Infantile Liver Giant Cells: Immunohistological Study of Their Proliferative State and Possible Mechanisms of Formation

The mechanism of liver giant cell formation is not clarified. Some authors consider the giant cells regenerative, others, degenerative. Paraffin sections of 10 archival cases of idiopathic neonatal hepatitis (INH), 8 of extrahepatic biliary atresia (EHBA), and 5 normal liver samples were immunostained with two well-characterized cell proliferation markers: anti-PCNA monoclonal antibody (MAb) (clone PC-10) and MAb MIB-1, which detects Ki-67, a nuclear proliferation-related antigen. In addition, polyclonal antibody to carcinoembryonic antigen (CEA) was used to identify remnants of canalicular, therefore hepatocytic, membranes in giant cells. Quantitative analysis of immunostaining was done by estimating PCNA and Ki-67 indices separately in giant cells and in nongiant hepatocytes. In normal samples, mean PCNA and Ki-67 indices were 1.22% and 0.74%, respectively. In the cases of INH and EHBA, only a small minority of giant cells showed PCNA or Ki-67 staining limited to occasional peripherally located nuclei. PCNA and Ki-67 indices were significantly higher in the non–giant cell compartment. CEA staining was seen only in rare giant cells as centrally located canalicular remnants bordered by polarized nuclei, suggesting that they had been formed from rosettes through dissolution of cell membranes. Other giant cells shared CEA-labeled canalicular membranes with mononuclear hepatocytes in rosettes. These findings indicate that the giant cells in INH and EHBA are not regenerative cells, they are not formed by amitotic division of nuclei in syncytia, and that fusion of rosette-forming hepatocytes is a possible mechanism of their formation. Received May 6, 1998; accepted July 23, 1998.     

EFFECT OF P-GLYCOPROTEIN EXPRESSION ON OUTCOME IN THE EWING FAMILY OF TUMORS

This study was designed to determine the prognostic significance of multidrug resistance, mediated by P-glycoprotein (Pgp) expression, in Ewing sarcoma. The clinical and laboratory features, treatment protocol, and outcome of 75 patients with Ewing sarcoma or peripheral neuroectodermal tumor treated between 1972 and 1997 were reviewed. Pgp expression was tested with the monoclonal antibody JSB-1. Thirty-four (64%) of the 53 tissue samples from untreated patients stained positive for Pgp. Progression-free and overall survival were 44 and 59%, respectively, in patients with negative findings, and 28 and 41% in those with positive findings; neither difference was significant. Of the 12 relapsed patients, 6 (50%) expressed more Pgp after chemotherapy than at diagnosis and 4 (33%) expressed less. Within these subgroups, 5 out of 6 and 3 out of 4 died from the disease. No correlation was found between Pgp and known prognostic factors of Ewing tumors. Pgp expression is probably an intrinsic factor of Ewing tumors but has no correlation to prognosis.     

EFFECT OF P-GLYCOPROTEIN EXPRESSION ON OUTCOME IN THE EWING FAMILY OF TUMORS

This study was designed to determine the prognostic significance of multidrug resistance, mediated by P-glycoprotein (Pgp) expression, in Ewing sarcoma. The clinical and laboratory features, treatment protocol, and outcome of 75 patients with Ewing sarcoma or peripheral neuroectodermal tumor treated between 1972 and 1997 were reviewed. Pgp expression was tested with the monoclonal antibody JSB-1. Thirty-four (64%) of the 53 tissue samples from untreated patients stained positive for Pgp. Progression-free and overall survival were 44 and 59%, respectively, in patients with negative findings, and 28 and 41% in those with positive findings; neither difference was significant. Of the 12 relapsed patients, 6 (50%) expressed more Pgp after chemotherapy than at diagnosis and 4 (33%) expressed less. Within these subgroups, 5 out of 6 and 3 out of 4 died from the disease. No correlation was found between Pgp and known prognostic factors of Ewing tumors. Pgp expression is probably an intrinsic factor of Ewing tumors but has no correlation to prognosis.     

