Detection of mRNAs for urokinase-type plasminogen activator, its receptor, and type 1 inhibitor in giant cell tumors of bone with in situ hybridization.

Although giant cell tumor of bone (GCT) is generally considered to be an uncommon benign neoplasm, it can pursue an aggressive course with local recurrence and metastasis. Attempts to predict the biological behavior of GCT with histopathological parameters, however, have not been successful. The urokinase-type plasminogen activation system has been implicated in tumor invasion and metastasis and abnormalities of the components of this system have been found in several malignancies. In this study we postulated that the urokinase-type plasminogen activation system associated with bone destruction and local invasion is present in GCT. We therefore evaluated the mRNA levels for urokinase-type plasminogen activator (u-PA), urokinase-type plasminogen activator receptor (u-PAR), and plasminogen activator inhibitor type 1 (PAI-1) by using Northern blot analysis and in situ hybridization in four cases of GCT and spindle-shaped mononuclear cells at the 35th passage from a GCT. Our results showed that giant cell tumors of bone contained variable levels of u-PA, u-PAR, and PAI-1 mRNA, respectively, 2.3, 1.4, and 3.2 kb in size. In situ hybridization showed that u-PA, u-PAR, and PAI-1 mRNA were expressed in both the mononuclear cells and the osteoclast-like giant cells; the signal for u-PA mRNA in the spindle-shaped mononuclear cells was more intense than that in the osteoclast-like multinuclear giant cells. Some spherical mononuclear cells (macrophage-like cells) expressed high levels of PAI-1 mRNA in comparison with the spindle-shaped mononuclear cells. In addition, the 35th passaged spindle-shaped mononuclear cells were used to study the gene expression of u-PA during cell proliferation. The results showed that the level of u-PA mRNA increases after adding 10% fetal calf serum to quiescent cells. The induction was maximal at 16 hours and remained high during 48 hours of treatment. In conclusion, even though osteoclast-like cells are ultimately responsible for the bone resorption of GCT, the mononuclear neoplastic cells of GCT may also be involved in degradation of the extracellular matrix during invasive growth by facilitating the urokinase plasminogen activation system. In addition, our observation of upregulation of u-PA mRNA in spindle-shaped mononuclear cells after serum stimulation indicated that u-PA production may be linked to tumor growth.     

Hepatocellular carcinoma with osteoclast-like giant cells: possibility of osteoclastogenesis by hepatocyte-derived cells

Giant cell tumor (GCT) of bone is a primary osteolytic tumor that is characterized by the formation of osteoclast-like giant cells. In addition to GCT of bone, extraskeletal GCT are known to be formed in several soft tissues. Giant cells in GCT of bone were suggested to be identical to osteoclasts, but the characterization of giant cells in extraskeletal GCT remains incomplete. In this study, a case of sarcomatoid hepatocellular carcinoma with osteoclast-like giant cells was analyzed. Immunohistochemistry revealed the expression of almost all markers of osteoclasts: tartrate-resistant acid phosphatase, CD68, CD51, CD54 and matrix metalloprotease-9, in osteoclast-like giant cells in the tumor. In situ hybridization revealed the expression of receptor activator of nuclear factor-kappa B (RANK) in the giant cells and receptor activator of nuclear factor-kappa B ligand (RANKL) in the tumor cells. The hepatic origin of the sarcomatoid hepatocellular carcinoma cells was confirmed by the expression of albumin. This is the first report suggesting that hepatocyte-derived cells possess the potential for osteoclastogenesis. In addition, these findings suggest that osteoclast-like cells in the hepatocellular carcinoma were formed by the same mechanism as osteoclastogenesis in bone.     

