Abstract
Background
Hyaline globules (HGs) in the cytoplasm of Kupffer cells (KCs) have been appraised for being a typical feature of autoimmune hepatitis (AIH). This study aimed to determine how useful Kupffer cell hyaline globules (KCHGs) are in diagnosing AIH vs. other causes of pediatric chronic liver diseases (PCLDs).
Materials and methods
This retrospective study recruited 124 children; 58 with AIH, 50 with chronic hepatitis C virus (HCV) infection, and 16 with Wilson’s disease (WD). Two pathologists retrieved paraffin blocks of liver biopsies and prepared new cut sections for Periodic acid–Schiff–Diastase (PAS-D) stain. They independently examined liver biopsies before starting treatment. Two pediatricians reviewed medical records for demographic, clinical, laboratory, and serological findings.
Results
Females represented 48.6% of the studied children with a median age of 5.8 (4.9) years. Pathologists identified KCHGs in 67.24%, 12.5%, and 6.0% of AIH, WD, and HCV affected children respectively, P < 0.001. A significantly higher proportion of seropositive than seronegative AIH patients had KCHGs (77.5% vs. 50.0%), (P < 0.05). In multivariate analysis, KCHGs and prolonged prothrombin time were the only significant predictors that differentiate between AIH and the other studied PCLDs. The odds ratio of having AIH increased 68 times if KCHGs were seen. Among children with AIH, the presence of KCHGs was associated with higher median levels of direct bilirubin 2.2 (1.3) vs. 1.2 (2.2), and immunoglobulin G 3.2 (1.9) vs. 2.0 (1.7), (P < 0.05), but not to histopathological findings or hepatic fibrosis and activity.
Conclusions
KCHGs are key indicators that can differentiate between AIH and other PCLDs, and between seropositive and seronegative AIH.
Keywords: liver histopathology, hyaline globules, Kupffer cells, autoimmune hepatitis, hepatitis C virus infection, Wilson’s disease
Abbreviations: AIH, Autoimmune hepatitis; ALP, Alkaline phosphatase; ALT, Alanine aminotransferase; AMA, Antimitochondrial antibody; ANA, Antinuclear antibody; antiLC1, Antiliver cytosol type 1 antibody; AST, Aspartate aminotransferase; GGT, Gamma-glutamyl transpeptidase; H&E, Hematoxylin and eosin; HCV, Hepatitis C virus; IAIHG, International Autoimmune Hepatitis Group; KCHGs, Kupffer cell hyaline globules; KCs, Kupffer cells; LKMA, Liver-kidney microsome 1 antibody; PAS, Periodic Acid Schiff; PAS-D, Periodic acid–Schiff–Diastase; PCLD, Pediatric chronic liver disease; SMA, Smooth muscle antibody; WBCs, White blood cell count; WD, Wilson’s disease
Autoimmune hepatitis (AIH) is a chronic immune-mediated liver injury that affects all ages, both sexes, and all ethnicities.1,2 Patients may present with different clinical manifestations due to the heterogeneous nature of AIH. Children with AIH may be asymptomatic, chronically ill, or rarely present with fulminant hepatic failure.3 All children with acute or chronic liver inflammation should be evaluated for AIH. Indeed, AIH is diagnosed using a combination of histological, laboratory, and clinical features. These diagnostic features may vary between patients with the same disease and can occur in other pediatric chronic liver diseases (PCLDs).2,4 The International Autoimmune Hepatitis Group (IAIHG) included liver biopsy for the definitive diagnosis ideally before starting the immunosuppressive treatment.5
In the serum of children suspected of having AIH, specific autoantibodies are crucial for diagnosing and identifying the disease subtype. Therefore, screening for antinuclear antibody (ANA), smooth muscle antibody (SMA), and liver-kidney microsome 1 (LKM1) antibody is often carried out whenever AIH is included in the differential diagnosis list of acute or chronic liver condition. Children with positive ANA and/or SMA are classified as type 1 AIH, while children with LKM1 antibody and/or antiliver cytosol (antiLC1) are classified as type 2 AIH.5
Unfortunately, more than 10% of children with AIH do not produce the aforementioned antibodies, and they are diagnosed as seronegative AIH.6 In this case, liver biopsy is mandatory as it may show the characteristic histopathological findings commonly seen in seropositive AIH like portal fibrosis, interface hepatitis, septal fibrosis, necrosis (pan lobular, bridging, and centrilobular), and plasma cells infiltration.7,8 Further evidence that aid in the diagnosis of seronegative AIH is the normalization of aminotransferases and of prothrombin time (PT) after starting immunosuppressive drugs.9