Immunocytochemical investigation of carcinoembryonic antigen expression in neuroblastoma with monoclonal and polyclonal antibodies

Immunocytochemical localization of two monoclonal anticarcinoembryonic antigen antibodies (11-285-14; 11-359-6) which recognize different CEA epitopes, and a polyclonal anti-CEA antibody (DAKO) recognizing both carcinoembryonic antigen and nonspecific cross-reacting antigen, was studied in 59 tumor tissue samples from 18 patients with neuroblastoma. No evidence of intact tumor cell staining was seen with any of the antibodies although the polyclonal antibody often stained necrotic tissue and infiltrating inflammatory cells. It is concluded that at the level of sensitivity of immunocytochemical localization, neuroblastoma cells do not express carcinoembryonic antigen.     

Monoclonal Antibody MB2: a Potential Marker for Ewing's Sarcoma and Primitive Neuroectodermal Tumor

The smalt round-cell tumors of childhood present difficulties in diagnosis when differentiation is not apparent. Immunohistochemislry is helpful; however, the only antigen consistently detected in Ewing's sarcoma is vimentin, which may also he detected in the other types of small-cell neoplasms. The monoclonal antibody (MAb) MB2 is marketed as a B-lymphocyte marker that can be used on paraffin-embedded tissue. To determine its specificity, we performed immunohistochemical staining on pediatric tumors with MB2. These included 55 cases of small round-cell tumors (lymphomas, Ewing 's sarcoma, peripheral primitive neuroectodermal tumors [PNET], neuroblastomas, rhabdomyosarcomas, and nephroblastomas). MB2 positivity was detected in all B-cell lymphomas and in seven of nine cases of Ewing's sarcoma and three of three PNET. In neuroblastomas only differentiating ganglion cells were positive. In rhabdomyosarcomas only large rhabdomyoblasls were positive. Blastema of nephroblastomas was negative. Thus, in cases of poorly differentiated small round-cell tumors, MB2 was positive in all B-cell lymphomas, most Ewing's sarcomas and all cases of PNET. Lymphomas were distinguished by staining for leukocyte-common antigen and PNET by neuron-specific enolase. Therefore, the addition of MB2 to a discrete panel of antibodies may prove useful in the diagnosis of Ewing's sarcoma and PNET.     

18F-fluorodeoxyglucose uptake of bone and soft tissue sarcomas in pediatric patients

A high (18)F-fluorodeoxyglucose (FDG) uptake by positron emission tomography/computed tomography (PET/CT) imaging in sarcomas of adults has been reported. The current study aimed at defining the degree of (18)F-FDG uptake of pediatric sarcomas. This retrospective study included 29 patients (23 males, 6 females; mean age 14 ± 5 years) with soft tissue (n = 9) or bone (n = 20) sarcomas. Twenty-two patients (76%) underwent (18)F-FDG PET/CT and 7 (24%) had dedicated (18)F-FDG PET studies. Tumor (18)F-FDG uptake was quantified by standard uptake value (SUV)(max) and tumor-to-liver ratios (SUV ratios; tumor SUV(max)/liver SUV(mean)). Tumor SUV(max) and SUV ratios were correlated with tumor Ki-67 expression. SUV(max) ranged from 1.4 to 24 g/mL (median 2.5 g/mL) in soft tissue sarcomas and 1.6 to 20.4 g/mL (median 6.9 g/mL) in bone sarcomas (P = .03), and from 1.6 to 9.2 g/mL (median 3.9 g/mL) and 3.5 to 20.4 g/mL (median 12 g/mL) in Ewing sarcoma and osteosarcoma, respectively (P = .009). Tumor SUV ratios ranged from 0.8 to 8.7 (median 1.9) in soft tissue sarcomas and 1.4 to 8.9 (median 3.8) in bone sarcomas (P = .08). Ewing sarcoma had a significantly lower tumor SUV ratio than osteosarcoma (P = .01). Ki-67 expression correlated significantly with the (18)F-FDG uptake in bone but not in soft tissue sarcomas. All sarcomas were visualized by (18)F-FDG PET/CT imaging. A higher (18)F-FDG uptake was observed in osteosarcoma than in Ewing and soft tissue sarcomas. The results of this study suggest that the degree of tumor (18)F-FDG uptake is sufficient to allow for monitoring of therapeutic responses in pediatric sarcomas.     