Giant cell tumor of the skin: a morphologic and immunohistochemical study of five cases

Giant cell tumor (GCT) of the skin is a rare entity that possesses similar gross and histologic features to GCT of bone. When located predominantly in the dermis GCT has been mistaken for benign fibrous histiocytoma and atypical fibroxanthoma. We report the clinical, morphologic, and immunohistochemical features of five cases of GCT of the skin. With one exception, all tumors are confined to the dermis. Patients' ages range from 6 to 78 years (median, 73 years) with a male to female ratio of 3:2. Gross and histologic features of the lesions are similar to those of GCT of bone (eg, brown fleshy tumor and a biphasic population of mononuclear cells admixed with osteoclast-like giant cells, respectively). The nuclei of the giant cells are similar to those of the mononuclear cells. A fascicular pattern with focal storiform arrangement of spindle neoplastic cells is noted in two cases. The osteoclast-like giant cells and some of the mononuclear cells are strongly positive for CD68, alpha-1-antitrypsin, and alpha-1-antichymotrypsin. Only the mononuclear cells express smooth muscle actin focally in one case. Both the osteoclast-like giant cells and the mononuclear cells are negative for cytokeratins (AE1/AE3 and CAM5.2) and S-100 protein in all cases. One patient developed lung metastases at presentation and local recurrence 4 months status post surgery. All patients are without evidence of disease 1 month to 12 years status post surgery. Cutaneous GCTs are low-grade sarcomas that can recur locally and infrequently metastasize. These tumors should be distinguished from a variety of cutaneous neoplasms that contain multinucleated giant cells.     

Giant cell tumor of the lung: An autopsy case report with immunohistochemical observations

Tumors resembling giant cell tumor (GCT) of bone are well known to occur in other organs and many cases have been reported to date. While GCT occurring as primary lesions in the lung are extremely rare, the authors experienced such a tumor at an autopsy of a 77 year old woman and subsequently performed histological and immunohistochemical examinations. The clinical and morphologic characteristics of this case are documented, and the literature concerning this type of tumor is reviewed. The present tumor of the lung was histologically characterized by proliferation of benign-looking osteoclast-like giant cells in association with slightly atypical mononuclear cells. The tumor cells were immunohistochemically positive for histlocytic markers but negative for epithelial markers. This case appears to be the first reported benign giant cell tumor of the lung in which histiocytic differentiation of mononuclear cells was suggested by immunohistochemistry.     

Primary giant cell tumor of soft tissues similar to bone giant cell tumor: A case report and literature review

In this report we describe a primary giant cell tumor (GCT) of soft tissues located in the left dorsal wrist of a 52-year-old man. Plain radiographs did not reveal any lesion in his carpal or hand bones. Although the tumor was clinically considered a ganglion initially, the microscopic features were identical to those found in classic GCT of bone. Light microscopy showed a lesion composed of a homogeneously mixed proliferation of spindle and polygonal mononucleated stromal cells and evenly distributed multinucleated, osteoclast-like giant cells. Whereas most bone tumors have an extraosseous counterpart, only 13 cases of GCT in soft tissues had been published until 1998. Moreover, 64 new cases have been reported in three series. Nevertheless, most major reviews and textbooks do not consider this tumor as a specific entity and regard it as a low grade variant of malignant GCT of soft tissue. We describe the clinical, histologic, and immunohistochemical features of this rare benign neoplasm emphasizing the differential diagnosis with its malignant soft tissue counterpart, an ominous tumor.     

Histochemistry of tartrate-resistant acid phosphatase and carbonic anhydrase isoenzyme II in osteoclast-like giant cells in bone tumours

Summary Using routinely processed, paraffin-embedded tissue specimens, osteoclast-like giant cells in giant cell tumour of bone (GCT), chondroblastoma, osteoblastoma and osteoblastic osteosarcoma were examined histochemically for osteoclast-specific enzymes tartrateresistant acid phosphatase (TRAP) and carbonic anhydrase isoenzyme II (CA-II). Osteoclast-like giant cells and some mononuclear cells possessed TRAP activity. These were further classified with respect to CA-II immunoreactivity, i.e. cells with CA-II were seen in GCT and chondroblastoma, while those in osteoblastoma and osteoblastic osteosarcoma were negative for CA-II. All the cellular components in malignant fibrous histiocytoma and various extraosseous inflammatory lesions including malignant giant cells and macrophage polykaryons were negative for both TRAP and CA-II. These results indicate that osteoclast-like giant cells in GCT, chondroblastoma, osteoblastoma and osteoblastic osteosarcoma are all osteoclasts and generated by fusion of mononuclear cells with the same histochemical characteristics as osteoclast-like giant cells. The difference in CA-II immunoreactivity suggests the functional or maturational difference between osteoclast-like giant cells in GCT and chondroblastoma and those in osteoblastoma and osteosarcoma.     

