A Rare Case of IgG4-related Disease Manifesting as an Ulcerated Gastric Cancer Complicated With Gastric Mucormycosis

IgG4-related disease (IgG4-RD) is a novel clinical entity that mimics many malignant, infectious, and inammatory disorders. Many organs or tissues are now known to be involved in IgG4-RD; however, there has been no report about IgG4-RD of stomach with fungal infection. We reported a 56-year-old male with IgG4-RD manifesting as ulcerated gastric cancer complicated with gastric mucormycosis. Both clinical and imaging manifestations were atypical, in clinical, it was highly suspected that the disease is a malignant tumor as ulcerated gastric cancer. However, no pathological evidence of malignancy could be found in repeated biopsies and surgically excised gastric specimen. Histological examination showed ulcer and inammatory granulation with lymphocytes and plasma cells in sheets, inammation of vascular wall, formation of new brous tissue and inammatory necrosis with hyphae and spores. The plasma cells had immunohistochemically strong staining for CD38, CD138, kappa, lambda, IgG and IgG4. And notably, IgG4/IgG positive plasma cell ratio was more than 40%, kappa/lambda positive staining ratio was approximately 1:1. Gastric mucormycosis was diagnosed with hyphae and spores conrmed by Gomori's methenamine silver staining and immunouorescence and confocal laser-scanning microscopy. IgG4-RD manifesting as ulcerated gastric cancer complicated with gastric mucormycosis was diagnosed by multidisciplinary team discussion from gastroenterology, radiology and pathology department. The patient vomited blood three times for the following month. An emergent exploratory laparotomy was conducted, and a mass with 10×9×8cm adhered closely to adjacent organs could be touched in lesser curvature of gastric antrum. Anastomotic leakage occurred after the surgery, symptomatic treatments (negative pressure drainage and antibiotics) were applied without steroids and anti-fungus drugs. The patient gradually recovered and was advised to consult gastroenterology department for further steroids and anti-fungus drugs treatment. PPI: proton pump IHC: immunohistochemistry; ESR: erythrocyte sedimentation rate; ENA: extractable nuclear antigen; ANA: antinuclear antibody; ANCA: anti-neutrophil cytoplasmic antibodies; PR3: anti-protease 3; MPO: anti-myeloperoxidase; GBM: glomerular basement membrane antibody; PCR: polymerase chain reaction; AIP: autoimmune pancreatitis; PBP: plasminogen-binding protein; UBR2: ubiquitin-protein ligase E3


Abstract Background
IgG4-related disease (IgG4-RD) is a novel clinical entity that mimics many malignant, infectious, and in ammatory disorders. Many organs or tissues are now known to be involved in IgG4-RD; however, there has been no report about IgG4-RD of stomach with fungal infection.

Case presentation
We reported a 56-year-old male with IgG4-RD manifesting as ulcerated gastric cancer complicated with gastric mucormycosis. Both clinical and imaging manifestations were atypical, in clinical, it was highly suspected that the disease is a malignant tumor as ulcerated gastric cancer. However, no pathological evidence of malignancy could be found in repeated biopsies and surgically excised gastric specimen.
Histological examination showed ulcer and in ammatory granulation with lymphocytes and plasma cells in sheets, in ammation of vascular wall, formation of new brous tissue and in ammatory necrosis with hyphae and spores. The plasma cells had immunohistochemically strong staining for CD38, CD138, kappa, lambda, IgG and IgG4. And notably, IgG4/IgG positive plasma cell ratio was more than 40%, kappa/lambda positive staining ratio was approximately 1:1. Gastric mucormycosis was diagnosed with hyphae and spores con rmed by Gomori's methenamine silver staining and immuno uorescence and confocal laser-scanning microscopy. IgG4-RD manifesting as ulcerated gastric cancer complicated with gastric mucormycosis was diagnosed by multidisciplinary team discussion from gastroenterology, radiology and pathology department. The patient vomited blood three times for the following month. An emergent exploratory laparotomy was conducted, and a mass with 10×9×8cm adhered closely to adjacent organs could be touched in lesser curvature of gastric antrum. Anastomotic leakage occurred after the surgery, symptomatic treatments (negative pressure drainage and antibiotics) were applied without steroids and anti-fungus drugs. The patient gradually recovered and was advised to consult gastroenterology department for further steroids and anti-fungus drugs treatment.

Conclusions
To our knowledge, it is the rst report about IgG4-related disease manifesting a gastric ulcer with mucormycosis infection.