Remarkably, the histopathological findings of liver biopsy are characteristics, but not pathognomonic features of AIH.10 However, liver biopsy is necessary to confirm the diagnosis and to assess the severity of liver tissue damage.1,11 In the absence of a single diagnostic test for AIH, the IAIHG has devised a diagnostic system that includes several positive and negative scores, the sum of which gives a value suggestive of probable or definite AIH.12
Kupffer cells (KCs) hyperplasia was demonstrated in liver tissue of children with AIH, although the function that these cells play was unknown in the disease process.13,14 Immunoglobulin’s deposition was demonstrated in the cytoplasm of KCs in AIH over 50 years ago by immunofluorescence techniques.14 These reports, on the other hand, did not mention any distinctive inclusions in KCs on hematoxylin and eosin (H&E) stained parts or any other special stains.3
The goal of the current work was to report the diagnostic value of Kupffer cell hyaline globules (KCHGs) in distinguishing AIH from other causes of chronic hepatitis. Furthermore, the researchers wanted to study whether there was an association between the presence of KCHGs and immunoglobulin level or the severity of the histopathological findings.
Patients and methods
Study Population
Paraffin blocks of liver biopsy from all children included in this study were retrieved from the archives of the Department of Pathology from January 2014 to December 2018. We excluded slides that contained less than six portal tracts. In total, we included 124 children. Children were classified into three groups: 58 with AIH, 50 with chronic hepatitis C virus (HCV) infection, and 16 with Wilson’s disease (WD). Medical records of children were reviewed for sociodemographic data (age and sex), clinical presentation, laboratory data [total bilirubin (TB); direct bilirubin (DB), platelets (PLT), alanine transaminase (ALT), aspartate aminotransferase (AST), PT, hemoglobin (Hb), and white blood cells (WBCs)], and autoimmune serological markers (ANA, SMA, and anti-LKM1 antibodies).
Histopathological Evaluation
Five-μm thick sections were cut, mounted on glass slides, and stained with H&E, Masson-Trichrome, Perls’ Prussian blue, Orcein, and periodic acid Schiff followed by diastase digestion Periodic acid–Schiff–diastase (PAS-D). Pathologists assesed the histopathological criteria of the AIH disease such as the nature and extent of lymphocytic infiltrate, hepatocyte rosette formation, emperipolesis, bile duct injury, and cholestasis. The stage of liver fibrosis and grade of necroinflammatory activity was evaluated according to the modified Ishak scoring system.15 Slides were examined on Olympus Microscope at ×20. Liver biopsies were assessed for the presence or absence of KCHGs. Generally, KCHGs were identified as round circumscribed homogenous pale-pink globules, separated from each other, and did not coalesce.
Immunohistochemistry
Immunohistochemistry was applied on 4 μm-thick sections prepared from paraffin blocks of liver biopsies. After deparaffinization and rehydration, hydrogen peroxide was applied to block nonspecific background staining. Heat-induced antigen retrieval was performed using citrate buffer solution high pH (pH 9 for 20 min at 97 °C). This was followed by incubation for an additional 20 min in the warm buffer. Tissue sections were incubated with a monoclonal CD68 antibody (clone KP1,0, DAKO A/S, Glostrup, Denmark, dilution 1: 50) overnight at 4 °C. Detection of the immunostaining was carried out using the EnVisionTM FLEX/HRP detection system (DAKO A/S, Glostrup, Denmark) with the 3-diaminobenzidine (DAKO) as the chromogen. After counterstaining with Mayer’s hematoxylin, the immunoreactivity for CD68 was identified as membranous brownish discoloration of macrophages. All histopathological examinations were done blindly by two pathologists,. and in case of discrepancy, slides were examined on a multihead microscope until agreement achieved. Histopathological findings of children with AIH were examined before starting treatment. These findings were compared with HCV infected individuals and WD patients who had similar inflammatory grades and a stage of fibrosis.