Clear cell sarcoma-like tumor with osteoclast-like giant cells in the small bowel: further evidence for a new tumor entity

Most mesenchymal neoplasms of the gastrointestinal tract belong to the category of gastrointestinal stromal tumors (GISTs) and are characterized by the immunohistochemical expression of KIT receptor. In cases without detectable KIT receptor expression several differential diagnoses have to be taken into consideration. Here, we report a case of a 41-year-old man with a tumor of the small bowel composed of large epithelioid tumor cells arranged in solid and alveolar sheets including scattered osteoclast-like multinucleated giant cells. Immunohistochemically, the tumor cells expressed strongly S-100 protein, vimentin, and to a lesser extent, bcl-2. HMB-45, melan-A, KIT receptor, desmin, smooth-muscle actin, and CD-34 were not detectable. Ki-67 index was 20%. The diagnosis was established by 2 different FISH strategies demostrating the presence of a t(12;22)(q13;q12) translocation, the diagnostic hallmark of clear cell sarcoma of soft parts. Our results provide further evidence for the existence of a new tumor entity designated gastrointestinal clear cell sarcoma with osteoclast-like giant cells. The diagnosis of this entity should be considered in the presence of S-100-positive tumors of the gastrointestinal tract containing multinucleated giant cells and can be established by FISH analysis.     

骨巨细胞瘤TIMP-3启动子甲基化的研究

目的 检测骨巨细胞瘤(GCT)中TIMP—3启动子甲基化及其蛋白表达，探讨该肿瘤组织TIMP—3蛋白表达缺失的原因和TIMP-3启动子甲基化与GCT分级和复发的关系。方法 用免疫组织化学SP法和蛋白免疫印迹检测TIMP—3在GCT组织中的表达，用甲基化特异的PCR(MSP)法检测TIMP—3基因启动子的甲基化状态。结果 TIMP—3主要在单核基质细胞和多核巨细胞的胞质表达，后者的表达具有明显的极向性。17例GCT中有5例(29．4％)TIMP—3蛋白丢失，其中4例的TIMP—3启动子发生异常甲基化，且均为组织学分级为Ⅱ级的病例。结论 GCT的局部骨质破坏，可能与TIMP—3丢失有关；而TIMP—3启动子的异常甲基化，是TIMP—3基因失活和蛋白丢失的重要机制。     

Histogenesis of giant cell tumors

The giant cell tumor of bone (GCT) is a local osteolytic tumor with variable degrees of aggressiveness. In rare cases pulmonary metastases can be observed. The lesion most frequently occurs in the epiphysis of long tubular bones of the knee region, predominantly affecting young adults after closure of the growth plate. The characteristic histological appearance of GCT displays a high number of osteoclast-like multinucleated giant cells, which resulted in the classification "osteoclastoma" or "giant cell tumor". Apart from the multinucleated giant cells, there are two mononuclear cell types in the GCT. The first one has a round morphology and resembles a monocyte. The second cell type is the spindle-shaped, fibroblast-like stromal cell. Cell culture experiments with GCT cells revealed the stromal cell to be the proliferating component of the GCT. The other two cell types, the monocyte and the multinucleated giant cell, were lost after a few cell culture passages. Furthermore, latest results from GCT reveal that the stromal cells secrete a variety of cytokines and differentiation factors, including MCP1, ODF and M-CSF. These molecules are monocyte chemoattractants and are essential for osteoclast differentiation, suggesting that the stromal cell stimulates blood monocyte immigration into tumor tissue and enhances their fusion into osteoclast-like, multinucleated giant cells. The multinucleated giant cell itself demonstrates properties of a normal osteoclast that is able to resorb bone leading to extended osteolysis. This new model of GCT genesis supports the hypothesis that the stromal cell is the neoplastic component whilst the monocytes and the multinucleated giant cells are just a reactive component of this tumor. Taking this into consideration, the nomenclature of the "giant cell tumor" needs to be reconsidered.     

