patients with Psoriasis Treated with Secukinumab in a Real-World Setting of Hepatitis B or Hepatitis C Infection.

Background and ObjectiveBiologics for psoriasis, especially anti-tumor necrosis factor-α therapies, may reactivate hepatitis B virus (HBV) or hepatitis C virus (HCV) infections, as well in inactive carriers or patients with occult infection. However, some biologics, including anti-interleukin-17 therapies such as secukinumab, seem to be less likely to cause hepatitis reactivation. This study assessed the safety of secukinumab treatment in patients with psoriasis with HBV or HBC infection.MethodsThis was a retrospective cohort study of patients with moderate-to-severe psoriasis treated with secukinumab at seven Italian centers. Patients serologically positive for one or more of the following viral hepatitis markers were included: HCV antibody (± HCV-RNA positivity) and/or hepatitis B surface antigen, and/or HBV core antibody and/or HBV surface antibody (± HBV-DNA positivity). Patients received secukinumab 300 mg subcutaneously at week 0/1/2/3/4 then every 4 weeks; prophylactic therapy before starting secukinumab was prescribed where indicated. The primary study endpoint was the reactivation of hepatitis viral infection, defined as conversion to HBV-DNA or HCV-RNA positivity, with or without elevation of transaminases.ResultsSixty patients (17 with concomitant psoriatic arthritis) were included. Thirteen subjects were hepatitis B surface antigen positive, 19 were HBV core antibody positive, and 30 were positive for the HCV antibody; however, all were HCV-RNA negative. After 53.5 ± 37.5 weeks of secukinumab therapy, hepatitis reactivation occurred in only one patient, who had a reactivation of both hepatitis B and hepatitis C. This patient had not undergone hepatitis B prophylaxis or hepatitis C treatment before secukinumab.ConclusionsThese real-world data support the safety of secukinumab in patients with positive markers of HBV or HCV infection, when administered together with dedicated prophylaxis.

The Investigation of Hepatitis B Vaccine Immune Responses in Occult Hepatitis B Virus-Infected Patients.

ObjectivesThere is no effective treatment for occult hepatitis B virus infection (OBI) patients, and immunotherapy may be one of the most promising options. We aim to investigate the underlying mechanism and therapeutic potential of hepatitis B vaccine immunotherapy for OBI patients.MethodsOutpatient OBI patients were screened and randomly divided into treatment (Group A) and control (Group B) groups. At weeks 0, 4, and 24, patients in Group A received a subcutaneous/intramuscular injection of hepatitis B vaccine (Engerix-B, 20 μg/time) according to the standard vaccination schedule; patients in Group B served as blank control. The patients were followed for 36 weeks, with clinical, biochemical, virological, immunological, and imaging data collected and analyzed at weeks 0, 12, 24, and 36, respectively, and the relation between the virology and immunology results was analyzed.ResultsOf the 228 OBI patients, 28 were excluded, and 200 were enrolled for observation. In the end, 44 patients were included in Group A and 39 in Group B after excluding lost cases. At week 0 (baseline), some patients in two groups had liver disease symptoms, HBV-related liver function damage, and liver fibrosis. 86.36% (38/44) and 82.05% (32/39) patients were positive for serum hepatitis B surface antibodies (anti-HBs) in Group A and Group B, respectively, with the median (quartile) of 42.47 (16.85, 109.1) and 39.27 (16.06, 117.4) mIU/ml, respectively. Reduced peripheral blood CD4+T, CD8+T, and B lymphocytes were found in some patients in two groups. These results were not statistically different between Group A and Group B (P>0.05). At week 36, all patients were serum anti-HBs (+) in Group A, with a median (quartile) of 1000 (483.9, 1000) mIU/ml, which was significantly higher than that at week 0 (P<0.05) and that in Group B (P<0.05). Compared to week 0, the number of CD8+ T and B lymphocytes increased significantly and were significantly higher than Group B at the same point. Two patients in Group B were found to have hepatitis B virus reactivation from week 12 to week 36.Correlation AnalysisAnti-HBs in Group A patients were positively correlated with B lymphocytes (r=0.3431, 0.3087, and 0.3041, respectively) and positively correlated with CD8+ T lymphocytes (r=0.4954, 0.3054, and 0.3455, respectively) at weeks 12, 24, and 36.ConclusionVirological reactivation is a risk for OBI patients. Serum hepatitis B surface antibodies were significantly increased after hepatitis B vaccine treatment, the same as the numbers of peripheral blood B and CD8+ T lymphocytes; changes in hepatitis B surface antibody levels were positively correlated with the changes in peripheral blood B and CD8+ T lymphocytes.