Highlights
First report about the diagnosed as an IgG4-related disease manifesting as ulcerated gastric cancers complicated with gastric mucormycosis Systematically reviewed pathological features in biopsy and surgical specimen of IgG4-related disease Multidisciplinary team discussion is necessary and indispensable for the diagnosis Page 3/13 Background IgG4-related disease (IgG4-RD) is a chronic multi-organ-involved broin ammatory condition characterized by tendency to form tumefactive lesions, increased serum levels of IgG4 (often but not always) and tissue in ltration by dense lymphocytes and IgG4-positive plasma cells. IgG4-RD is always confused with neoplasm, infectious and in ammatory disorders clinically and radiologically [1,2] . Many organs and tissues are now known to be involved in IgG4-RD including pancreas, biliary duct, lacrimal/salivary glands, lymph node, lung, liver, kidney, thyroid gland, gastrointestinal tract, prostate, hypophysis, stomach, skin, aorta, joint, retroperitoneum, meninges and pleura [2][3][4][5][6] . To our knowledge, there has been no report about IgG4-RD of stomach with fungal infection. Here we report a case of IgG4-RD manifesting as an ulcerated gastric cancer complicated with gastric mucormycosis. expression of CK in normal gastric epithelial cells, but no tumor cell was found again. Worthy to mention was that IgG4/IgG was more than 40%. Mucormycosis was diagnosed with hyphae and spores con rmed by Gomori's methenamine silver staining ( Fig. 5A and Fig. 5B) and immuno uorescence and confocal laser-scanning microscopy (Fig. 5C). Due to the suspicions of lymphoma in clinical and image, we further completed gene rearrangement of B lymphocytes, and the result was negative. Therefore, it was highly suspected for this patient of IgG4-RD presenting as an atypical gastric ulcer complicated with gastric mucormycosis through the multidisciplinary team discussion.

Presentation Of Cases
The patient vomited blood three times for the following month (dark-red, approximately 150mL, 10mL and 70mL respectively). An emergent exploratory laparotomy was conducted, and a mass with 10×9×8cm adhered closely to adjacent organs could be touched in lesser curvature of gastric antrum ( Supplementary Fig. 1A and 1B). The gastrectomy, abdominal lymph node biopsy and jejunal nutrition tube placement were then conducted.
Pathological examination showed the in ammatory granulation with lymphocytes and plasma cells in sheets ( Supplementary Fig. 1C), in ammation of vascular wall ( Supplementary Fig. 1D), formation of new brous tissue (Supplementary Fig. 1E) and multinuclear giant cells (Supplementary Fig. 1F). The plasma cells had immunohistochemically strong staining for CD38 ( Supplementary Fig. 1G), CD138 ( Supplementary Fig. 1H), kappa (Supplementary Fig. 1I), lambda (Supplementary Fig. 1J), IgG ( Supplementary Fig. 1K) and IgG4 (Supplementary Fig. 1L). And the IgG4/ IgG ratio was more than 40%; kappa/lambda ratio was approximately 1:1; Ki-67 index was 70%. Tuberculosis was negative by polymerase chain reaction (PCR). Anastomotic leakage occurred after the surgery, treated with negative pressure drainage and antibiotics (Meropenem, following Levo oxacin) for a month. During this period, no steroids and anti-fungus drugs were used. The latest laboratory tests revealed IgG4 22.4g/L one month after the surgery. The patient was advised to consult gastroenterology department for further steroids and anti-fungus drugs treatment.