骨巨细胞瘤中PTEN基因与PCNA的表达

目的 探讨PTEN与骨巨细胞瘤中破骨样多核巨细胞的关系。方法 采用免疫组化染色方法检测27例骨巨细胞瘤PTEN蛋白与PCNA的表达、PCR检测骨巨细胞瘤PTEN基因的外显子。结果 77．8％(21／27例)骨巨细胞瘤中仅破骨样多核巨细胞PTEN蛋白呈强表达，PCNA不表达；而间质细胞基本不表达PTEN蛋白，但具有不同比例的PCNA阳性细胞。在所分析的8例骨巨细胞瘤中，PCR均检测到PTEN基因的外显子。结论 PTEN在GCT破骨样多核巨细胞的过表达可能是骨巨细胞瘤形成原因之一。     

Hepatic giant cell carcinoma. An ultrastructural and immunohistochemical study

Hepatocellular carcinoma with osteoclast-like giant cells (hepatic giant cell carcinoma [HGCC]) is a rare entity, with only three cases reported. The tumor is histologically similar to giant cell tumor (GCT) of bone, and the origin of the multinucleated giant cells and mononuclear stromal cells has not been determined. The purpose of this report is to present a case of this rare tumor and compare its ultrastructural and immunohistochemical features with those of a conventional GCT of bone. Histologically, the HGCC consists of sheets of osteoclast-like giant cells with a background of mononuclear cells. The giant cells lack the pleomorphism seen in hepatocellular carcinomas with anaplastic giant cells. At the light microscopic level, most of this tumor was nearly identical to a GCT of bone, but several microscopic fields (less than 5% of the tumor) had the histologic appearance of a "usual" hepatocellular carcinoma. The hepatic tumor was negative for HAM 56, epithelial cytokeratins, muramidase, and alpha-1-antitrypsin, with only focal positivity for chymotrypsin in mononuclear and giant cells. The GCT was strongly positive for alpha-1-antitrypsin and chymotrypsin in both the mononuclear and giant cells and showed focal, weak staining for AE1 and AE3 in the mononuclear stromal cells. Ultrastructurally, both mononuclear and giant cells of the HGCC showed features typical of hepatocellular carcinoma. Although the patient presented in this report died, the pattern of growth was different from most hepatocellular carcinomas. The overall histologic features of this tumor are distinctive and appear to justify separating this variant from other types of hepatocellular carcinoma.     

Cytochemical and Ultrastructural Changes in the Osteoclast-like Giant Cells of Giant Cell Tumor of Bone Following Bisphosphonate Administration

Giant cell tumor of bone (GCT) is a local aggressive neoplasm of bone characterized by expansive osteolytic lesions at the epiphysis of long bones. Bisphosphonates have been used to prevent bone resorption in secondary osteolytic tumors because of their strong anti-osteoclastic action. The authors studied the apoptosis and ultrastructural changes induced in osteoclast-like giant cells of GCT, following treatment with the aminobisphosphonate pamidronate in 16 patients with GCT of bone. Transmission electron microscopy (TEM) was used to identify ultrastructural changes, indicative of apoptosis, in the cytoplasm and the nucleus of the giant cells. Significant changes were observed in tumor samples from all 16 patients. In the cytoplasm these changes were characterized by abundant large tubular vesicles containing a central electrodense core scattered through the cytoplasm. In addition, mitochondria in the sections from pamidronate-treated patients appeared to be edematous when compared with sections from untreated patients. Nuclear changes in the giants cells were characterized by the formation of dense chromatin material scattered throughout the nucleus. The TUNEL labeling assay indicated that the mean pretreatment apoptotic index of 7.8% increased to 53% following pamidronate treatment. This was statistically significant ( p <.001) and correlated well with the ultrastructural changes noted on TEM. The formation of abundant tubular vesicles in giant cells following bisphosphonate treatment may reflect disturbed vesicular trafficking and may affect the bone resorbing activity of giant cells.     