Factors predicting neuronal surface antibodies in the elderly with new-onset and unknown seizures.

ObjectiveTo investigate risk factors of neuronal surface antibodies (NSAbs) and develop a nomogram that could identify patients at the odds of NSAbs among the elderly (aged 60 years or older) with new‐onset seizures of unknown etiology.MethodsClinical data for aged ≥60 years diagnosed with new‐onset seizures of unknown etiology were retrospectively reviewed. A nomogram based on multivariable logistic regression was constructed. Model performance of nomogram was evaluated using area under the curve (AUC), calibration curve, decision curve analysis (DCA), and clinical impact curve (CIC). Meanwhile, it was internally validated by bootstrap validation in current cohort.ResultsOf 147 patients included in final analysis, 68 (46.3%) had NSAbs‐mediated encephalitis. Six factors were identified: duration of seizures less than 3 months (OR:14.259; 95% CI: 4.480–45.386), focal‐onset seizures (OR:12.457; 95% CI: 2.710–57.261), psychiatric deficits (OR:10.063; 95% CI: 3.231–31.343), sleep disorders (OR:3.091; 95% CI: 1.011–9.454), hyponatremia (OR:6.252; 95% CI: 1.445–27.043), and medial temporal lobe (MTL) lesions on MRI (OR:4.102; 95% CI: 1.382–12.169). The nomogram had a good discrimination with an AUC of 0.916 and with a corrected AUC of 0.881 after the bootstrapping validation, our model also exhibited a better predictive performance than scoring systems commonly used clinically. Additionally, the calibration curve showed that predicted NSAbs‐positive rates of nomogram were closely aligned with actual observed results. Moreover, the nomogram achieved well on clinical utility by using the DCA and CIC.InterpretationOur nomogram may provide a convenient and useful tool for identifying the elderly with new‐onset seizures of unknown etiology who are at risk of NSAbs‐mediated encephalitis, which would allow these patients receive earlier immunotherapy.

Dynamic Changes of Cytokine Profiles and Virological Markers Associated With HBsAg Loss During Peginterferon Alpha-2a Treatment in HBeAg-Positive Chronic Hepatitis B Patients.

ObjectiveTo explore dynamic changes of cytokines and virological markers associated with hepatitis B surface antigen (HBsAg) loss during peginterferon alpha-2a (PEG-IFN α-2a) treatment in hepatitis B e antigen (HBeAg) positive chronic hepatitis B (CHB) patients.MethodsIt was a single-center prospective cohort study. HBeAg-positive CHB patients were prospectively and consecutively enrolled. Cytokines were detected at baseline, week 12 and 24 of PEG-IFN treatment. HBsAg disappearance rate was the primary evaluation index at 48 weeks of PEG-IFN treatment.ResultsAmong 100 patients who completed the 48-week PEG-IFN α-2a treatment, 38 patients achieved serum HBeAg disappearance, 25 patients achieved HBeAg seroconversion, 9 patients achieved functional cure, 37 patients had HBsAg decline of ≥1 log IU/ml, and 8 patients produced hepatitis B surface antibody (HBsAb). Albumin (ALB), fms-like tyrosine kinase 3 ligand (FLT3-L) and interferon-alpha2 (IFN-α2) in the clinical cure group were significantly lower than those in the non-clinical-cure group at baseline. After 12 weeks of treatment, HBsAg in the clinical cure group was significantly lower than that in the non-clinical-cure group (median 1.14 vs. 3.45 log10IU/ml, Z=-4.355, P < 0.001). The decrease of HBsAg and hepatitis B virus desoxyribose nucleic acid (HBV DNA) in the clinical cure group was significantly higher than that in non-clinical-cure group (median: HBsAg 1.96 vs. 0.33 log10IU/ml, Z=-4.703, P< 0.001; HBV DNA 4.49 vs.3.13 log10IU/ml, Z=-3.053, P=0.002). The increase of IFN-α2 in the cure group was significantly higher than that in the non-clinical-cure group (497.89 vs. 344.74, Z=-2.126, P=0.034). After 24 weeks of treatment, HBsAg, HBeAg, Flt3-L, and IL-10 in the clinical cure group were significantly lower than those in the non-clinical-cure group (median: HBsAg 0.70 vs. 3.15 log10IU/ml, Z=-4.535, P< 0.001; HBeAg 1.48 vs. 13.72 S/CO, Z = 2.512, P = 0.012; Flt3-l 0.00 vs 2.24 pg/ml, Z = 3.137, P=0.002; IL-10 0.70 vs. 2.71 pg/ml, Z=-4.067, P < 0.001). HBsAg decreased significantly in the clinical cure group compared with non-clinical-cure group (median 3.27 vs. 0.45, Z=-4.463, P < 0.001).ConclusionDynamic changes of cytokines and virology markers during early PEG IFN α-2a treatment were associated with HBsAg loss in HBeAg-positive CHB patients.

