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The survival benefit of neoadjuvant chemotherapy (NAC) was assessed in patients with FIGO stage III or IV ovarian cancer, and the prognostic value of various therapeutic factors was determined. In patients treated for stage III or IV ovarian malignancies at the Department of Obstetrics and Gynecology of Nagoya University or related institutions between 1987 and 1996, 119 had a histologic diagnosis of serous cystadenocarcinoma. For this group, the long-term outcome was compared between 96 patients receiving conventional adjuvant chemotherapy following standard surgery and 23 patients treated with NAC, both followed by a second cytoreductive surgery. In a total of 29 patients with all histologic types of malignancy, the tumor response to NAC and survival were analyzed on the basis of histology, chemotherapy regimen, residual tumor size after the second cytoreductive operation, and the dose intensity of cisplatin. The long-term outcome (5-year survival rate) was better in patients treated with conventional adjuvant chemotherapy than in patients receiving NAC, although the difference was not significant. Overall survival did not differ significantly in relation to tumor histology or chemotherapy regimen. With respect to residual tumor size after the second surgery, patients with a residual tumor < or = 2 cm in diameter had a significantly better prognosis than those with a residual tumor >2 cm. A better prognosis was also associated with a higher dose intensity of cisplatin, and patients treated at >or = 18 mg/m(2)/week survived significantly longer than those receiving <18 mg/m(2)/week.

Background: Although Cushing's syndrome arises from various neuroendocrine tumors secreting adrenocorticotropin (ACTH) ectopically, ovarian carcinoma rarely causes this syndrome., Case: A 66-year-old woman presented with facial swelling and skin pigmentation. She manifested hypercortisolemia, high plasma ACTH, and lack of dexamethasone suppression. MRI showed a solid ovarian tumor and resection of the tumor led to normalization of ACTH and cortisol levels. In addition, elevated serum vasopressin (ADH) and alpha-fetoprotein (AFP) were found, which were also normalized after removal of tumors. Pathological diagnosis was serous adenocarcinoma with neuroendocrine and hepatoid features. Immunohistochemistry detected immunoreactivity of chromogranin A, ACTH, ADH, and AFP in tumor cells., Conclusion: This is a very rare case of successful treatment of Cushing's syndrome arising from an ovarian adenocarcinoma secreting multiple endocrine substances.

Angiotensin II (Ang II) is a bioactive peptide of the renin-angiotensin system, exerting its actions not only as a vasoconstrictor, but also as a growth promoter. In human placenta, type 1 Ang II receptors (AT(1)R) are predominantly expressed in trophoblasts, and we previously reported that aminopeptidase A (APA), a cell surface peptidase that converts Ang II to Ang III, is also expressed in both normal and neoplastic trophoblasts. However, the roles of Ang II and APA in trophoblast function remain to be clarified. In the present study we examined the effects of Ang II on proliferation and APA expression in trophoblast-like BeWo choriocarcinoma cells. Treatment of BeWo cells with Ang II significantly increased DNA synthesis in a dose-dependent manner. Ang II also enhanced APA mRNA and cell surface expression in BeWo cells analyzed by Northern blotting, flow cytometry, and enzyme activity assay. The Ang II-induced proliferation and APA up-regulation were blocked by the AT(1)R antagonist candesartan, but not by the AT(2)R antagonist PD123319. Furthermore, these Ang II effects were abolished by the protein kinase C inhibitor bisindolylmaleimide I and the MAPK inhibitor PD98059. Immunohistochemistry using choriocarcinoma tissues demonstrated that APA was expressed on the cell surface of AT(1)R-positive cytotrophoblastic cells in vivo. With these findings we demonstrate that Ang II stimulates the proliferation of trophoblastic cells via AT(1)R that are linked to protein kinase C /MAPK-dependent signaling pathways, and that the Ang II-degrading enzyme APA is up-regulated during Ang II-induced cell proliferation. These observations suggest the possible regulatory mechanism by the local renin-angiotensin system, especially the Ang II-AT(1)R-APA system, for the growth of human choriocarcinoma cells.

Adipocyte-derived leucine aminopeptidase (A-LAP) is a novel zinc-metallopeptidase involved in angiotensin II (AngII) metabolism, cell migration and antigen presentation. These functions are implicated in the progression of cancer, whereas A-LAP expression and involvement have not been studied in any type of cancer. We investigated the expression of A-LAP in endometrial cancer as well as its association with angiogenesis and clinicopathological features. Immunohistochemical staining of 58 endometrial endometrioid adenocarcinoma specimens revealed that 37 were A-LAP immunoreactive. We also found that A-LAP staining correlated with histological tumor grade in a significant and reverse manner. In addition, serum CA-125 levels in patients with A-LAP positive cancers were significantly higher. However, contrary to our hypothesis that A-LAP suppresses angiogenic activity via AngII metabolism, A-LAP expression was not associated with the microvessel count determined by CD34 immunostaining. Our results suggest that A-LAP is involved in endometrial cancer cell growth and differentiation. However, further studies, especially of the biological roles of A-LAP, are required to confirm this notion., (Copyright 2003 S. Karger AG, Basel)

Dipeptidyl peptidase IV (DPPIV), cell surface peptidase, works as an adhesion molecule as well as an enzyme. To investigate the role of DPPIV as an adhesion molecule of ovarian carcinoma cells to mesothelial cells, ovarian carcinoma (SKOV3 and NOS4) cells were transfected with a sense and an antisense cDNA coding human DPPIV. The adhesion potency of these transfected cells to fibronectin- and collagen-coated plates and mesothelial cells were examined with or without fibronectin. Flow cytometry and immunohistochemistry demonstrated DPPIV in ovarian carcinoma cells. The adhesion rate of DPPIV-transfected SKOV3 (SKDPIV) cells to fibronectin-coated plates was significantly higher than SKOV3 cells, while there was no difference in the adhesion rate to non-coated plates between SKDPIV and SKOV3 cells. The adhesion rates of vector-transfected SKOV3 (SKpcDNA) cells to coating and noncoating plates were similar to those of SKOV3 cells. SKDPIV cells showed twofold adhesion potency to mesothelial cells compared to SKOV3 cells. Furthermore, an addition of soluble fibronectin resulted in a dose-dependent increase in the adhesion rate of SKDPPIV cells, but not in either SKOV3 or SKpcDNA cells. Antisense-transfected NOS4 cells decreased the adhesion potency compared to NOS4 and vector-transfected NOS4 cells. In conclusion, ovarian carcinoma cells express DPPIV and adhere to human mesothelial cells in part by DPPIV. This adhesion mechanism of DPPIV is mediated by immobilized and soluble fibronectin., (Copyright 2003 Wiley-Liss, Inc.)