Cytochemical and ultrastructural changes in the osteoclast-like giant cells of giant cell tumor of bone following bisphosphonate administration

Giant cell tumor of bone (GCT) is a local aggressive neoplasm of bone characterized by expansive osteolytic lesions at the epiphysis of long bones. Bisphosphonates have been used to prevent bone resorption in secondary osteolytic tumors because of their strong anti-osteoclastic action. The authors studied the apoptosis and ultrastructural changes induced in osteoclast-like giant cells of GCT, following treatment with the aminobisphosphonate pamidronate in 16 patients with GCT of bone. Transmission electron microscopy (TEM) was used to identify ultrastructural changes, indicative of apoptosis, in the cytoplasm and the nucleus of the giant cells. Significant changes were observed in tumor samples from all 16 patients. In the cytoplasm these changes were characterized by abundant large tubular vesicles containing a central electrodense core scattered through the cytoplasm. In addition, mitochondria in the sections from pamidronate-treated patients appeared to be edematous when compared with sections from untreated patients. Nuclear changes in the giants cells were characterized by the formation of dense chromatin material scattered throughout the nucleus. The TUNEL labeling assay indicated that the mean pretreatment apoptotic index of 7.8% increased to 53% following pamidronate treatment. This was statistically significant (p<.001) and correlated well with the ultrastructural changes noted on TEM. The formation of abundant tubular vesicles in giant cells following bisphosphonate treatment may reflect disturbed vesicular trafficking and may affect the bone resorbing activity of giant cells.     

Pancreas microenvironment promotes VEGF expression and tumor growth: novel window models for pancreatic tumor angiogenesis and microcirculation

Pancreatic cancer has a poor prognosis, and treatment strategies based on preclinical research have not succeeded in significantly extending patient survival. This failure likely stems from the general lack of information on pancreatic tumor physiology, attributable to the difficulties in developing relevant, orthotopic models that accurately reflect pancreatic cancer in the clinic. To overcome this limitation, we developed abdominal wall windows suitable for intravital microscopy that allowed us to monitor angiogenesis and microvascular function noninvasively during tumor growth in vivo. We used two complementary tumor models in mice: orthotopic (human ductal pancreatic adenocarcinoma, PANC-1, grown in the pancreas), and ectopic (PANC-1 grown in the abdominal wall). We found that orthotopic PANC-1 tumors grew faster than the ectopic tumors and exhibited metastatic spread in the late stage similar to advanced pancreatic cancer in the clinic. Orthotopic PANC-1 tumors expressed vascular endothelial growth factor (VEGF)(121) and VEGF(165), contained higher levels of tumor cell-derived VEGF protein, and maintained vascular density and hyperpermeability during exponential tumor growth. Orthotopic PANC-1 tumors showed lower leukocyte-endothelial interactions in the early stage of growth. In addition, both VEGF(121) and VEGF(165) promoted the growth of PANC-1 cells in vitro. Finally, Anti-VEGF neutralizing antibody inhibited angiogenesis and tumor growth of PANC-1 tumors in both sites. We conclude that the orthotopic pancreas microenvironment enhances VEGF expression, which stimulates growth of PANC-1 tumors (compared with ectopic tumors). The mechanism is autocrine and/or paracrine and also is involved in the maintenance of blood vessels. This comparative system of orthotopic and ectopic pancreatic cancer will provide the rigorous understanding of pancreatic tumor pathophysiology needed for development of novel therapeutic strategies.     

Giant cell tumor of bone: Frequent actin immunoreactivity in stromal tumor cells

Although giant cell tumor (GCT) of bone is a well-recognized neoplasm with distinctive clinical and histopathological features, the origin of tumor cells, particularly of mononuclear cells, has not yet been established. An immunohistochemicai study was carried out on 11 cases of GCT of bone to examine the cellular natures of stromal mononuclear cells. In all cases, stromal cells were positive for muscle actin (HHF35) or α-smooth muscle actin, and in eight of 11 cases, positivist was intense and extensive. The cell margin of osteoclast-like giant cells (OGC) was stained positively by muscle actin, In addition to Intense and diffuse positive staining of the cytoplasm for KP1 (CD68), whereas α-smooth muscle acting exhibited a negative reaction on the OGC. In conclusion, the tumor cells with muscle actin and α-smooth muscle actin proclivities are not rare but frequently numerous In the GCT of bone; whereas further observation Is necessary to elucidate whether the stromal cells exhibit myoflbroblastlc cell differentiation exactly.     

Osteoclast-rich undifferentiated carcinomas of the urinary tract.

