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Japanese Journal of Clinical Oncology Pages 436-440

Interleukin-1[alpha] Producing Synovial Sarcoma with Prolonged Fever: a Case Report
Introduction
Case Report
Materials And Methods
Results
   Pathological Findings
   RT-PCR and Sequence of IL-1[alpha]
Discussion
References
Interleukin-1[alpha] Producing Synovial Sarcoma with Prolonged Fever: a Case Report

Interleukin-1[alpha] Producing Synovial Sarcoma with Prolonged Fever: a Case Report

Shunzo Osaka1, Yukiko Fujimoto1, Hideoki Yamazaki1, Koyu Suzuki2, Shigemasa Sawada3 and Nakanobu Hayashi4

1Department of Orthopedic Surgery, 2Department of Pathology and 3Department of Internal Medicine, Nihon University School of Medicine and 4Omgen Inc., Tokyo, Japan

The patient had a synovial sarcoma of the monophasic fibrous type accompanied by prolonged spike fever. Wide excision of the tumor resulted in the disappearance of her fever. The tumor cells showed interleukin-1[alpha] (IL-1[alpha]) expression immunohistochemically. IL-1[alpha] cDNA was detected in the tumor by RT-PCR, the sequences of which were identical with those of normal human IL-1[alpha] molecules. These results indicated that IL-1[alpha], one of the fever-inducing cytokines, was expressed by sarcoma cells in the present case, which were thought to be a causative factor of the fever.

Key words: synovial sarcoma - fever - interleukin-1[alpha] - reverse transcriptase polymerase chain reaction

INTRODUCTION

Fever in mesenchymal tumors, especially in soft tissue sarcomas, is a relatively rare event. Some smooth muscle tumors have been reported to be accompanied by fever of unknown origin (1-3). Recently, cytokines such as interleukin-1[alpha] (IL-1[alpha]), interleukin-1[beta] (IL-1[beta]), interferon-gamma (IFN-[gamma]), interleukin-6 (IL-6) and macrophage inflammatory protein-1 (MIP-1) have been shown to be related to fever (4-7). These cytokines are released in various conditions by lymphocytes, macrophages and other sources, resulting in the development of fever (8,9). So-called tumor-related fevers are observed fairly commonly in leukemias, lymphomas and carcinomas. It is suspected that tumor cells may produce cytokines which cause fever in patients (10-12). Here, we report a case of synovial sarcoma accompanied by a prolonged fever which was thought to be associated with the expression of IL-1[alpha] by tumor cells.

Case Report

The patient, a 66-year-old female, who came to our hospital on December 12, 1996, noticed a mass in the right proximal thigh. A CT scan and MRI showed a large tumor (9 × 12 cm) in the tensor fascia lata and gluteus medius muscle (Fig. 1). An angiogram showed a vascular mass of tumor stain. A needle biopsy revealed a high-grade spindle cell sarcomatous tumor. Data on admission were Hb 10.0 g/dl, Ht 29.7%, platelets 561 × 103/µl, CRP 13.4 mg/dl and Fe 35 µg/dl. Thyroid tests and tumor markers (CEA, AFP and others) were normal. Gastroscopic and colonoscopic examinations, radiographs of the chest, bone marrow and genital examinations were normal. Neither thyroid enlargement nor lymphadenopathy was noted. Blood cultures were repeatedly negative. The patient had a spiking fever. She received a preoperative arterial infusion of 100 mg of cisplatin. The tumor continued to grow, so chemotherapy was stopped. The patient received six units of blood before surgery. A wide excision including the greater trachanter and the capsule of the hip joint was performed and reconstructed with an anterolateral thigh flap and autobone graft. The patient's fever disappeared immediately after surgery. Six months follow-up showed no recurrence of the tumor or the fever.


Figure 1. CT scan revealed a mass of low density (A) and MRI in the T2-weighted mode a mass of high intensity (B).