Dipeptidyl peptidase IV (DPPIV/CD26) is a multifunctional cell surface aminopeptidase that is widely expressed in different cell types. Our previous study demonstrated a possible link between DPPIV expression and decreased i.p. dissemination and loss of invasive potential of ovarian carcinoma. In this report, we examined the mechanisms of the anti-invasive ability of DPPIV in greater detail. Expression of E-cadherin and beta-catenin was positively correlated with DPPIV expression among five independent ovarian carcinoma cell lines. The introduction of DPPIV cDNA into an ovarian carcinoma cell line (SKOV3) with low DPPIV expression enhanced the expression of E-cadherin and beta-catenin, with a cellular morphological change from a fibroblastic and motile phenotype to an epithelial phenotype. In addition, matrix metalloproteinase 2 and membrane type 1 matrix metalloproteinase, important markers associated with invasive and metastatic potential, were remarkably reduced. In contrast, tissue inhibitors of matrix metalloproteinases were up-regulated by DPPIV transfection. Furthermore, suppression of the phosphorylation levels of mitogen-activated protein kinase isoform, extracellular signal-regulated kinase, was observed in DPPIV-overexpressing cells. To our knowledge, this is the first evidence that increasing DPPIV expression may contribute to prolonged survival by up-regulation of E-cadherin and tissue inhibitors of matrix metalloproteinases.

This study investigated the immunohistochemical expression and localization of neutral endopeptidase (NEP) (CD10), which plays a functional role by degrading bioactive peptides, in ovarian tumors. In normal ovaries and benign cystadenomas, NEP was not detected in any epithelial or stromal cells. In borderline tumors, NEP was detected in the stromal cells in 6 of 7 serous tumors, but not in those from mucinous tumors. In ovarian carcinomas, NEP in the stromal cells was observed in 13 of 20 serous, 8 of 10 endometrioid, and 7 of 10 clear-cell adenocarcinomas. NEP was weakly detected in only 1 of 9 mucinous adenocarcinomas. The staining intensity of stromal NEP was decreased in grades 2 and 3 serous carcinomas compared with that in grade 1 serous carcinomas. In conclusion, NEP was specifically expressed in the stroma of borderline and malignant ovarian tumors, but not in adenomas. Furthermore, stromal NEP was downregulated as the histological grade advanced. These results suggest that NEP may play a role in the regulation of neoplastic transformation and tumor differentiation in epithelial ovarian carcinomas.

Purpose: Oxytocin (OT) was reported to inhibit the proliferation of various neoplastic tissues and cells, however, the regulation system remains unclear. This study examined the role of OT and its regulatory ability in endometrial adenocarcinoma., Experimental Design: To investigate the possible function of placental leucine aminopeptidase (P-LAP) in endometrial adenocarcinoma, we transfected P-LAP cDNA into A-MEC cells, showing the lowest enzyme activity of P-LAP. Also we examined P-LAP protein expression in human endometrial adenocarcinoma., Results: We demonstrated the presence of P-LAP, which is identical to cystine aminopeptidase as oxytocinase, in human endometrial adenocarcinoma tissues and found that the expression of P-LAP increase with advances in the grade. Exposure of endometrial adenocarcinoma cell lines to OT caused dose- and time-dependent inhibition of growth. Treatment with 10(-7) M OT for 72 h reduced cell growth by 62, 25, and 30% in A-MEC, HEC1A, and Ishikawa cells, respectively. P-LAP-transfectant cells not only partially recovered from OT-induced growth inhibition but also showed a higher growth rate than parental cells under condition without OT. An OT receptor antagonist and a protein kinase A inhibitor blocked OT-induced growth inhibition in A-MEC and A-MEC-pc cells but not in A-MEC-LAP cells., Conclusions: These findings suggested that P-LAP might be functionally positive on carcinoma cell growth by degrading suppressive peptides such as OT.

Objectives: Dipeptidyl peptidase IV (DPPIV)/CD26 is a cell surface aminopeptidase. This study investigated the expression and localization of DPPIV in endometrial endometrioid adenocarcinomas of different grades., Study Design: Immunohistochemical analysis was performed by using DPPIV and regulated on activation, normal T-cell expressed and secreted (RANTES) specific monoclonal antibodies. Cell proliferation was evaluated by bromodeoxyuridine (BrdU) uptake assay., Results: Immunohistochemical analyses showed that DPPIV was strongly or moderately stained in glandular cells of the normal secretory phase. In endometrial adenocarcinoma, the DPPIV expression decreased with advancing grade (P <.01). Furthermore, RANTES, one of the possible DPPIV substrates, was highly expressed in all grades of endometrial adenocarcinoma cells. The addition of RANTES to endometrial adenocarcinoma cells increased proliferation in a concentration-dependent manner., Conclusion: DPPIV is expressed in normal endometrial glandular cells, but its expression in endometrial adenocarcinoma is down-regulated with increasing grade. Our data also suggest a regulatory role of this ectoenzyme in neoplastic transformation and progression of endometrial adenocarcinomas possibly by degrading certain bioactive peptides such as RANTES.

Aims: Membrane-bound aminopeptidases in human placenta are thought to be involved in maintaining homeostasis during pregnancy by metabolizing bioactive peptides such as oxytocin and angiotensin at the interface between the fetus and mother. Because determining the precise localization of these enzymes is required to support this notion, we investigated the ultrastructural localization of two principal enzymes, aminopeptidase A (APA; EC 3.4.11.7)/angiotensinase and placental leucine aminopeptidase (P-LAP; EC 3.4.11.3)/oxytocinase in human first trimester and full-term placenta., Methods: Immunohistochemical analysis using anti-P-LAP and anti-APA antibodies was performed on ultrathin frozen sections of fixed human placental villi., Results: Transmission immunoelectron microscopy revealed that both enzymes were expressed on the surface of apical microvilli of syncytiotrophoblast cells and, to a lesser extent, on the basal infoldings. The location of the two enzymes did not vary between the first trimester and full-term placenta sections, while the staining intensities were slightly enhanced in full-term villi., Conclusions: Our observation that P-LAP and APA are present on the microvilli, which is a site of interaction between the mother and fetus, suggests possible involvement of these enzymes in cleaving peptide hormones from the fetus and mother in order to regulate bioactivity.

Objective: There are a few long-term follow-up studies of patients with stage I ovarian carcinoma. The objective of this study was to evaluate prognostic factors for long-term survival., Methods: Two hundred fourteen stage I ovarian carcinomas (registered by the Tokai Ovarian Tumor Study Group from 1988 to 1999) were analyzed. Staging according to the International Federation of Gynecology and Obstetrics was set without considering the pathologic findings of the lymph nodes. Stage Ic was divided into Ic(b) and other Ic., Results: Seventy-four patients were stage Ia, 2 were stage Ib, and 138 were stage Ic, consisting of 75 Ic(b) and 63 other Ic patients. The ratios of stage Ic to Ia and stage Ic(b) to Ic were highest in clear-cell adenocarcinoma among histologies analyzed. There were significant differences in disease-free survival curves between each group. In clear cell adenocarcinoma, there was a significant difference in disease- free survival curves between stage Ic(b) and other Ic (p = 0.0373), while this tendency was also observed in other histologies except for mucinous carcinoma. Multivariate analysis demonstrated that substage was a significant prognostic factor of both disease-free and overall survival., Conclusions: Stage Ic(b) patients showed poorer survival than stage Ia patients but better than survival in other Ic. Thus, cautious treatment of ovarian tumor during surgery is quite important to avoid rupture., (Copyright 2003 S. Karger AG, Basel)