Osteoclast-like giant-cell neoplasms of the urinary tract are rare. They are composed of ovoid or spindle-shaped mononuclear cells with evenly spaced osteoclast-like giant cells. Terminology, histogenesis, and biologic behavior of these tumors remain controversial. Six cases of osteoclast-like giant-cell neoplasms of the urinary tract were identified from the consultation files of two of the authors. Patients were all male and elderly (range 65-82), with the exception of one 39-year-old male. In all, 3/6 tumors developed in the bladder and 3/6 in the renal pelvis. Size ranged from 5 to 11 cm. One bladder and three renal pelvis tumors were high stage (pT3) at time of presentation. Adjacent to the osteoclast-like giant-cell neoplasm in the same specimen, all patients had urothelial carcinoma in situ and/or high-grade papillary urothelial carcinoma. Multinucleated cells had identical morphological and immunohistochemical properties of osteoclasts; positive for CD-68, LCA, CD51 and CD54, and negative for cytokeratins and EMA. Varying percentages of mononuclear cells expressed alpha-smooth muscle actin (4/6), desmin (1/6), S-100 (4/6), LCA (2/6) and CD68 (6/6). However, mononuclear cells were also positive for epithelial markers in 4/6 tumors (cytokeratins AE-1/AE-3, Cam 5.2, CK7 and/or EMA). p53 stained mononuclear tumor cells in three cases, paralleling the staining on the accompanying urothelial carcinoma. Ki-67 stained mononuclear tumor cells, but not osteoclast-like giant cells. Follow-up data were available in five cases. One patient developed recurrence of noninvasive urothelial carcinoma and is still alive. Four patients were dead due to disease within 15 months, three with distant metastases. The intimate association of these tumors with urothelial carcinoma along with their immunohistochemical profile supports an epithelial origin for the mononuclear cells and non-neoplastic reactive histiocytic lineage for the osteoclast-like giant cells.     

Combined hepatocellular carcinoma and osteoclast-like giant cell tumor of the liver: Possible clue to histogenesis

Hepatic giant cell tumor is extremely rare, and only five cases have been reported of overt hepatocellular carcinoma, thus its histogenesis is controversial. Herein is reported a case of simultaneous hepatocellular carcinoma and osteoclast-like giant cell tumor in a single tumor. A liver tumor was found in a 74-year-old woman. Histologically the tumor consisted of two distinct components: mononuclear and multinuclear giant cells with osteoclastic giant cells, and a conventional hepatocellular carcinoma. The boundary between the two components showed transitional features. Immunohistochemistry showed that the osteoclast-like giant cells were CD68 and vimentin positive, but cytokeratin and AFP negative, while spindle-shaped cells were positive only for vimentin. In a portion of the hepatocellular carcinoma the cells were cytokeratin-8 and AFP positive. Ki-67 positivity was 10% for the hepatocellular carcinoma, 60% for the spindle-shaped cells, and 0% for the giant cells. It is possible that the tumor might have had a hepatocellular carcinoma origin, given the more highly proliferative sarcomatous changes and reactive osteoclast-like cells. This case provides a clue to the histogenesis of hepatic giant cell tumors.     

Analysis of chromosome 12 aneuploidy in interphase cells from human male germ cell tumors by fluorescence in situ hybridization.

The i(12p) marker chromosome has been found to be a highly nonrandom chromosome abnormality associated with germ cell tumors (GCTs). We have previously shown that a chromosome 12 centromere specific alpha-satellite DNA probe detects the i(12p) by virtue of differences in the size of the signal originating from the i(12p) and normal chromosome 12 centromeres after fluorescence in situ hybridization (FISH) in metaphase and interphase cells of cultured GCT cell lines. We have now extended this analysis to 72 fresh GCT tumor biopsy specimens. Banded cytogenetic analysis was attempted on each of these tumors, 45 of which were found to be clonally abnormal. Data on i(12p) and chromosome 12 copy number obtained by FISH agreed well with those obtained by cytogenetic analysis. In addition, the FISH method made possible the detection and determination of i(12p) and the chromosome 12 copy number in cases in which conventional cytogenetic analysis was unsuccessful. We found the incidence of i(12p) in seminomas to be low (7%) compared to that in nonseminomas (75%) when tumor biopsy specimens were studied by FISH. Our results show that the FISH technique can be used reliably for detection of the diagnostically and prognostically useful i(12p) marker in GCT tumor biopsy specimens.