MATERIALS AND METHODS

The resected tissues were fixed with 10% buffered formalin, routinely processed and embedded in paraffin. Sections were examined with hematoxylin and eosin and immunohistochemistry. The latter was carried out with the antibodies listed in Table 1, with both the avidin-biotin peroxidase complex (ABC) technique with an ABC kit (Vector Laboratories, Burlingame, CA) and also with the indirect immunoperoxidase method (13) using formalin-fixed, paraffin-embedded tissue sections or frozen sections, respectively. Appropriate positive and negative controls were used for all antibodies.

Table 1. Results of immunohistochemical study
Antibody Monoclonal/polyclonal Dilution Source* Methods[dagger] Results[Dagger]
Cytokeratin M 1:200 Lipshaw ABC +
Epithelial membrane antigen (EMA) M 1:100 Nichirei ABC +
Vimentin M 1:200 Nichirei ABC +++
S 100 protein (S-100) P 1:500 Biomeda ABC ++
Desmin M 1:100 Nichirei ABC -
Smooth muscle specific actin M 1:100 Dakopatts ABC -
CD34 M 1:50 Nichirei ABC -
IL-1[alpha] M 1:20 Oncogene Science IP +++
IL-1[beta] M 1:20 Oncogene Science IP -
IL-6 M 1:20 Oncogene Science IP -
TNF-[alpha] M 1:20 Oncogene Science IP -
*Lipshaw, Pittsburgh, PA; Nichirei, Tokyo, Japan; Biomeda, Foster City, CA; Dakopatts, Glostrup, Denmark; Oncogene Science, Cambridge, MA. [dagger]ABC, avidin-biotin peroxidase complex technique; IP, indirect immunoperoxidase method. [Dagger]+++, many tumor cells stained; ++, some tumor cells stained; +, few tumor cells stained; -, no tumor cells stained.

Small tissue blocks from the formalin-fixed specimen were submitted for ultrastructural examination. They were fixed with 2% osmium tetraoxide and embedded in Epon 812. Ultrathin sections were cut and stained with uranyl acetate and lead citrate and observed with a Hitachi H-800 electron microscope (Hitachi, Tokyo, Japan).


Figure 2. Spindle tumor cells are arranged in fascicles. Mitoses are seen. H & E, original magnification ×400.


Figure 3. Immunostaining of IL-1[alpha]. Tumor cells show positive staining in the cytoplasm. Original magnification ×400.

Total RNA was extracted from snap-frozen tissue as described previously (14). The reverse transcriptase polymerase chain reaction (RT-PCR) was performed using nested primer sets. First and second primer sets were 5[prime]-CATTTCATTGGCGTTTGAGTCAGC-3[prime], 5[prime]-AAATCGAAGAAGTACATGTACCAT-3[prime] and 5[prime]-AGAAGTCAAGATGGCCAAAGTTCC-3[prime], 5[prime]-GTATACTTGACAGTTGTGACGTGT-3[prime], respectively. The primer sequences were based on the IL-1[alpha] sequences in the gene bank library (accession No. M28983). The amplified cDNA fragments (874 bp) were analyzed by electrophoresis in a 2% agarose gel. The PCR products were subcloned into p-GEM-T vectors. Positive clones which had an insert of the correct size were subjected to sequence examination (ABI 377 sequencer).

RESULTS

Pathological Findings


Figure 4. Ultrastructure shows microvilli (A) ×5000 and inclusion bodies in the cytoplasm of the tumor cells (B) ×5000.

The resected specimen consisted of a solid gray-white tumor measuring 11.5 × 6 × 10 cm, with central necrosis, attached to the muscles of the thigh. The tumor showed fascicles of spindle cells having spindle to ovoid nuclei and a moderate amount of pale eosinophilic cytoplasm (Fig. 2). The cells were relatively uniform in size and shape. The cellularity was high and mitoses were frequent (more than five mitoses/10 high power fields). The tumor cells were diffusely and strongly positive for vimentin S-100, which was detected in a moderate number of cells. EMA and cytokeratin were stained in a small number of cells. IL-1[alpha] was clearly stained in the cytoplasm of the tumor cells (Fig. 3). Other cytokines were negative. Ultrastructually, there were microvilli and inclusion bodies in the cytoplasm of the tumor cells (Fig. 4). A diagnosis of synovial sarcoma, monophasic fibrous type, was made, based on the histopathology and results of immunohistochemistry and electron microscopy.