Background and Aim: Preoperative hepatic function was evaluated using technetium-99 m-diethylenetriaminepenta-acetic acid-galactosyl-human serum albumin (Tc-GSA) and a scintillation camera to detect hepatic Tc-GSA uptake by the asialoglycoprotein receptor., Methods: Sixty-two preoperative patients with liver cancer, including hepatocellular and cholangiocellular carcinomas, were studied, using two-parameter two-compartment model analysis. This model is simpler than either the three- or five-compartment model, both of which are accurate but which require complicated analysis and enormous calculation. The parameters k1 and k2 represented the transfer rate constant from the blood to the liver and from the liver to the blood, respectively. We calculated k1, k2, and k1/k2 from time-radioactivity curves of the heart and liver as well as VLmg, which represented the maximal amount of Tc-GSA in the liver., Results: The results were compared to those of conventional liver function analysis using Tc-GSA (the index of blood clearance (HH15) and the receptor index (LHL15)) or indocyanine green (ICGR15). Both HH15 and LHL15 were significantly correlated with k1, k1/k2, and VLmg. In addition, they closely correlated with the results of ICGR15 and some serum hepatic function tests (aspartate aminotransferase, choline esterase, albumin, platelet). The pathological grading for liver cirrhosis also correlated with k1, k1/k2, and VLmg. From a clinical point of view, VLmg had a significant correlation with the Child-Pugh score., Conclusions: These results suggest that this new compartment analysis will be useful in evaluating liver function, as it is accurate, simple and convenient., (Copyright 2003 Blackwell Publishing Asia Pty Ltd)

The presence of an extrahypothalamic gonadotropin releasing hormone (GnRH) in human placenta is well known and this decapeptide is presumed to play an important role in the regulation of the function and growth of human placenta. Immunohistochemistry showed that neutral endopeptidase 24.11 (NEP), a candidate of the responsible enzyme of GnRH degradation, is highly expressed on the cell surface of trophoblasts. Hydrolysis of GnRH by human villi was studied by measuring liberated amino acids using high performance liquid chromatography. The GnRH degrading activity was 1.53 times higher after incubation with the membrane fraction of first trimester villi than that after incubation with the membrane fraction of term villi. Phosphoramidon, a potent inhibitor of NEP, reduced the liberated amino acids to about a half, suggesting that NEP is a responsible enzyme for GnRH degradation. Ubenimex, which can inhibit several aminopeptidases, also reduced the liberated amino acids to about 50 per cent. O-phenanthroline, EDTA, and thiorphan could inhibit GnRH degradation but inhibitors of post proline endopeptidase could not. Furthermore, GnRH degrading activity of the membrane fraction was reduced remarkably after the membrane fraction was immunotitrated by anti NEP and anti placental leucine aminopeptidase (P-LAP) IgG. In conclusion, NEP and P-LAP are responsible enzymes for GnRH degradation in human villi.

Neutral endopeptidase 24.11 (NEP) is identical to CD10, which is a differentiation antigen for early B-lymphoid progenitors in the B-cell differentiation pathway. This ectoenzyme is known to have a key role in the control of growth, differentiation, and signal transduction of many cellular systems by regulating bioactive peptides and cytokines. Recently, we demonstrated that NEP/CD10 is upregulated during forskolin-induced choriocarcinoma cell differentiation, suggesting that NEP/CD10 is a trophoblast differentiation marker. The purpose of this study was to clarify the enhancement of NEP/CD10 expression and its signal transduction pathway during phorbol ester (PMA)-induced differentiation of BeWo choriocarcinoma cells. PMA-induced differentiation of BeWo cells was confirmed by morphological change and human chorionic gonadotropin (hCG) secretion, which was completely blocked by a protein kinase C (PKC) inhibitor, Bisindolylmaleimide I (Bis I). On immunoblot analysis, PMA enhanced NEP/CD10 expression in a dose- and time-dependent manner, which was completely abolished by Bis I and a mitogen-activated protein kinase (MAPK) kinase (MEK) inhibitor, PD98059. PMA also induced phosphorylation of p44/p42 extracellular signal-regulated kinases (ERK) 1 and 2. These observations indicated that activation of PKC by PMA induced differentiation of BeWo cells, and that PMA activated MAPK/ERK, which resulted in the enhancement of NEP/CD10 expression. Furthermore, immunocytochemical analysis showed that NEP/CD10 expression was detected on the membranes of PMA-treated differentiated BeWo cells. In summary, we demonstrated that NEP/CD10 was enhanced during PMA-induced differentiation of BeWo choriocarcinoma cells through a PKC-dependent MEK/ERK signalling pathway. Our findings also suggest that NEP/CD10 may play a functional role in the process of trophoblast differentiation.

Dipeptidyl peptidase IV (DPPIV) is a multifunctional cell surface aminopeptidase with ubiquitous expression. Recent studies have suggested that DPPIV plays an important role in tumor progression in several human malignancies. In the present study, we investigated the correlation between DPPIV expression and progressive potential in ovarian carcinoma. We demonstrated that ovarian carcinoma cell lines with higher DPPIV expression were less invasive. Furthermore, DPPIV overexpression in SKOV3 cells, derived from serous cystadenocarcinoma, with little DPPIV expression induced a dramatic change in cellular morphology and a significant decrease in the abilities of both migration and invasion. In addition, we have also shown that nude mice inoculated with DPPIV-transfected SKOV3 cells showed significantly less peritoneal dissemination and longer survival time than those receiving the parental or vector-only transfected cells (mean survival time, 64.9 +/- 4.7, 35.7 +/- 2.8, and 36.6 +/- 1.8 days, respectively). This evidence implies that DPPIV may functionally suppress peritoneal dissemination in ovarian carcinoma.

Objective: Although the ovaries are common sites of metastases from a variety of primary neoplasms excluding carcinomas of the genital tract, there were few reports concerning survival and prognostic factors. The objective of this study was to assess the clinical factors affecting survival., Methods: Fifty-three secondary ovarian carcinomas excluding metastases from genital tract carcinoma were registered by the Tokai Ovarian Tumor Study Group from 1989 to 1999. FIGO staging was set without considering the pathologic findings of the lymph nodes and the primary tumors., Results: Twenty-four patients were stage I, 11 were stage II, 16 were stage III, and 2 were stage IV. There were significant differences in the survival curves between the early stages (I and II) and advanced stages (III and IV). The 5-year survival rate for patients without residual tumors was 39.9%, while all patients with residual tumors after surgery died within 33 months. The most frequent primary tumor was large intestinal carcinoma, and the second was gastric carcinoma. The 5-year survival rate for patients with large intestinal primary tumors was significantly better than that with stomach tumors. Multivariate analysis demonstrated that tumor stage and primary tumor sites were significant prognostic factors., Conclusion: FIGO staging without considering lymph node involvement and the primary tumor was a significant prognostic factor, and prognostic factors for primary ovarian carcinomas can be applied to secondary ovarian carcinomas. Furthermore, the primary tumor site was also an important prognostic factor for survival., (Copyright 2002 S. Karger AG, Basel)

This study evaluated whether preoperative findings are useful prognostic indicators in patients with non-gynecologic carcinomas metastasizing to the ovaries. Fifty-three patients were analyzed. Primary sites of carcinoma were discovered after gynecologic surgery in 20 patients. In 10 patients, primary sites of carcinoma were discovered during gynecologic surgery. Thirty-one patients received surgery on the primary site before gynecologic surgery. The primary site could not be identified in 2 patients. The 5-year survival rate was 40.0% in patients, who underwent surgery on the primary site before gynecologic surgery. However, the 5-year survival rate was 12.2% in patients, who underwent surgery on the primary site during or after gynecologic surgery. The CEA level was significantly lower in the stomach primary group than in the intestine primary group. However, CA125 was the only marker significantly affecting survival. In comparison with primary ovarian carcinoma, the sensitivity of CEA was significantly higher for non-gynecologic ovarian carcinoma, while that of CA125 was significantly less sensitive for non-gynecologic ovarian carcinoma. In conclusion, the prognosis was better in patients, who had undergone resection of primary carcinoma before gynecologic surgery on non-gynecologic ovarian carcinoma. CEA is a useful marker to distinguish non-gynecologic from primary ovarian carcinoma, while CA125 is the only significant prognostic marker for non-gynecologic ovarian carcinoma., (Copyright 2002 S. Karger AG, Basel)