Ethics
The current research was approved by the Ethical Review Board Committee at the National Liver Institute, Menoufia University.
Statistical Analysis
Median and interquartile ranges were used to present quantitative data. The categorical data were presented in numbers and percentages. The statistician used the Mann–Whitney and Kruskal Wallis tests to compare the median values of different variables. For categorical data, significance was tested by χ2-test or Fisher’s exact test. Results were considered significant if P value < 0.05. Statistical analysis was performed using Statistical Package for the Social Sciences (SPSS) software version 25 (SPSS Inc, Chicago, IL, USA). A binary logistic regression model was constructed to identify the independent predictors for diagnosis (AIH vs. not AIH). All variables with P < 0.15 in the univariate analysis were included in the model. The final model included age, total bilirubin, liver enzymes (AST/ALT), total protein, serum albumin, hemoglobin, WBCs, platelet count, PT, and KCHGs. The Odds Ratio (OR) and 95% confidence interval (CI) were reported for all variables.
Results
Females comprised 48.6% of the recruited sample (56.9% and 32.0% of children with AIH and chronic HCV infection, respectively, while an equal number of both sexes were allocated to WD). Children in this study had a median age of 5.8 (4.9) years. Refer to supplementary data Table 1, Table 2 for more information about the characteristics and clinical profile of the studied population.
Table 1.
Distribution of Hyaline Globules Among Patients With Different Chronic Liver Disease and Different Types of Autoimmune Hepatitis.
| Total sample (n = 124) | AIH (n = 58) |
HCV (n = 50) |
Wilson (n = 16) |
|||
|---|---|---|---|---|---|---|
| Positive | n | 39 | 3 | 2 | 61.3 | <0.001∗ |
| % | 67.2 | 6.0 | 12.5 | |||
| Negative | n | 19 | 47 | 14 | ||
| % | 32.8 | 94.0 | 87.5 |
| AIH type (n = 58) | Seropositive (n=40) |
Seronegative (n=18) |
|||
|---|---|---|---|---|---|
| Positive | n | 31 | 9 | 3.929 | 0.047∗ |
| % | 77.5 | 50.0 | |||
| Negative | n | 9 | 9 | ||
| % | 22.5 | 50.0 |
AIH: autoimmune hepatitis, HCV: hepatitis C virus, Positive: contain 1, or 2 hyaline globules, ∗P < 0.05.
Table 2.
Binary Logistic Regression Model to Identify the Predictors That Differentiate Autoimmune Hepatitis From Other Chronic Liver.
| Predictors | Univariate |
Multivariate |
|||
|---|---|---|---|---|---|
| P-value | Odd ratio | (95% CI) | P-value | ||
| Constant | 0.59 | ||||
| Age (days) | <0.001 | 0.82 | 0.60–1.12 | 1.12 | 0.21 |
| TB (mg/dl) | <0.001 | 1.05 | 0.46–2.39 | 2.39 | 0.91 |
| ALT (IU/L) | <0.001 | 1.01 | 0.98–1.04 | 1.04 | 0.41 |
| AST (IU/L) | <0.001 | 1.01 | 0.98–1.05 | 1.05 | 0.42 |
| TP (gm/dl) | 0.02 | 0.30 | 0.03–2.74 | 2.74 | 0.29 |
| Albumin (gm/dl) | <0.001 | 2.73 | 0.36–20.50 | 20.50 | 0.33 |
| HB (gm/dl) | <0.001 | 0.59 | 0.27–1.25 | 1.25 | 0.17 |
| WBC ( × 103/ml) | 0.03 | 1.13 | 0.66–1.93 | 1.93 | 0.65 |
| PLT ( × 103/ml) | <0.001 | 0.99 | 0.98–1.01 | 1.01 | 0.24 |
| Hyaline globules | <0.01 | 60.95 | 4.42–841.13 | 841.13 | 0.00∗ |
| PT (Sec) | <0.001 | 3.00 | 1.08–8.34 | 8.34 | 0.04∗ |
TB: total bilirubin; PLT: platelets; ALT: alanine transaminase; AST; aspartate aminotransferase; PT: prothrombin time; Hb: hemoglobin; WBCs: white blood cells, ∗ P value < 0.05.