Diagnosis of tuberculous meningitis by detection of antigen and antibodies in CSF and sera

OBJECTIVE: To evaluate diagnostic potential of three immunological tests, namely, detection of H37Rv antigen of M. Tuberculosis in CSF, detection of antibodies (IgG) against H37Rv in CSF and detection of antibodies (IgG) against H37Rv in serum for diagnosis of tuberculous meningitis in children. SUBJECTS: 50 children diagnosed as patients of tuberculous meningitis were included as cases and 48 children with CNS diseases of nontubercular etiology [pyogenic meningitis (n = 31), encephalitis (n = 10), seizure disorder of unknown etiology (n = 5), brain tumor (n = 2)] served as controls. METHODS: H37Rv antigen of M. tuberculosis was detected in CSF by Dot ELISA, and antibodies (IgG) against H37Rv in CSF and serum were detected by Plate ELISA. RESULTS: Detection of H37Rv antigen in CSF was the most sensitive (90%) and specific (95.83%) with positive and negative predictive values of 95.74% and 90.19%, respectively, followed by detection of antibodies in CSF (sensitivity-74%, specificity-89.58%, positive predictive value-88.10%, negative predictive value-76.78%). Detection of antibodies in serum had low sensitivity (50%), specificity (91.67%), positive predictive value (86.21%) and negative predictive value (63.76%). CONCLUSIONS: Detection of antigen in CSF is a rapid, sensitive and specific test for diagnosis of tuberculous meningitis in children. Detection of antibody in CSF may be useful in some cases but needs further evaluation. Detection of antibody in serum does not appear to be useful for diagnosis of tuberculous meningitis.     

Selection of human antitumor single-chain Fv antibodies from the B-cell repertoire of patients immunized against autologous neuroblastoma

BACKGROUND: We used phage display technology to clone human recombinant antitumor antibodies from the antibody repertoire of neuroblastoma patients immunized with cytokine-gene transduced tumor cells. PROCEDURE: Lymphocytes obtained from neuroblastoma patients either at diagnosis or after immunization with an autologous interleukin-2 gene transduced tumor vaccine were used to construct two human single-chain Fv (scFV) phage libraries. Tumor-reactive phage were characterized using ELISA, flow cytometry, and sequencing analysis. RESULTS: The initial screening after panning on neuroblastoma cells yielded a substantially higher proportion of selectivity tumor-binding phage clones derived from the immunized patients library (12.9%) than from the unvaccinated patients library (0.8%). The antibodies stained the cells from several additional pediatric malignancies, including Ewing sarcoma and rhabdomyosarcoma, in the absence of binding to any normal tissue cultures or epithelial tumor cell lines. The pattern of reactivity was different from that of antibodies recognizing other widely distributed neuroblastoma-associated antigens, suggesting recognition of a novel shared tumor antigen. CONCLUSION: The human recombinant scFV antibodies reported here appear to represent a tumor-specific B-cell response induced by autologous tumor immunization and are potentially useful targeting moieties for the treatment of selected childhood tumors.     

Cyclooxygenase-2 expression does not correlate with outcome in osteosarcoma or rhabdomyosarcoma

PURPOSE: To determine if expression of cyclooxygenase (COX)-2, an inducible enzyme with known tumor-promoting activity, correlates with outcome in pediatric sarcomas. COX-2 overexpression correlates with more aggressive disease in a variety of adult solid tumors. METHODS: Archived human osteosarcoma, rhabdomyosarcoma, and Ewing sarcoma tumors were retrospectively evaluated, blinded to outcome, for COX-2 expression by immunohistochemistry and correlated with patient characteristics and survival. RESULTS: COX-2 expression was detected in 94 of 142 (66%) tumors (osteosarcoma, 66/99 [67%]; rhabdomyosarcoma, 21/35 [60%]; Ewing sarcoma, 7/8 [88%]) and 51 of 85 (60%) diagnostic biopsies (osteosarcoma, 26/45 [58%]; rhabdomyosarcoma, 21/35 [60%]; Ewing sarcoma, 4/5 [80%]). COX-2 expression did not vary with clinicopathologic features and was not predictive of prognosis in these cases. CONCLUSIONS: This study does not support the use of COX-2 expression as an upfront prognostic variable in patients with osteosarcoma or rhabdomyosarcoma.Results from the small number of patients studied with Ewing sarcoma suggest a similar lack of predictive value for COX-2 expression. However, COX-2 inhibitors are not entirely dependent upon enzyme expression for their antitumor effects; this study does not necessarily preclude the use of COX-2 inhibitors for the treatment of pediatric sarcomas.     