An explanatory analytics model for identifying factors indicative of long- versus short-term survival after lung transplantation

Due to the shortage of available organs compared to the number of patients on waitlists, the organ allocation process has always been challenging and calls for an equitable and optimized allocation system. This system demands minimizing the waitlist mortality and improving transplantation benefits (e.g., survival time and quality of life). According to prior research, lung recipients’ long-term survival time is lower than other solid organs recipients. This paper proposes and implements an explanatory analytics framework to study the most prominent factors contributing to long-term survival after lung transplantation. We collect data from the United Network for Organ Sharing registry database. It contains more than 44 thousand unique patients undergoing lung transplantation in the US since 1987. In our proposed framework, we first employ several machine learning (ML) algorithms for classification and choose the best-performing one for further analysis of factors. Then, we utilize a state-of-the-art explainable artificial intelligence method with the chosen ML algorithm for the model explanation and interpretation. The proposed framework lists the most critical factors influencing long-term survival after lung transplantation and their corresponding importance measures. For instance, our results suggest that Hepatitis B surface antibody (HBV_SURF) and forced expiratory volume in one second (FEV1) are among the important factors but have not been well examined by lung transplant researchers. Our framework is also able to provide the main contributing factors for each patient individually. Medical scholars and practitioners can use our findings for further analysis and improvement of the lung allocation system.

Role of the pathologist in the diagnosis of autoimmune encephalitis.

Autoimmune encephalitis is a group of non-infectious immune-mediated inflammatory disorders manifesting with epilepsy and encephalitis syndromes that are associated with autoantibodies in the serum and/or cerebrospinal fluid (CSF). Pathogenic autoantibodies have been discovered against intracellular onconeural antigens, surface neuronal, or synaptic antigens with distinctive pathogenesis that underlie differences in response to immunotherapy. The onconeural antigens incite cytotoxic T-cell-mediated neuronal destruction, whereas surface antigens trigger direct damage by autoantibodies via complement mediated pathways, and hence respond well to immunomodulatory therapy, in contrast to poor response in the former. Neuroimaging, electroencephalogram, and CSF findings being non-specific, detection of autoantibodies is essential for a confirmatory diagnosis. Detection methods available include tissue-based assay, cell-based assays, immunoblot, cell culture, flow cytometry, and enzyme-linked immunosorbent assays. In this review, we discuss the various testing modalities available for onconeural and cell surface antibodies, their sensitivity and specificity and the emerging role of the pathologist in the diagnosis of autoimmune encephalitis. Early diagnosis is crucial for instituting treatment and preventing morbidity and mortality.

The effects of dietary zinc supplement on intracellular zinc concentration in bovine immune cells

Abstract Zinc (Zn) is one of the most abundant metals in our body and is critical for innate and adaptive immune functions. In humans and mice, Zn deficiency may lead to immunodeficiency, reduced reproductive health, and increased disease susceptibility. However, trace mineral requirements and evidence of Zn dysregulation are poorly defined in cattle. This study aimed to evaluate intracellular Zn concentrations in immune cells from young growing steers with different Zn dietary supplements. Eighteen Angus crossbred steers were stratified by body weight and randomly assigned to two dietary treatments groups: 1) control, no supplemental Zn, or 2) high Zn; 150 ppm of Zn delivered in premixes in total mixed rations. To evaluate the intracellular Zn concentration of immune cells, whole blood cells were loaded with FluoZinTM-3, a fluorescent cell-permeant Zn indicator, then stimulated with Zn pyrithione (100 μM) or TPEN [N,N,N′,N′-tetrakis(2-pyridinylmethyl)-1,2-ethanediamine] (50 μM) to measure maximum and minimum mean fluorescent intensity. Cells were stained with surface antibodies to identify immune cell populations and data were collected by flow cytometry. The intracellular Zn concentrations of circulating CD8 T cells and γδ T cells, but not other populations, differed significantly between treatment groups. These results indicate that intracellular Zn concentration can be influenced by dietary Zn intake, and that certain immune populations are differentially affected by Zn status. This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2020-06540 from the USDA National Institute of Food and Agriculture.