Distribution of hyaline globules among different pediatric chronic liver diseases
Histopathological examination revealed that more than two-thirds (67.2%) of children diagnosed as AIH had at least one KCHG, compared to 6.0% and 12.5% of children with chronic HCV infection and WD, respectively, P < 0.001. Figure 1a,b CD68 positive KCs were observed hypertrophied in AIH liver tissue as shown in Figure 1c. KCHGs were negative in contrast with the phagolysosome granules in the KCs, which exhibited strong positivity (Figure 1d). Table 1.
Figure 1.
Liver tissue section from a child with AIH. a) D-PAS stain showing many KCs demonstrating hyaline globules (arrows), b) Higher magnification of the previous section revealing the hyaline globules in Kupffer cells (arrow), c) Immunostaining of CD68 highlighting Kupffer cells (arrow), d) granular appearance in the cytoplasm of KCs demonstrating phagolysosomes (arrow). Original mag.×400.
Distribution of hyaline globules among different types of autoimmune hepatitis
Furthermore, a higher proportion of children with AIH, regardless of its type, had at least one KCHG compared to seronegative AIH; 77.5% (31/40) vs. 50.0% (9/18), P < 0.05. Table 1 In total, 27 of 35 (77.1%) children with type 1 AIH had been tested positive for KCHGs. Similarly, 80.0% (4/5) of type 2 AIH had at least one KCHG. This difference between type 1 AIH, type 2 AIH, and seronegative AIH was not statistically significant. Supplementary Table 3.
All significant predictors identified in univariate analysis between AIH and non-AIH (HCV and WD) children were used to build a binary logistic regression model to differentiate between AIH and other studied PCLD. Interestingly, the presence of KCHGs and PT were the only significant predictors of AIH. The OR of having AIH increased by 84 times if the liver biopsy contains one or more KCHGs, Table 2 Regarding the developed regression model, the area under the curve was 0.87%, and R2 was 91%; this means that about 91% of the dependent variable variance (being diagnosed as AIH or not) was explained by the regression model. Figure 2.
Figure 2.
A receiver operating characteristic curve of the regression model used to predict the type of chronic liver disease.
Different laboratory and histopathological findings based on the presence of hyaline globules
Among the 58 children with AIH, the median total bilirubin, direct bilirubin, ALT, AST, ALP, Albumin level, GGT, TP, HB, WBCs, and platelets were [3.0 (3.2), 1.6 (2.7), 242.5 (418.0), 269.0 (432.8), 287.0 (164.7), 58.5 (72.8), 6.7 (0.8), 3.2 (1.4), 16.5 (5.9), 2.6 (2.0), 10.7 (1.8), 7.2 (3.9), 147.5 (122.3)] respectively. Regarding autoantibodies, about 31.1% (18 children) were tested negative for autoantibodies, positive ANA alone or ANA with SMA were diagnosed in 43.1% (25 children) and 15.5% (9 patients) respectively, SMA and anti-LKM1 antibodies were detected in 1.7% (1 child) and 8.6% (5 children) respectively.
It is worthy to note that all the studied variables were not statistically significant in-between children with positive and negative KCHGs except for the median score of direct bilirubin level and gamma globulin levels. The median direct bilirubin level was significantly higher if KCHGs were seen 2.2 (3.1) compared to KCHGs negative children 1.2 (2.2), P = 0.044. Similarly, the median serum level of immunoglobulin G (IgG) was significantly higher among KCHGs positive than KCHGs negative children [3.2 (1.9) and 2.0 (1.7)], P = 0.01. Table 3.
Table 3.
Laboratory Results of Patients With AIH Based on the Presence or.