Cell surface aminopeptidases play an important role in biological processes through degradation of small peptides. There are many bioactive peptides in ascites and these peptides are involved in carcinoma cell dissemination and infiltration. In human mesothelial cells dipeptidyl peptidase IV (DPPIV) shows the highest expression mostly in four cell surface aminopeptidases: aminopeptidase A, neutral endopeptidase 24-11, aminopeptidase N and DPPIV. Since mesothelial cells are always in contact with ascites, we examined the influence of malignant ascites on DPPIV. DPPIV enzyme activity in mesothelial cells was enhanced by the addition of ascites obtained from ovarian carcinoma patients in a time- and concentration-dependent manner, and flow cytometry and immunocytochemistry also revealed an increased expression of DPPIV on the cell surface of mesothelial cells. The <3-kD fraction of malignant ascites increased the DPPIV enzyme activity to the same level as the total ascites. Northern hybridization demonstrated that DPPIV mRNA was increased 3-fold by the addition of the <3-kD malignant ascites. In conclusion, DPPIV is highly expressed in human mesothelial cells and was regulated by ascites., (Copyright 2002 S. Karger AG, Basel)

Background: Gestational choriocarcinoma is a malignant tumor that frequently metastasizes to the highly vascularized organs such as the lung, brain, and liver via hematogenous spread. However, this tumor rarely metastasizes to the eye and only a few cases of metastasis to the choroid have been reported., Case: A 17-year-old woman presented with visual field defects, decreased vision, and increasing pain in her left eye. She had undergone evacuation of a complete hydatidiform mole 32 months prior to the presentation. Ophthalmologic evaluation revealed a metastatic choroidal tumor, and a CT scan showed a metastatic tumor in the left lung. The serum hCG level was elevated at 7780 mIU/ml. A clinical diagnosis of metastatic gestational choriocarcinoma involving the choroid and lung was made. The patient received 13 courses of combination chemotherapy, resulting in complete remission. Radiotherapy and surgical treatment were unnecessary., Conclusion: This is a very rare case of the successful treatment of gestational choriocarcinoma metastatic to the choroid using systemic chemotherapy alone., ((c)2001 Elsevier Science.)

In human placenta aminopeptidase A (APA), a principal enzyme that converts angiotensin II to angiotensin III, seems to be involved in angiotensin II metabolism during pregnancy. In this study, we investigated the possible effects of progesterone and estrogen on APA mRNA and protein levels in choriocarcinoma cells as a model for placenta. By RNase protection assay, progesterone induced higher APA mRNA levels than estrogen at the same concentration. Progesterone exhibited dose-dependent stimulation of APA mRNA, 1.8-fold increase at 10(-6) m for 24 h treatment. Progesterone at 10(-6) m increased APA mRNA levels within 12 h and in time-dependent fashion up to 24 h. Fluorescence-activated cell sorting analysis and measurements of APA activities revealed the induction of APA protein by progesterone. Expression of progesterone receptors (PR) and glucocorticoid receptors (GR) were determined in these cells by RT-PCR, which suggested that the progesterone's actions might be displayed through PR and/or GR. These findings may serve as a useful model to study the effects of progesterone on angiotensin II metabolism in placenta, although the physiological validity of these studies remains to be clarified., (Copyright 2001 Harcourt Publishers Ltd.)

Neutral endopeptidase (NEP)/CD10, a cell-surface peptidase degrading various bioactive peptides, is mainly present in syncytiotrophoblasts in the human placenta. However, the change in NEP expression upon trophoblast differentiation remains to be clarified. In the present study, we examined the expression of NEP in the differentiating trophoblast using the BeWo choriocarcinoma cell line as a model system. Under the normal culture conditions, NEP was very weakly expressed on most proliferating cytotrophoblastic BeWo cells, while a minority of the cell population (less than 5 per cent ), consisting of giant, multinucleated cells, clearly expressed NEP at the cell membrane. Treatment of BeWo cells with forskolin (FSK) for 48-72 h resulted in an 11- to 44-fold increase in the level of hCG secretion and induced cell fusion leading to the formation of multinucleated syncytiotrophoblasts, indicating functional and morphological differentiation. Fluorescence-activated cell sorting (FACS) analysis revealed that treatment with FSK significantly increased the cell-surface protein expression of NEP on differentiating BeWo cells. Consistently, there was a significant increase in the NEP enzymatic activity after FSK treatment. The level of hCG secretion from the FSK-treated cells was further enhanced when the cells were treated in the presence of the NEP inhibitor phosphoramidon. Immunohistochemical analysis of normal chorionic villi and choriocarcinoma tissues revealed the localization of NEP in syncytiotrophoblastic cells, as opposed to weak or negative staining in cytotrophoblastic cells. These data demonstrate that induction of choriocarcinoma cell differentiation is associated with an increase of NEP/CD10 expression at the cell surface, suggesting a role of this enzyme in regulating differentiated trophoblast functions such as hCG secretion. NEP/CD10 may also be a new cellular differentiation marker of both the normal and neoplastic trophoblast., (Copyright 2001 Harcourt Publishers Ltd.)

We recently found that mobilized peripheral blood stem cell (PSC) products (from both cancer patients and normal donors) contain high levels of CD14+ monocytes, which can inhibit the proliferation of allogeneic and autologous T cells. We found in our studies that using CD14+ monocytes from mobilized PSC products (from normal and cancer patient donors), normal apheresis products or normal peripheral blood (PB) can affect lymphocyte function and apoptosis-dependent T cell activation. However, it appears that the apoptosis is dependent on the frequency of monocytes, which is increased by both mobilization and apheresis. Both phytohemagglutinin (PHA)- and interleukin (IL)-2-induced proliferation of steady-state peripheral blood mononuclear cells (PBMC) were markedly inhibited by co-culture with irradiated CD14+ monocytes, although inhibition was significantly greater with PHA than with IL-2 stimulation. IL-2 (predominately CD56+ NK cells) or anti-CD3 monoclonal antibody (mAb) and IL-2-expanded lymphocytes (activated T cells) were inhibited by PSC monocytes to a significantly greater level as compared to steady-state lymphocytes. Indeed, no inhibition of T cell proliferation was observed when lymphocytes were co-cultured in the absence of mitogenic or IL-2 stimulation. In contrast, an increased proliferation was observed in co-cultures of CD14+ monocytes and steady-state or activated lymphocytes without mitogenic stimulation. Cell cycle analysis by flow cytometry revealed a significant increase in hypodiploid DNA, in a time-dependent manner, following co-culture of monocytes and PBMC in PHA, suggesting that T cell apoptosis occurred during PHA-induced activation. These results demonstrate that PSC-derived monocytes inhibit T cell proliferation by inducing the apoptosis of activated T cells and NK cells, but not steady-state cells. This suggests a potential role for monocytes in the induction of peripheral tolerance following stem cell transplantation.