RT-PCR and Sequence of IL-1[alpha]

RT-PCR products were mobilized at 874 base pairs on the gel corresponding to the IL-1[alpha] fragment (Fig. 5). Nucleotide sequences of 10 positive clones were identical. They were completely homologous with those seen in the normal human IL-1[alpha] cDNA sequence.


Figure 5. Identification of IL-1[alpha] by RT-PCR. M = 100 bp DNA marker (Gibco BRL). Lane 1: amplification product of 874 bp corresponds to IL-1[alpha].

DISCUSSION

Malignant cases with prolonged fever have been reported for over 30 years, especially in hematological malignancies and carcinomas (11,15). In mesenchymal tumors, fever is a relatively rare event and, so far, only a few cases have been reported of neuroblastoma (16) and leiomyosarcoma (3,17,18). Irrespective of tumor histogenesis, however, the mechanism and causative factors of these fevers were usually undetermined. In the present case, the patient manifested a prominent prolonged fever with a spiking characteristic. The tumor was diagnosed as a synovial sarcoma of monophasic spindle cell type, backed up with the findings of histopathological, immunohistochemical and ultrastructural analyses. Hence the present case was revealed to be an example of tumor-related fever in a malignant mesenchymal tumor. The fever disappeared following resection of the tumor, suggesting that the tumor or sarcoma cells produced substances which caused the fever.

Recently, it has been reported that the induction of fever and anemia (18,19) was closely related to cytokines such as IL-1[alpha], IL-1[beta], IL-6, TNF, IFN-[alpha], IFN-[beta] and IFN-[gamma] (4-7). Studies have indicated a strong relationship between the malignant tumor and release of these cytokines. For example, glioblastoma (20) and osteosarcoma (21) produced IL-6, which was enhanced by TNF and IL-1[beta].

Among the cytokines, IL-1 is thought to be one of the strongest factors in fever development through the reaction of prostaglandin, as was observed in animal experiments (22,23). Further, IL-1 is a polypeptide that has a wide variety of action such as being a mitogen for cultured fibroblasts (9), smooth muscle cells (24), endothelial growth (25), promoting keratinocyte differentiation, inhibiting erythropoiesis (26) and production during 180-sarcoma growth in rats (27). However, this was not observed in human sarcomas. These are usually observed in inflammation and immunity (5,28). In the present case, IL-1[alpha] was abundantly present in the sarcoma cells, as demonstrated by immunohistochemistry, although IL-1[beta], IL-6 and TNF-[alpha] were negative. RT-PCR revealed the expression of mRNA of IL-1[alpha], which was thought to be consistent with the production of this cytokine in the tumor. The fever disappreared after the excision of the sarcoma. This indicated that IL-1 was produced by sarcoma cells and was released in the blood, causing the patient to become febrile. Cloning of the IL-1[alpha] demonstrated that the IL-1[alpha] sequences examined were identical with those seen normally in humans. Therefore, the IL-1[alpha] molecules observed in the present sarcoma were probably intact. It is speculated that the expression of IL-1[alpha] was enhanced by an alteration of the regulation of IL-1[alpha] production in the tumor cells. The tumor may have produced other fever-stimulating factors in addition to IL-1[alpha].

In summary, the present case showed an IL-1[alpha] producing synovial sarcoma in which prolonged fever was probably caused by the over-production of IL-1[alpha] by the tumor cells. It is suggested that cytokine examinations may be of value in tumor cases presenting stubborn fever.

References

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Received January 30, 1998; accepted April 24, 1998
For reprints and all correspondence: Shunzo Osaka, Department of Orthopaedic Surgery, Nihon University Nerimahikarigaoka Hospital, 2-11-1, Hikarigaoka, Nerima-ku, Tokyo 179, Japan
Abbreviations: IL-1[alpha], interleukin-1[alpha]; IFN-G, interferon-gamma; MIP-1, macrophage inflammatory protein-1; ABC, avidin-biotin peroxidase complex; RT-PCR, reverse transcriptase polymerase chain reaction.

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