| Item |
Total |
Hyaline globules |
Hyaline globules |
Test statistics | P- |
|---|---|---|---|---|---|
| Median (IQR) | (n = 58) | Positive (n = 39) | Negative (n = 19) | ||
| TB (mg/dl) | 3.0 (3.2) | 3.1 (3.7) | 2.7 (3.4) | −1.3 | 0.205 |
| DB (mg/dl) | 1.6 (2.7) | 2.2 (3.1) | 1.2 (2.2) | −2 | 0.044∗ |
| ALT (IU/l) | 242.5 (418.0) | 386.0 (627.0) | 221.0 (418.0) | −1.2 | 0.25 |
| AST (IU/l) | 269.0 (432.8) | 358.0 (553.0) | 221.0 (418.0) | −1.3 | 0.188 |
| ALP (IU/l) | 287.0 (164.7) | 306.0 (131.0) | 267.0 (168.0) | −1.7 | 0.089 |
| GGT (IU/l) | 58.5 (72.8) | 71.0 (96.0) | 58.0 (74.0) | −0.2 | 0.836 |
| TP (gm/dl) | 6.7 (0.8) | 6.6 (0.6) | 6.8 (0.8) | −1.8 | 0.07 |
| Albumin (gm/dl) | 3.2 (1.4) | 2.8 (0.9) | 3.4 (1.6) | −1.6 | 0.115 |
| PT (Sec) | 16.5 (5.9) | 16.9 (5.0) | 16.4 (5.0) | −0.6 | 0.573 |
| IgG | 2.6 (2.0) | 3.2 (1.9) | 2.0 (1.7) | −2.6 | 0.01∗ |
| Hb (gm/dl) | 10.7 (1.8) | 10.2 (1.8) | 10.8 (1.5) | −0.1 | 0.921 |
| WBC (×103/ml) | 7.2 (3.9) | 7.5 (3.7) | 6.4 (4.4) | 0 | 0.993 |
| PLT (×103/ml) | 147.5 (122.3) | 149.0 (186.0) | 186.0 (122.0) | −0.3 | 0.778 |
| Autoantibodies n (%) | |||||
| ANA | 25 (43.1) | 19 (48.7) | 6 (31.6) | 3.98 | 0.416 |
| ASMA | 1 (1.7) | 1 (2.6) | 0 (0.0) | ||
| LKMA1 | 5 (8.6) | 4 (10.3) | 1 (5.3) | ||
| ANA + SMA | 9 (15.5) | 6 (15.4) | 3 (15.8) | ||
| Seronegative | 18 (31.1) | 9 (23.1) | 9 (47.7) | ||
| Activity score Median (IQR) |
15.0 (4.3) | 13.0 (5) | 15.0 (4.0) | −0.4 | 0.689 |
| Ishak score Median (IQR) |
3.0 (2.0) | 3.0 (2.0) | 3.0 (2.0) | −0.9 | 0.355 |
| Plasma cells Median (IQR) |
2.0 (2.0) | 2.0 (1.0) | 2.0 (2.0) | −1.2 | 0.23 |
| Stage of fibrosis n (%) | |||||
| No | 7 (12.1) | 6 (15.4) | 1 (5.3) | 1.65 | 0.713 |
| Mild | 18 (31.0) | 11 (28.2) | 7 (36.8) | ||
| Moderate | 28 (48.3) | 18 (46.2) | 10 (52.6) | ||
| Severe | 5 (8.6) | 4 (10.3) | 1 (5.3) | ||
| Disease activity fibrosis n (%) | |||||
| No activity | 4 (6.9) | 3 (7.7) | 1 (5.3) | 3.42 | 0.345 |
| Mild activity | 27 (46.6) | 17 (43.6) | 10 (52.6) | ||
| Moderate activity | 21 (36.1) | 13 (33.3) | 8 (42.1) | ||
| Severe activity | 6 (10.3) | 6 (15.4) | 0 (0.0) | ||
| Confluent necrosis (yes) | 36 (62.1) | 26 (66.7) | 10 (52.6) | 1.07 | 0.301 |
| Hepatocellular necrosis (yes) | 50 (86.2) | 34 (87.2) | 16 (84.2) | 0.75 | 0.54 |
| Plasma cell | 52 (89.7) | 36 (89.7) | 16 (84.2) | 3.57 | 0.055 |
| Interface hepatitis | |||||
| No | 14 (24.1) | 8 (20.5) | 6 (31.6) | 3.6 | 0.55 |
| Score 1 | 15 (25.9) | 10 (25.6) | 5 (26.3) | ||
| Score 2 | 16 (27.9) | 13 (33.3) | 3 (15.8) | ||
| Score 3 | 11 (19.0) | 6 (15.4) | 5 (26.3) | ||
| Score 4 | 2 (3.4) | 2 (5.1) | 0 (0.0) | ||
TB: total bilirubin; DB: direct bilirubin; PLT: platelets; ALT: alanine transaminase; AST; aspartate aminotransferase; ALP: Alkaline phosphatase; GGT: gamma-glutamyl transferase; TP: total protein; PT: prothrombin time; IgG: Immunoglobulin G; Hb: hemoglobin; WBCs: white blood cells; SMA: smooth muscle antibody ANA: antinuclear antibody; LKM1: liver-kidney microsome 1 antibody; IQR: interquartile range; IQR: interquartile range, ∗ P value < 0.05.