We formulate a field theory for the Haldane-Rezayi quantum Hall state and its variants. A new essential ingredient is a class of super Chern-Simons field.

Objectives: Neutral endopeptidase 24.11 (NEP)/CD10 is a cell-surface peptidase that degrades various bioactive peptides including endothelin-1 (ET-1). This enzyme is known to play a role in maintaining ET-1-regulated vascular homeostasis in the normal human endometrium. The purpose of the present study was to investigate the expression and localization of NEP and ET-1 in neoplastic endometria, and also to clarify the correlation of their expression with the tumor grade of endometrial carcinoma., Methods: Immunohistochemical analysis for NEP and ET-1 expression was performed on paraffin-embedded tissue sections of 7 normal endometria (after menopause), 5 atypical endometrial hyperplasias (AEH), and 32 endometrial endometrioid adenocarcinomas., Results: In normal endometrium and AEH, NEP immunoreactivity was detected in stromal cells, but not in glandular cells. In contrast, ET-1 immunoreactivity was detected in both glandular and stromal cells. In the stromal cells of grade 1 endometrial adenocarcinoma, NEP was detected at high or moderate quantities. However, significantly decreased NEP immunoreactivity was observed in the stromal cells of grade 2 and 3 adenocarcinomas. However, NEP was not immunostained in adenocarcinoma cells except for the lesions of squamous differentiation. ET-1 immunoreactivity was weakly detected in the stromal cells of grade 1 adenocarcinoma, but the intensity of ET-1 staining increased with advancing tumor grade. The ratio of the staining scores of stromal ET-1 to stromal NEP was positively correlated with the tumor grade., Conclusions: These findings demonstrate that NEP expression in the stromal cells of endometrial adenocarcinoma is downregulated, while stromal ET-1 is upregulated, with increasing tumor grade. The present findings also suggest that NEP may play a role in the regulation of neoplastic transformation, tumor progression, and differentiation in endometrial neoplasms, possibly by degrading certain peptide growth factors such as ET-1., (Copyright 2001 S. Karger AG, Basel)

Using sonography, we classified the adnexal masses of 292 patients into 4 patterns. Pattern A was benign cystic tumors; B was benign mixed tumors (cysts with a smooth solid component); C was malignant mixed tumors (cysts with an irregular solid component or thickened septum), and D was solid tumors. We diagnosed tumors showing patterns A and B as benign, while patterns C and D represented tumors with low malignant potential or actual malignancy. The sensitivity and specificity of sonography was 82.2 and 82.1%, respectively, and these values were superior to those for tumor markers (CA125, CA19-9, CA72-4). Both the sensitivity and specificity of intraoperative frozen sections were the highest, showing that this is the most reliable examination. However, 15 of 191 patients undergoing frozen section were upgraded by the final pathological diagnosis. If sonography is performed by an experienced gynecologic oncologist, this examination is more reliable than tumor markers. However, intraoperative frozen section should still be performed during surgery for patients with ovarian tumors., (Copyright 2001 S. Karger AG, Basel)

We report herein the case of a ruptured liver abscess that resulted in pneumoperitoneum. A patient with diabetes mellitus presented with symptoms of acute abdomen. The plain abdominal radiograph and computed tomography findings revealed abdominal free air and a gas-containing liver abscess, whereby a diagnosis of a ruptured liver abscess was made. An emergency operation was performed, and the abscess was drained followed by peritoneal lavage and the administration of appropriate antibiotics. To the best of our knowledge, very few cases of spontaneous pneumoperitoneum occurring secondary to the rupture of a gas-containing liver abscess have been encountered in Japan.

High dose therapy (HDT) and stem cell transplantation (SCT) results in alterations in the immunologic network, thymic re-education and the induction of peripheral tolerance. The changes to the immunoregulatory cascade and tolerance induction associated with autotransplants have been investigated in a series of studies focused on leukocyte reconstitution and function following HDT and autologous SCT. In these studies, we observed a significant decrease in the CD4:CD8 T cell ratio post-transplantation compared to normal peripheral blood (PB) donors due to a decrease in CD4+ cells. In addition, T cell function (phytohemagglutinin (PHA) mitogenesis) was consistently depressed compared to samples obtained from normal PB donors. The loss of T cell function was associated with an increased frequency of circulating monocytes, their expression of Fas ligand (FasL) and a high frequency of apoptotic CD4+ T cells. Indeed, 28-51% of circulating CD4+ T cells were observed to be apoptotic during the first 100 days following HDT and SCT. These studies suggest that 'primed' or activated Fas+ CD4+ lymphocytes interact with FasL+ monocytes, resulting in apoptosis, leading to the preferential deletion of CD4+ T cells, a decrease in the CD4:CD8 T cell ratio and depressed T cell function. Further, as discussed herein, the T cells are activated with a predominantly type 2 phenotype, which may also contribute to the maintenance of the immunosuppressive condition. Therefore, there is the potential to regulate immune recovery by stem cell product manipulation or post-transplantation cytokine administration.

Neutral endopeptidase 24.11 (NEP)/CD10 is a cell-surface peptidase that hydrolyzes various bioactive peptides. NEP is distributed in both normal and neoplastic cells and plays a functional role by modulating cellular responses to peptide substrates. Recently, NEP has been shown to be expressed in normal placental trophoblasts, suggesting its physiological role during pregnancy. In the present study, we investigated the expression of NEP in hyperplastic and anaplastic trophoblasts in gestational trophoblastic diseases (GTDs). Flow cytometric analysis demonstrated that NEP was expressed in all choriocarcinoma cell lines examined. The NEP enzyme activity in these cell lines correlated with cell-surface protein levels and was abolished by the NEP inhibitor phosphoramidon. On immunoblot analysis, NEP protein was detected in both hydatidiform mole and choriocarcinoma tissues as a double band of 95 and 100 kDa similar to that of the normal placental tissues. Immunohistochemical analysis revealed that NEP was present on syncytiotrophoblasts, while no or very faint NEP immunoreactivity was observed on cytotrophoblasts in the normal placenta. Similarly, NEP in hydatidiform mole and invasive mole was localized on the membrane of syncytiotrophoblasts, but not on hyperplastic cytotrophoblasts. In contrast, in choriocarcinoma, NEP was highly expressed not only on syncytiotrophoblastic cells but also on invading anaplastic cytotrophoblasts. In addition, NEP was also expressed on intermediate trophoblasts in placental site trophoblastic tumors. In summary, this is the first study demonstrating the expression of NEP/CD10 in GTDs. The differential localization of NEP among various trophoblastic tumors suggests that NEP may play a functional role in the regulation of trophoblast transformation and human chorionic gonadotropin secretion.