Discussion
IgG4-RD is a systemic disorder characterized by tendency to form tumefactive lesions, elevated levels of serum IgG4 (often but not always) and tissue in ltration by dense lymphocytes and IgG4-positive plasma cells in the affected organs. Autoimmune pancreatitis (AIP) is one of the most common disease features. Terumi Kamisawa and his colleague rstly proposed that AIP was not simply pancreatitis but that it was a pancreatic lesion involved in IgG4-RD with extensive organ involvement, and they de ned IgG4-RD as a new clinicopathological entity in 2003 [7] . Actually, IgG4-RD has been described in virtually every organ system including pancreas, biliary duct, lacrimal/salivary glands, lymph node, lung, liver, kidney, thyroid gland, gastrointestinal tract, prostate, hypophysis, stomach, skin, aorta, joint, retroperitoneum, meninges and pleura [2][3][4][5][6] .
It is widely believed that hereditary susceptibility, autoimmunity and infectious agents are potential factors to lead to IgG4-RD [5] . Shigeyuki Kawa and his colleagues show that the frequencies of DRB1*0405 and DQB1*0401 are signi cantly higher in patients with AIP compared with chronic calcifying pancreatitis [8] . Their group also reveals that the HLA-linked genetic basis for AIP seems to be controlled by the DRB1*0405-DQB1*0401 mini-haplotype and the ABCF1 gene area [9] . Besides, plasminogen-binding protein (PBP) of Helicobacter pylori and ubiquitin-protein ligase E3 component nrecognin 2 (UBR2) expressed by acinar cells of the pancreas are homologous in peptide. 94% patients with autoimmune pancreatitis (33/35) can be detected antibodies against the PBP peptide, while can be detected by 5% patients with pancreatic cancer (5/110), indicating that infectious factors and molecular mimicry may initiate IgG4-RD [10] . Moreover, the innate and the adaptive immune system might synergize to promote the progression of IgG4-RD and related tissue or organ brosis [11] . The microenvironment disorders are of critical importance for the pathogenesis of IgG4-RD, including immune cells including regulatory T cell, regulatory B cell, T follicular helper 2 cell and M2 macrophage [12][13][14] . In ammatory cytokines also participate in the IgG4-RD such as IFN-γ, IL-4, IL-10, IL-5, IL-13 and TGF-β [11] . In fact, there is no speci c pathogenic factors for IgG4-RD and its cognitive process is dynamic. Although genetic factor, infection and autoimmunity disorder may be involved in the pathogenesis of IgG4-RD, every hypothesis alone is not su cient to explain it.
Since the incidence of IgG4-RD is quite low with estimation of 1.4 cases per 100,000 people (statistics from Japan) [15] , and sometimes diagnosis may be missed or erroneous. The multi-disciplinary involved method may be valuable for the diagnosis of IgG4-RD. There are no speci c clinical symptoms or signs for IgG4-RD, usually common in other diseases or exerting symptoms of complications. Abdominal pain, nausea, vomiting and jaundice are more common in males, whereas swelling of the lacrimal glands and submandibular glands occurs more often in females. In males, the most commonly involved organs are salivary glands, pancreas and lacrimal glands, while they are salivary glands, lacrimal glands and sinuses in female [16] . The serum IgG4 level is elevated in most patients with increased ESR, CRP, serum IgG level and total IgE level, but there are still some patients presenting normal serum IgG4 levels (largely varied in different researches, from 2.5-40%) [17,18] . However, increased IgG4 level may occur in other diseases such as allergy and lymphoma. Organ enlargements of involved tissues are the most common imaging ndings. If lungs are involved, honey-combing or ground-glass opaci cation, pulmonary nodules, bronchovascular bundle thickening may be found [17] . Both clinical and imaging manifestations are atypical, to some extent, which are likely to be misleading and confusing, especially for single or infrequent involved organ. Pathological examination is necessary and indispensible for the diagnosis of IgG4-RD. The typical microscopic ndings are diffused lymphoplasmacytic in ltration with irregular and storiform brosis, and obliterative phlebitis, the former two of which occurs in most cases. Phlebitis without obliteration of the lumen and elevated numbers of eosinophils also exist sometimes, but no sensitive or speci c effects for the diagnosis of IgG4-RD. Necrosis, discrete granuloma and extensive neutrophilic in ltration are the important clues to exclude IgG4-RD. Immunohistochemically, staining for IgG and IgG4 is very important. The absolute number of IgG4 positive plasma cells should reach a certain standard in corresponding organs or tissues, moreover, IgG4/IgG positive plasma cell ratio should be more than 40% [18] . Although pathological examination is quite important for the diagnosis of IgG4-RD, it can't be con rmed by microscopy alone; multidisciplinary collaboration and comprehensive consideration with clinical and imaging manifestations along with laboratory tests is really important especially for single or infrequent involved organ.
To our knowledge, IgG4-RD rarely involves stomach. In the few reports available, almost every case is presenting multiple organs involvement accompanied by stomach related, and no case with fungal infection. We summarize all literature on the gastric involvement of IgG4-RD (Supplementary Table 1)[]. In the above table, the majority of cases are multiple organs involved or con rmed by surgical specimen; no patient associates fungal infection. Microscopically, there are four zones in the base of the ulcer of surgical specimen: in ammatory exudates, brinoid necrosis, granulation tissue and mature brous tissue. However, pathologists always accept biopsy tissues surrounding the base of the ulcer in the clinical practice for fear that gastric perforation and unnecessary operation, which undoubtedly brings bene t for patients. At the same time, biopsy specimen make diagnosis for IgG4-RD involving stomach di cult, especially presenting with ulcer lesion. Pathologists must build up general impression based on the local ndings. In this case, fungal infection makes diagnosis more confusing and challenging. Gastric mucormycosis may cause secondary changes for ulcer under microscope; besides, the identi cation of speci c fungus will in uence clinical decision and treatment methods.

Conclusion
As far as we know, it is the rst report about IgG4-related disease manifesting as an ulcerated gastric cancer complicated with gastric mucormycosis. Multidisciplinary team discussion from gastroenterology, radiology and pathology department is necessary and indispensable for the disease diagnosis.    The typical IHC staining for this case (biopsy specimen). The plasma cells had immunohistochemically strong staining for CD38 (A, 100x), CD138 (B, 100x), kappa (C, 100x), lambda (D, 100x), IgG (E, 100x), and IgG4 (F, 100x). The typical images for hyphae and spores of mucormycosis of this case. Gomori's methenamine silver staining for hyphae and spores of mucormycosis (A, 100x, B, 400x). Immuno uorescence and confocal laser-scanning microscopy were used to characterize hyphae and spores of mucormycosis (C, 200x).