Immunopathology of brain tumors

The various classifications of brain tumors are characterized by a rather disturbing diversity of tumor designations. This diversity results from the heterogeneity of human brain tumors, but also from the lack of knowledge about the histogenesis of many of these tumors. The histogenesis of some of the different types of tumor could be resolved by the application of electron microscopical studies. New aspects are gained from immunohistochemical investigations using mono- and polyclonal antibodies against intermediate filaments (GFAP, vimentin, cytokeratins), neuron-specific enolase (NSE), protein S-100, carcinoembryonic antigen (CEA), myelin-associated glycoprotein (Leu 7), synaptophysin and nuclear proliferating antigen (Ki-67). The present paper gives a survey on how immunohistochemistry can be advantageously used in the diagnosis of brain tumors.     

18F-FluoroDeoxyGlucose Uptake of Bone and Soft Tissue Sarcomas in Pediatric Patients

A high ¹⁸F-fluorodeoxyglucose (FDG) uptake by positron emission tomography/computed tomography (PET/CT) imaging in sarcomas of adults has been reported. The current study aimed at defining the degree of ¹⁸F-FDG uptake of pediatric sarcomas. This retrospective study included 29 patients (23 males, 6 females; mean age 14 ± 5 years) with soft tissue (n = 9) or bone (n = 20) sarcomas. Twenty-two patients (76%) underwent ¹⁸F-FDG PET/CT and 7 (24%) had dedicated ¹⁸F-FDG PET studies. Tumor ¹⁸F-FDG uptake was quantified by standard uptake value (SUV)_max and tumor-to-liver ratios (SUV ratios; tumor SUV_max/liver SUV_mean). Tumor SUV_max and SUV ratios were correlated with tumor Ki-67 expression. SUV_max ranged from 1.4 to 24 g/mL (median 2.5 g/mL) in soft tissue sarcomas and 1.6 to 20.4 g/mL (median 6.9 g/mL) in bone sarcomas (P = .03), and from 1.6 to 9.2 g/mL (median 3.9 g/mL) and 3.5 to 20.4 g/mL (median 12 g/mL) in Ewing sarcoma and osteosarcoma, respectively (P = .009). Tumor SUV ratios ranged from 0.8 to 8.7 (median 1.9) in soft tissue sarcomas and 1.4 to 8.9 (median 3.8) in bone sarcomas (P = .08). Ewing sarcoma had a significantly lower tumor SUV ratio than osteosarcoma (P = .01). Ki-67 expression correlated significantly with the ¹⁸F-FDG uptake in bone but not in soft tissue sarcomas. All sarcomas were visualized by ¹⁸F-FDG PET/CT imaging. A higher ¹⁸F-FDG uptake was observed in osteosarcoma than in Ewing and soft tissue sarcomas. The results of this study suggest that the degree of tumor ¹⁸F-FDG uptake is sufficient to allow for monitoring of therapeutic responses in pediatric sarcomas.     

Reactivation of TWIST1 contributes to Ewing sarcoma metastasis

1 Background

Ewing sarcoma is a cancer of bone and soft tissue. Despite aggressive treatment, survival remains poor, particularly in patients with metastatic disease. Failure to treat Ewing sarcoma is due to the lack of understanding of the molecular pathways that regulate metastasis. In addition, no molecular prognostic markers have been identified for Ewing sarcoma to risk stratify patients.

2 Procedure

Ewing sarcoma patients were divided into high or low Twist1 gene expression and survival curves were generated using the R2 microarray‐based Genomic Analysis platform ( http://r2.amc.nl ). Tumors from Ewing sarcoma patients were also evaluated for TWIST1 expression by immunohistochemistry. Ewing sarcoma xenografts were established to evaluate the role of TWIST1 in metastasis. The effects of Twist1 on migration and invasion were evaluated using migration and invasion assays in A673 and RDES cells.