Histopathological features associated with the presence of hyaline globules among children with AIH
The median AIH activity score was 15.0 (4.3), with a median Ishak score of 3.0 (2.0). The median number of plasma cells was 2.0 (2.0). Regarding the stage of fibrosis, about one-third (31.0%) of the examined biopsies had mild fibrosis, while moderate fibrosis and severe fibrosis were noted in 48.3% and in 8.6%. Active disease was diagnosed among 93.1% of the studied biopsies; mild disease activity was the most predominant, 46.6%, followed by moderate and severe, 36.1% and 10.3%, respectively. More than three-fifths (62.1%) of AIH had confluent necrosis, 86.2% had hepatocellular necrosis, and 89.7% had plasma cells. A nearly equal proportion of children with AIH had either grade 1 or grade 2 interface hepatitis or no grade. Figure 3 All the studied histopathological features of AIH were not statistically significant in-between positive and negative KCHGs children. Table 3.
Figure 3.
Liver tissue sections from different cases of AIH. (a) and (b) Hematoxylin and eosin-stained sections showing interface hepatitis (dashed arrow), plasma cell-rich infiltrate (circle), hepatocyte rosette formation (arrow) and spotty necrosis (arrow head). (c) Masson-trichrome stained liver section showing concentric fibrosis around bile duct (bd) branch (arrow). D) Orcein stained liver section showing brown coloration of old fibrosis in portal tracts and fibrous septa. No copper-associated granules were noticed. Original mag × 200.
Discussion
Compared to adults, AIH is more aggressive in children and progresses rapidly unless immunosuppressive therapy started early. Certainly, the early diagnosis might improve the outcome of the disease.16 Diagnosis of AIH is based on the exclusion of other etiologies of PCLDs that may share the same serological and histological features such as viral hepatitis B, C, and E, Wilson disease, steatohepatitis, and drug-induced liver injury.1,2,12
The histopathological features in AIH are relatively nonspecific and may be present in other forms of chronic liver injury.17,18 It has been noted that the defective function and hyperplasia of KCs may be involved in the pathogenesis of AIH.19 However, studies on AIH in this field are scarce.10,14,20 Himoto et al.20 denied the diagnostic significance of KCHGs to be specific in AIH. Recently, a modified scoring system for the diagnosis of AIH was developed by Gurung et al. 201821 in which “typical” features require both the presence of prominent plasma cells and KCHGs, while “Compatible” features included prominent plasma cells but lack KCHGs.
In this work, about two-thirds (67.0%) of children with AIH had at least one KCHG, compared to 6.0% and 12.5% among children with chronic HCV infection and WD, respectively. This finding was statistically significant across the different studied groups. We speculate that the existence of KCHGs may be a novel histopathological hallmark of AIH. In concordance with our result, Lotowska et al.22 reported that the proportion of children with AIH who were positive for KCHGs was 71.4%. The main issue in this study was that the sample size was relatively small; they studied only 14 children. In the same line, Cortes-Santiago and his colleagues23 indicated that the KCHGs were identified in 37% of children with HCV infection compared to 85% of children with interface hepatitis. Less promising result was reported by Tucker et al.13 They reported that the proportion of AIH who had KCHGs was only 50% (15/30). Interestingly, they found that none of the HCV-infected children had KCHGs (0/30), but one of thirty children with chronic hepatitis B virus infection (HBV) did.
In the current work, there was a significantly higher proportion of seropositive than seronegative AIH tested positive for the presence of KCHGs. Conversely, Tucker et al.,10 reported that there is no significant association between the seropositive and seronegative AIH and the presence of KCHGs. Noteworthy, this difference in KCHG positivity was not shown when type 1 and type 2 AIH were compared. In this study, researchers compared only five children with type 2 AIH with 34 children with type 1 AIH. We speculate that the if larger sample size is recruited in another pair-wise comparison results will be more informative.