Objective: In the present studies, we examined the role of immunoregulatory cytokine gene expression in immune reconstitution following high-dose chemotherapy and peripheral stem cell transplantation., Materials and Methods: We analyzed the steady-state mRNA cytokine levels and the immune phenotype and function in the peripheral blood mononuclear cells from intermediate-grade non-Hodgkin's lymphoma patients prior to and following high-dose chemotherapy and peripheral stem cell transplantation., Results: Significantly higher mRNA levels of both type 1 and type 2 cytokines and monokines were observed in patients undergoing high-dose chemotherapy and peripheral stem cell transplantation as compared with normal healthy individuals. Pretransplant mRNA levels of interleukin-2, -4, -8, -10, interferon-gamma and tumor necrosis factor-alpha were significantly higher than in normal individuals. In addition, on days 30 and 100 following transplantation interleukin-10 levels were significantly increased compared with pretreatment levels. In contrast, the levels of interleukin-2 mRNA and interferon-gamma were decreased significantly on day 365 compared with pretransplant levels., Conclusions: The high levels of cytokine mRNA transcripts, both prior to and following peripheral stem cell transplantation, were not due to an increased cellular frequency; rather, they appear to be due to abnormal cellular activation. However, T-cell function is significantly depressed compared with normal donors, which is associated with significantly higher levels of cellular-dependent T cell inhibitory activity and, we suggest herein, high levels of interleukin-10, a type 2 cytokine.

Aminopeptidase A (AP-A) is a cell surface metallopeptidase which specifically cleaves the amino-terminal acidic residue from peptide substrates such as angiotensin II. AP-A is identical to the differentiation-related antigen, murine BP-1 or human kidney gp160, and is involved in regulating cell differentiation and/or neoplastic transformation of certain normal and transformed cells. We examined expression of AP-A in premalignant and malignant lesions of the uterine cervix, and investigated whether its expression was related to disease progression and neoplastic transformation. Formalin-fixed, paraffin-embedded tissue sections including 14 cervical intraepithelial neoplasms (CIN) and 23 invasive squamous cell carcinomas (SCC) were immunohistochemically evaluated. AP-A was localized in the basal cell layer in normal squamous epithelium. In CIN, AP-A expression was found on dysplastic cells, and increased with the severity of the precancerous lesions. In invasive cancer, 18 of 19 non-keratinizing-type SCCs and none of 4 keratinizing-type SCCs expressed AP-A. In addition, AP-A immunoreactivity was significantly correlated with proliferating cell nuclear antigen expression in both CIN and SCC cases. Furthermore, angiotensin II type 1 receptor was present in all AP-A-positive SCCs. These results indicate that AP-A is upregulated as the lesion progresses toward carcinoma in the cervical epithelium, and suggest that AP-A may play a regulatory role in neoplastic transformation and disease progression in cervical neoplasms and may serve as a potential tumor marker during cervical neoplasia development., (Copyright 2000 S. Karger AG, Basel)

Aminopeptidase A (AP-A), a cell-surface metallopeptidase hydrolyzing peptide with N-terminal acidic residues, has been proved to be identical to the B cell differentiation antigen BP-1 and to the kidney differentiation antigen gp160, suggesting recognition of AP-A as a differentiation-related marker on certain normal and transformed cells. AP-A has also been purified from human placenta and been shown to be localized in the trophoblasts. In the present study, we examined the expression and enzymatic activity of AP-A in human gestational choriocarcinoma, a neoplastic transformant from trophoblasts which comprises a heterogenous population of trophoblastic cells in different stages of differentiation. Flow cytometry and immunoblot analysis demonstrated that AP-A was expressed in five choriocarcinoma cell lines which were secreting low or moderate levels of human chorionic gonadotropin (hCG), while two high hCG-secreting cell lines lacked AP-A expression. The AP-A enzymatic activity correlated with cell-surface levels of AP-A and was abrogated by amastatin, an inhibitor of AP-A. Immunohistochemical analysis revealed that AP-A was present in seven of eight choriocarcinoma tissues and was localized on the cell membrane of cytotrophoblastic choriocarcinoma cells, but not on cells with syncytiotrophoblast-like features. These results demonstrate that AP-A is expressed on most choriocarcinomas and its expression is restricted to low hCG-secreting, cytotrophoblastic cells and down-regulated as a function of cell differentiation, suggesting an involvement of AP-A in the differentiation/maturation process of neoplastic trophoblasts., (Copyright 2000 Harcourt Publishers Ltd.)

In two separate lymphoma populations, we examined immune reconstitution following high dose chemotherapy (HDT) and bone marrow transplantation (BMT). In the first study we followed immune reconstitution for one year after HDT and BMT. In the second study we examined the ability of the orally active immunomodulator, Bestatin to augment immune reconstitution following HDT and BMT. The studies on immune reconstitution following HDT and BMT were undertaken in a cohort of non-Hodgkin's lymphoma (NHL) patients (n = 35) and examined the peripheral blood (PB) leukocyte subsets and their in vitro functions. Our results demonstrate that monocyte and natural killer (NK) cell engraftment occurred more rapidly then did T cell reconstitution. We also observed a significant decrease in the CD4:CD8 ratio post-transplantation as compared to normal PB donors due to a decrease in CD4+ cells. In addition, following HDT and BMT, measures of T cell function (phytohemagglutinin [PHA] mitogenesis) and T helper cell activity (pokeweed mitogen [PWM] mitogenesis) were consistently depressed as compared to cells from normal PB. Further, we demonstrate a correlation between the loss of T cell function and the frequency of circulating monocytes, suggesting a cause-effect relationship. Despite the dysfunction in T cells following HDT and BMT, immune-modulating agents can still augment the immune function. One such drug is Bestatin (ubenimex), an inhibitor of aminopeptidase (AP) that binds to CD13 on macrophage/monocytes. To examine its immune modulatory activity after HDT and BMT, a dose finding (10, 30, 90 and 180 mg/day) phase Ib trial was conducted with 30 Hodgkin's disease (HD) and NHL patients who received no drug (control), or Bestatin daily for 60 days following BMT. In these studies, Bestatin administration was initiated when the absolute neutrophil count was greater than 250/mm3 on two consecutive days. These studies revealed that Bestatin significantly increased the PHA and PWM responses in a dose-dependent manner. Flow cytometric analysis revealed a significant increase in NK cells (CD56+), B cells (CD19+), as well as the CD4:CD8 cell ratio. The latter observation was associated largely with a depression in the percent of CD8+ T cells as opposed to an increase in CD4+ T cells. We conclude that despite the peripheral tolerance observed following HDT and BMT, Bestatin could significantly increase some, but not all, immune surrogates.

We report the preferential expression of Fas on CD4+ T cells and Fas ligand (FasL) on monocytes and their potential role in the selective loss of CD4+ T cells in breast cancer patients undergoing high-dose chemotherapy and peripheral blood stem cell transplantation (PSCT). A high frequency of apoptotic CD4+ T cells (28-51%) is observed during the first 100 days after PSCT concomitant with a significant increase in monocyte frequency and FasL expression (11.6-23%) on monocytes. The preferential expression of Fas on CD4+ T cells (73-92%) in the peripheral blood (PB) of these patients is associated with a significantly higher frequency of CD4+ T-cell apoptosis compared with CD8+ T cells (28-47%) and CD4+ T cells (46 +/- 5.7%) in normal PB. These data suggest that "primed" Fas+ CD4+ lymphocytes interact with activated monocytes that express FasL, resulting in apoptosis, leading to deletion of CD4+ T cells, an inversion in the CD4:CD8 T-cell ratio, and immune dysfunction. The prevention of CD4+ T-cell apoptosis and improved immune reconstitution by the manipulation of PB stem cell products, blockade of Fas-FasL interactions, or cytokine support after transplantation may be important adjuvant immunotherapeutic strategies in patients undergoing high-dose chemotherapy and PSCT.