3 Results

Twist1 expression was a negative prognostic marker for overall survival in a public Ewing sarcoma patient data set based on Twist1 mRNA levels and in patient tumor samples based on Twist1 immunohistochemistry. TWIST1 is detected in significantly higher percentage of patients with metastatic diseases than localized disease. Using Ewing sarcoma tumor xenografts in mice, we found that suppressing TWIST1 levels suppressed metastasis without affecting primary tumor development. Knockdown of Twist1 inhibited the migration and invasion capability, while overexpression of Twist1 promoted migration and invasion in Ewing sarcoma cells.

4 Conclusion

These results suggest that TWIST1 promotes metastasis in Ewing sarcoma and could be used as a prognostic marker for treatment stratification; however, further validation is required in a larger cohort of patients.     

Evaluation of cytarabine against Ewing sarcoma xenografts by the pediatric preclinical testing program

Treatment with the nucleoside analog cytarabine has been shown to mimic changes in gene expression associated with downregulation of the EWS-FLI1 oncogene in Ewing sarcoma cell lines, selectively inhibit their growth in vitro, and cause tumor regression in athymic nude mice. For this report cytarabine was studied in vitro against a panel of 23 pediatric cancer cell lines and in vivo against 6 Ewing sarcoma xenografts. Acute lymphoblastic leukemia cell lines were the most sensitive to cytarabine in vitro (median IC(50) 9 nM), while Ewing sarcoma cell lines showed intermediate sensitivity (median IC(50) 232 nM). Cytarabine at a dose of 150 mg/kg administered daily 5× failed to significantly inhibit growth of five xenograft models, but reduced growth rate of the A673 xenograft by 50%. Cytarabine shows no differential in vitro activity against Ewing sarcoma cell lines and is ineffective in vivo against Ewing sarcoma xenografts at the dose and schedule studied.     

Haploidentical hematopoietic cell transplantation for disseminated Ewing sarcoma

We describe the case of a 13‐year‐old girl with multifocal disseminated Ewing sarcoma family of tumor (ESFT) who received a 5/8 human leukocyte antigen‐matched haploidentical hematopoietic cell transplantation to generate a graft‐versus‐tumor effect. The patient had grade 2 acute graft‐versus‐host disease (GVHD) of the skin and chronic GVHD nausea and abdominal pain that required prednisolone for 17 months, but has been free from ESFT for 3 years 10 months after therapy. The present case suggests a beneficial effect of haploidentical hematopoietic cell transplantation in disseminated ESFT.     

Monoclonal antibody MB2: a potential marker for Ewing's sarcoma and primitive neuroectodermal tumor

The small round-cell tumors of childhood present difficulties in diagnosis when differentiation is not apparent. Immunohistochemistry is helpful; however, the only antigen consistently detected in Ewing's sarcoma is vimentin, which may also be detected in the other types of small-cell neoplasms. The monoclonal antibody (MAb) MB2 is marketed as a B-lymphocyte marker that can be used on paraffin-embedded tissue. To determine its specificity, we performed immunohistochemical staining on pediatric tumors with MB2. These included 55 cases of small round-cell tumors (lymphomas, Ewing's sarcoma, peripheral primitive neuroectodermal tumors [PNET], neuroblastomas, rhabdomyosarcomas, and nephroblastomas). MB2 positivity was detected in all B-cell lymphomas and in seven of nine cases of Ewing's sarcoma and three PNET. In neuroblastomas only differentiating ganglion cells were positive. In rhabdomyosarcomas only large rhabdomyoblasts were positive. Blastema of nephroblastomas was negative. Thus, in cases of poorly differentiated small round-cell tumors, MB2 was positive in all B-cell lymphomas, most Ewing's sarcomas and all cases of PNET. Lymphomas were distinguished by staining for leukocyte-common antigen and PNET by neuron-specific enolase. Therefore, the addition of MB2 to a discrete panel of antibodies may prove useful in the diagnosis of Ewing's sarcoma and PNET.     