Serum IgG levels are usually raised in both types of AIH, although a normal range of IgG is commonly observed in type 2 AIH (25%) than in type 1 AIH (15%), particularly when the disease presents acutely.4 Generally, serum IgG is used mainly as one of the criteria to diagnose AIH and to monitor treatment response. Recently, a new role of IgG was addressed by Hartel and his team24 in a large multicenter study conducted on 1318 patients with AIH. They found that patients with normal IgG levels had similar histological, biochemical, and disease severity; however, they might have a higher chance of treatment withdrawal compared to patients with high IgG. Interestingly, in this study, the IgG level among children with positive KCHGs was significantly higher than in children with negative KCHGs. In the same vein, three studies conducted by Lotowska et al.,22 Tucker et al.,13 and Cortes-Santiago et al.,23 reported that the presence of KCHGs in liver tissue of children with AIH was associated with higher serum IgG level. They suggested that these droplets had a significant immunological nature. Lotowska et al.22 considered the glassy droplet inclusions to be identical with Russel bodies of plasma cellsand these droplets may show accumulated IgG depositions and excess of immunoglobulins produced by plasma cells may be taken up and recycled by KCs [36]. On the contrary, Himoto and his colleagues20 did not report any significant association between KCHGs and the level of IgG. Consequently, they came to the conclusion that the presence of KCHGs was independent of the autoimmune process observed in AIH.
In this work, we compared the type of autoantibodies identified and the key pathological findings, including hepatic fibrosis and activity, based on whether the KCHGs were present or not. There was no significant association between KCHGs and type of autoantibodies or histopathological findings. Correspondingly, Himoto and his colleagues20 did not report any significant association between KCHGs and the histopathological findings of AIH, and they reported that the existence of KCHGs was independent of the seropositivity for ANA.
Strength and limitation of the study
One of the strong points of this study was the relatively large number of children with different PCLDs included for identification of KCHGs. The main limitation was that this study did not include children with other autoimmune liver diseases, like overlapping syndrome or children with sclerosing cholangitis, or other causes of chronic liver diseases such as HBV infection.
In conclusion, hyaline globules in Kupffer cells in liver tissues of children with AIH could be a useful diagnostic tool to distinguish AIH from other causes of pediatric chronic liver injury. Further research should be conducted to assess the feasibility of adding this feature to the diagnostic histopathological criteria of the AIH scoring system. Although the presence of KCHGs was associated with higher IgG level, it was neither associated with characteristic histopathological findings, fibrosis stage, and activity nor the presence of autoantibodies. More research is needed in the future to provide robust evidence for the function of KCHGs, as well as to investigate the association between the existence of KCHGs and response to immunosuppressive drugs.
CREDIT AUTHORSHIP CONTRIBUTION STATEMENT
All authors have contributed equally:
Mohammed Khedr: Study design, clinical management of patients, review of the literature, drafting of the article.
Nermin Adawy: patients’ recruitment and management.
Tahany Salim: clinical diagnosis and follow up of patients, review of literature, help in writing the draft of the article.
Menen Salem: patients’ recruitment and management.
Ramy Ghazy: review of literature, writing the article, and statistical analysis.
Ahmed Alharoun: data acquisition.
Mervat Sultan: histopathological examination of hyaline globules by d-PAS and interpretation of immunostaining of CD68.
Nermine Ehsan: histopathological evaluation of all cases included in the study, histopathological examination of hyaline globules in KC and writing the article.
Supportive foundation
None.
Authors contribution
All authors contributed equally to this work.
Conflicts of interest
The authors have none to declare.
Acknowledgement
Authors would like to thank Dr. Ramy Shaaban, Assistant Professor of Instructional Technology and Learning Sciences, Utah State University, Department of Surgical Services, USA, and Nancy Ali, Doctoral Candidate of Communications Media and Instructional Technology at Indiana University of Pennsylvania for proofreading and revising the manuscript.
Footnotes
Supplementary data to this article can be found online at https://doi.org/10.1016/j.jceh.2021.04.013.
Appendix A. Supplementary data
The following is the Supplementary data to this article:
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