Background: A new endoscopic classification of gastroesophageal reflux disease (GERD) has been proposed, and the term mucosal break has been introduced to describe mucosal damage. This new classification was evaluated by endoscopists with different levels of experience., Methods: Fifty endoscopic photographs for each of 20 randomly selected patients with GERD were assessed retrospectively by three groups of seven endoscopists classified by experience: group 1 (100 to 500 procedures), group 2 (500 to 3000), and group 3 (more than 3000). The new classification was modified by adding grade 0 to describe healed mucosal breaks, so that there were five grades. All photographs were assessed twice at an interval of more than 1 week, and kappa statistics were used to determine observer variation., Results: Interobserver variation within group 3 (kappa = 0.39, n = 21) and between groups 3 and 2 (kappa = 0.36, n = 49) was significantly different (p < 0.01) from that between groups 3 and 1 (kappa = 0.26, n = 49). Intraobserver variation in group 1 (kappa = 0.39, n = 7) was significantly different (p < 0.01) from that in group 2 (kappa = 0.51, n = 7) and group 3 (kappa = 0.54, n = 7)., Conclusions: Observer variation depends on level of endoscopic experience. Only experienced endoscopists should use the new classification for grading of GERD.

Previous reports showed the abnormal activation of immune cells in growth factor-mobilized peripheral blood stem cell (PBSC) products, which might be responsible for depressed T cell responsiveness to mitogens compared with normal peripheral blood mononuclear cells (PBMC). In the present study, the mRNA expression levels of interleukin (IL)-2, IL-4, IL-10, and interferon-gamma (IFN-gamma) were significantly higher in CD4+ and CD8+ T cells from mobilized PBSC products compared with CD4+ and CD8+ cells from normal peripheral blood (PB). The mRNA expression levels of IL-4 and IL-10 were significantly higher in CD8+ compared with CD4+ cells from PBSC products. However, the expression of IL-2 and IFN-gamma mRNA transcripts was similar in the CD4+ and CD8+ T cells from PBSC products. The levels of IL-10, IL-8, and tumor necrosis factor (TNF)-alpha mRNA were also significantly higher in monocytes isolated from PBSC products compared with monocytes isolated from normal PB. Expression of IL-10-specific mRNA in monocytes also was significantly higher than the levels observed in CD8+ cells from PBSC products. We suggest that both CD4+ and CD8+ cells in the PBSC products are highly activated. However, their response to phytohemagglutinin (PHA) mitogenesis is depressed in part because of IL-10 expression by CD8+ cells and monocytes in addition to the higher levels of monocyte-dependent T cell inhibitory activity. These data demonstrate that aberrant IL-10 expression in the CD8+ T cells and monocytes present in PBSC products may represent a possible mechanism of immune dysfunction in patients after high-dose chemotherapy (HDT) and peripheral blood stem cell transplantation (PBSCT).

In these studies, we compared the phenotype, function, and expression of type 1, type 2, and monocyte-associated cytokine mRNA transcripts in autologous bone marrow (BM) and growth factor-mobilized peripheral blood stem cell (PSC) products. These studies demonstrate that lymphocytes and monocytes in stem cell products are abnormally activated, expressing significantly higher levels of interleukin (IL)-2, 4 and 10, interferon gamma (IFN-gamma), and tumor necrosis factor alpha (TNF-alpha), but not IL-8, as compared to normal peripheral blood mononuclear cells (PBMC). In addition, the levels of IL-2, IL-10 and TNF-alpha are significantly higher in mobilized PSC as compared to BM products. The high cytokine levels are unexpected as T cell function in stem cell products is depressed. PSC products have high levels of T cell inhibitory activity, which directly correlates with IL-10 expression, both of which are mechanisms that might be involved in the immune dysfunction within stem cell products used for autologous stem cell transplantation. These data demonstrate that: (1) immune cells in autologous BM and PSC products are activated with the expression of high levels of type 1 and type 2 cytokines as well as monokines; (2) PSC products contain a high frequency of monocytes which mediate T cell inhibitory activity; and (3) despite the high levels of cytokine expression, T cell function in stem cell products is depressed. The significance of these immune abnormalities within stem cell products for myeloid and lymphoid recovery following autologous stem cell transplantation remains to be determined.

The expansion of T and natural killer (NK) cells in growth factor-mobilized peripheral blood stem cell (PSC) products with interleukin-2 (IL-2) requires a reduction in monocyte frequency. Monocytes are enriched with stem cells during apheresis and, in this series of growth factor-mobilized PSC products from breast cancer patients, represented 36 +/- 6% of the cells in the product. Immunophenotyping studies revealed that monocytes inhibited the proliferation of NK cells (CD56+ and CD3- CD8+ CD56+ cells) and T cells (CD3+, CD4+, and CD8+ cells) during IL-2 co-culture for 7, 14, or 21 days. A reduction in monocytes resulted in 61-fold expansion of CD3- CD8+ CD56+ cells compared with a 3.7-fold increase of CD3+ cells by day 21. In addition, following IL-2 co-culture, cells from PSC products with a reduced frequency of monocytes had a significantly increased T cell mitogenic response and NK cell activity in PSC products compared with intact products. We suggest that monocytes inhibit the IL-2-dependent proliferation and augmented function of NK and T cells from growth factor-mobilized PSC products.

Adenovirus infection of CD34+ hematopoietic stem/progenitor cells is dependent on the multiplicity of infection (MOI), time of incubation, the volume in which the co-incubation occurs and the presence or absence of growth factors. Studies revealed that a brief co-incubation (1-8 hours), resulted in low levels of transgene expression, suggesting that adenovirus infection of CD34+ cells occurs slowly, and optimal transduction requires a 24 hour exposure to adenovirus. Infection by Ad/beta-gal or Ad/p53 at a MOI of 500:1 provided a high transduction efficiency but inhibited hematopoietic function. However, treatment at a MOI of 50-100 resulted in efficient transduction (10.7-15.7% positive) without detectable toxicity. Secondary proof of adenovirus transgene expression was demonstrated by detection of mRNA for p53 in Ad/p53 infected stem cells. We conclude that a 24 hour exposure to recombinant adenovirus encoding p53 or beta-gal, at a MOI of 50-100 is optimal for in vitro gene transfer to BM cells and has no significant effect on hematopoietic function. Adenovirus-mediated transduction of BM cells can also be modulated by growth factors (IL-3, GM-CSF and G-CSF) with improved gene delivery and maintenance of hematopoietic function. In summary, adenovirus vectors can be used to transiently transduce stem cells, and conditions have been defined to maximize expression and limit inhibitory effects on CD34+ cells. These data support continued investigation of this vector for local cytokine delivery and purging of stem cell products.

We report a case of a giant malignant phyllodes trmor examinde by &sup99m; Tc-sestamibi (MIBI) mammoscintigraphy. The patient was a 51 yera-old woman who complainedof bleeding from a large mass in her right breast. The tumor was well circumscribed, with an ulcerized surface. The accumulation of &sup99m; Tc-MIBI in the tumor was recognized on &sup99m; Tc-MIBI scintigraphy. A standard radical mastectomy was performed with a wide margin of skin. The resected specimen measured 20x17x13cm, weighed 2100g and was histologically diagnosed as a malignant phyllodes tumor. The skin defect was reconstructed by a rectus abdominis musculocutaneous flap, with good cosmetic results. &sup99m; Tc-MIBI scintigraphy may have the potential to distinguish a malignant from benign phyllodes tumors.