Concentration of vascular endothelial growth factor (VEGF) in the serum of patients with malignant bone tumors

BACKGROUND: Vascular endothelial growth factor (VEGF) is recognized as an important stimulator of angiogenesis. Formation of new blood vessels by angiogenic factors occurs in many biological processes, both physiological and pathological, among others in growth of primary solid malignant tumors and metastasis. This implies that the inhibition of angiogenic factors like VEGF would result in a suppression of tumor growth and metastasis formation. The aim of the present study was to compare preoperative serum VEGF levels of patients having malignant bone tumors with healthy controls to identify serum VEGF levels as a tumor marker. PROCEDURE: Blood sera from patients with high-grade osteosarcoma (n = 17), chondrosarcoma (n = 4) and Ewing sarcoma (n = 6) were taken at the time of diagnosis before biopsy and compared with sera from 129 healthy persons. To measure VEGF levels in serum, a commercially available ELISA was used (Quantikine Human VEGF Immunoassay; R&D Systems). RESULTS: The observed geometric mean VEGF levels and 95% confidence intervals are 232.0 pg ml(-1) (168.9-318.5) for patients with high-grade osteosarcoma, 325.5 pg ml(-1) (169.3-625.8) for patients with chondrosarcoma, 484.3 pg ml(-1) (284.0-826.0) for patients with Ewing sarcoma, as compared to 216.2 pg ml(-1) (192.8-242.5) for healthy individuals. CONCLUSIONS: While the sample means for the three groups of sarcoma patients were higher than the respective mean for the healthy controls, only the mean for the group with Ewing sarcoma is statistically significantly higher than the mean for the healthy controls. Despite the significant difference, VEGF levels are not suitable as a marker for Ewing sarcoma.     

儿童脑胶质瘤Ki-67抗原表达及其意义

目的 研究Ki-67抗原在儿童脑胶质瘤中的表达与不同病理分类分级、肿瘤生长部位的关系.方法 对62例不同病理分类分级的儿童脑胶质瘤进行Ki-67免疫组织化学染色,观察Ki-67标记指数及染色程度.结果 胚胎性肿瘤的Ki-67标记指数(Ki-67 LI)显著高于其他腩胶质瘤(P<0.05);幕上胶质瘤的Ki-67 LI显著低于幕下胶质瘤(P<0.05);Ki-67 LI随脑胶质瘤恶性程度级别增高呈上升趋势,其中Ⅰ、Ⅱ、Iil级的Ki-67 LI显著低于Ⅳ级(P<0.05),且Ⅰ级的Ki-67 LI显著低于Ⅲ级(P<0.01);低级别胶质瘤(Ⅰ级和Ⅱ级)的Ki-67 LI显著低于高级别胶质瘤(Ⅲ级和Ⅳ级)(P<0.01);低级别胶质瘤(Ⅰ级和Ⅱ级)的Ki-67阳性细胞核染色程度明显弱于高级别胶质瘤(Ⅲ级和Ⅳ级)(P<0.01).结论 Ki-67抗原是反映儿童脑胶质瘤细胞增殖活性较可靠的细胞增殖指标,可以为小儿脑胶质瘤病理分类分级提供一种有效的辅助方法,有助于更加客观地评价息儿的预后.     

Ewing sarcoma family of tumors express adenovirus receptors and are susceptible to adenovirus-mediated oncolysis

PURPOSE: Attenuated viruses derived from adenoviruses (Ad) that kill tumor cells (oncolysis) are currently in clinical trials for selected cancers. Some cancers have proven resistant to Ad infection due to low expression of viral receptors. The authors sought to determine whether members of the Ewing sarcoma family of tumors (ESFTs) express Ad receptors and are sensitive to Ad-mediated oncolysis. METHODS: Using flow cytometry, the authors tested a panel of cell lines derived from ESFTs for expression of both the Ad receptor, coxsackie-adenovirus receptor (CAR), and the cellular mediator of Ad uptake, alpha(v)-integrins, as well as for Ad-mediated gene transduction. Cell survival assays were used to assess the sensitivity to Ad-mediated oncolysis. Immunohistochemistry was used to assess CAR expression in primary tumors. mRNA levels of CAR in cell lines and tumor samples were also queried from a cDNA expression database. RESULTS: The ESFT cell lines expressed CAR and alpha(v)-integrins, showed high levels of gene transduction, and were highly sensitive to viral oncolysis. Primary tumor samples were positive for CAR expression by immunohistochemistry. Microarray analysis confirmed CAR expression in ESFT cell lines and tumors. CONCLUSIONS: Ewing sarcoma cells express the Ad receptors and are sensitive to Ad oncolysis. Treatment of Ewing sarcoma using conditionally replicative adenoviruses should be explored.