Dietary Zinc supplementation impact on relative frequency and innate function of peripheral bovine immune cells

Abstract Zinc (Zn) is an essential micronutrient which is critical for optimum innate and adaptive immune function. While Zn deficiency leads to reduced growth and feed intake, the impact of Zn deficiency and supplementation on immune responses in beef cattle has not been properly addressed. To investigate to impact of dietary Zn supplementation on peripheral immune responses of growing cattle, 27 Angus crossbred steers were stratified by body weight and randomly assigned to three dietary treatments groups: 1) CON, no supplemental Zn, 2) Zn100, 100ppm of Zn or 3) Zn150, 150ppm of Zn delivered in premixes in total mixed rations. Whole blood obtained at 14 (D14) and 58 (D58) days after the start of the treatments was plated and stimulated with TLR agonists LPS (TLR4), PAM3CSK4 (TLR2/1), and Poly-I:C+Imiquimod (TLR3/7) for 48 h, then pro-inflammatory cytokine secretion was measured by ELISA on the supernatants. Additionally, whole blood was stained with surface antibodies to determine relative frequency of peripheral immune cell populations. We observed diminished IL-1ß secretion in response to Poly-I:C+Imiquimod in the CON group when compared to Zn150 (p=0.09) and Zn100 group (p=0.013) at D14 and D58, respectively. We observed no differences in IL-6 secretion upon TLR agonist stimulation between groups at any time point. Further, although no significant changes in CD4+, CD8+, TCRδ+ T cells, or CD21+MHCII+ B cells were observed, CD5+MHCII+ B cells had a higher relative frequency in the CON group when compared to Zn100 (p 0.03) and Zn150 (p 0.10) groups at D58. Future studies will address how Zn supplementation impacts the immune response to infection and disease susceptibility. This project was supported by Agriculture and Food Research Initiative Competitive Grant no. 2020-06540 from the USDA National Institute of Food and Agriculture

The Clinical Spectrum of Autoimmune-Mediated Neurological Diseases in Paediatric Population.

Neurological autoimmune diseases have various origins and pathogeneses. Specific antibodies are associated with paraneoplastic syndromes, other infectious agents, or inherited disorders. We aim to evaluate the relation between the autoantibodies, the chosen symptoms, demographic characteristics, and infection history. We retrospectively analysed 508 children during neurological diagnostics. We investigated serum antineuronal, IgG, IgM anti-ganglioside, and anti-aquaporin-4 in both the serum and cerebrospinal fluid (CSF) anti-cell surface and anti-synaptic protein antibodies in 463, 99, 44, 343, and 119 patients, respectively. The CSF polymerase chain reaction detection of Herpesviridae, enterovirus, B19 parvovirus, adenovirus, and parechovirus involved 261 patients. We included available clinical information and electroencephalographic, radiologic, and microbiological results. The IgM anti-ganglioside antibodies increased the risk of tics and positive symptoms (p = 0.0345, p = 0.0263, respectively), the anti-glutamic acid decarboxylase particle of paresis (p = 0.0074), and anti-neuroendothelium of mutism (p = 0.0361). Anti-neuroendothelium, IgM anti-ganglioside, and CSF anti-N-methyl-D-aspartate antibodies were more often associated with consciousness loss (p = 0.0496, p = 0.0044, p = 0.0463, respectively). Anti-myelin antibodies co-occured with Herpes simplex virus (HSV)-2 IgG (p = 0.0415), anti-CV2 with HSV-1 IgM (p = 0.0394), whereas anti-glial fibrillary acidic protein was linked with past Epstein-Barr virus infection. The anti-ganglioside IgM and anti-myelin particles were bilaterally correlated (p = 0.0472). The clinical pictures may overlap, requiring specialistic diagnostics. We noticed the links between the infection aetiology and the specific autoantibody’s positivity.