Rapid immune reconstitution is observed following autologous peripheral blood stem cell transplantation (PSCT) as compared to autologous bone marrow transplantation (ABMT), although it is depressed compared to that observed in normal individuals. The immune dysfunction occurs despite the restoration of normal lymphoid cell numbers and may be associated with the immunologic characteristics of the infused peripheral blood stem cell (PSC) product. We report herein that the in vitro T cell proliferation and NK activity in PSC products of breast cancer patients are significantly increased following the removal of CD14+ monocytes (33 +/- 2% of the PSC product) by carbonyl iron magnetic cell isolation (CI). In vitro expansion of PSC cells cultured for 7-21 days in the presence of interleukin-2 (IL-2) is also significantly increased by depletion of the phagocytic cells. The PHA and IL-2 mitogenic responses, as well as NK activity of the expanded cells, was also significantly increased by the depletion of the phagocytes. In summary, the depletion of phagocytic monocytes from PSC products restores the proliferative and functional properties of T and NK lymphocytes and may facilitate adoptive cellular therapy, as well as rapid immunologic reconstitution post-PSCT.

Granulocyte-macrophage colony-stimulating factor, mobilized peripheral blood stem cell (PSC) products, and peripheral blood leukocytes posttransplantation contain cells that cause allogeneic and autologous T cell apoptosis. Isolation and characterization of these cells demonstrated that they were low-density (Percoll fractionation) CD14+ monocytes. T cells in PSC products have a depressed phytohemagglutinin (PHA) mitogenic response; however, purified CD4+ or CD8+ T cells exhibit a statistically normal mitogenic function. Furthermore, no T cell inhibitory activity was observed in CD14+, CD4+, and CD8+ cell-depleted fractions enriched in CD4- CD8- TCR alpha/beta+ T cells. Inhibition of T cell function by CD14+ monocytes required cell-cell contact, and the analyses of DNA fragmentation by Southern and TUNEL analysis demonstrates an activation-induced T cell apoptosis in the presence of CD14+ monocytes. Reverse-transcriptase polymerase chain reaction studies suggested that high levels of interleukin-10 or tumor necrosis factor gene transcripts in the PSC products may contribute to the inhibition of T cell function.

We report a case of a giant malignant phyllodes tumor examined by(99m)Tc-ses-tamibi (MIBI) mammoscintigraphy. The patient was a 51 year-old woman who complained of bleeding from a large mass in her right breast. The tumor was well circumscribed, with an ulcerized surface. The accumulation of(99m)Tc-MIBI in the tumor was recognized on(99m)Tc-MIBI scintigraphy. A standard radical mastectomy was performed with a wide margin of skin. The resected specimen measured 20 X 17 X 13 cm, weighed 2100 g and was histologically diagnosed as a malignant phyllodes tumor. The skin defect was reconstructed by a rectus abdominis musculocutaneous flap, with good cosmetic results.(99m)Tc-MIBI scintigraphy may have the potential to distinguish a malignant from benign phyllodes tumors.

A majority of patients with intermediate or high-grade non-Hodgkin's lymphoma (NHL) who are treated with high-dose chemotherapy (HDT) and hematopoietic stem cell transplantation subsequently relapse. Until recently, transplantation was associated with high morbidity and mortality and the focus was on improving the safety of this procedure. However, the use of growth factors and other supportive measures has successfully reduced treatment mortality to less than 5%. Therefore, new strategies need to be developed to eliminate the growth of any occult tumor cells reinfused with the stem cell products and the tumor cells remaining in the patient. One approach is to improve the immune function of the patients by a more rapid immune reconstitution and augmentation of effector cell function. We report studies comparing immune recovery following HDT and autologous peripheral stem cell transplantation (PSCT) as compared to autologous bone marrow transplantation (ABMT). These studies examined patients with intermediate and high-grade non-Hodgkin's lymphoma (NHL) who were treated with HDT and PSCT (n = 56) or ABMT (n = 60). The PSCT patients had a significantly faster recovery of circulating monocytes (CD14+ cells), natural killer ((NK) CD56+) cells, T helper (CD4+) cells, TCR gamma/delta cells, and naive T lymphocytes (CD45RA+). Following ABMT there was a significantly more rapid increase in the frequency of T suppressor/effector (CD8+) cells, B (CD19+) cells, CD34+ cells, polymorphonuclear leukocytes (PMN) and memory T lymphocytes (CD45RO+). The CD4:CD8 and CD45RA:CD45RO ratios were consistently higher in the PSCT group as compared to ABMT suggesting an improved ratio of T helper to T effector/suppressor cells and naive T cells. The differences in cellular phenotype translated into improved T cell function (PHA mitogenesis) and T cell help (pokeweed mitogenesis). In addition, there as an accelerated reconstitution of NK cell activity following PSCT as compared to ABMT. The more rapid reconstitution of NK and T cells in patients rescued with PSCT as compared to ABMT may contribute to an improved clinical outcome. Further, patients receiving a PSCT may be more responsive to adjuvant immunotherapy following transplantation.

Bestatin (ubenimex), an inhibitor of aminopeptidase, is an oral immunomodulator that binds to CD13 (aminopeptidase N) on macrophages/monocytes. To examine its immunomodulatory effect after high-dose therapy and autologous bone marrow transplantation (BMT), a dose-finding phase Ib trial was conducted with 30 Hodgkin's disease and non-Hodgkin's lymphoma patients who received no drug (control), 10 and 30 mg (low dose), or 90 and 180 mg (high dose) of bestatin daily for 60 days following autologous BMT. Bestatin administration was initiated when the absolute neutrophil count was greater than 250/mm3 on 2 consecutive days. The serum neopterin levels, an indicator of monocyte/macrophage activation, increased in the high-dose group compared to the control group (not significantly) and the low-dose group (significantly). Similarly, the colony-stimulating activity in the sera was significantly increased in the high-dose group compared to the control and low-dose groups. We also examined the expression of cell-surface markers on monocytes in these patients by fluorescent cytometry analysis. There was no significant difference either in the frequency or absolute number of monocytes (CD14+) among the three groups at any time. However, a significant increase in the frequency of CD16(FcgRIII)-positive monocytes (a marker of activation) was observed in the high-dose group compared to controls from day 14 to day 60 after the start of bestatin administration. Further, the frequency of HLA-DR+ monocytes (another marker of activation) was significantly increased in the high-dose group. These results indicate that bestatin at higher doses (90 and 180 mg daily), but not lower doses, activates macrophages/monocytes, as demonstrated by phenotypic marker (HLA-DR and CD16) up-regulation, and this provides augmentation of neopterin and colony-stimulating activity in the serum of patients following autologous BMT.

We examined four choriocarcinoma cell lines, NaUCC-1, NaUCC-3, NaUCC-4 and BeWo, for the presence of epidermal growth factor (EGF) by enzyme immunoassay and reverse transcription and polymerase chain reaction, and for EGF receptor (EGFR) by 125I-EGF binding assay. Specific EGF binding and EGF proteins were detected in these four choriocarcinoma cell lines. On the cell lines examined, NaUCC-4 had the greatest EGF binding capacity (18 x 10(5) sites/cell) and the highest amount of immunoreactive EGF (142 pg/ml). These results prompted us to assess the significance of EGF/EGFR autocrine mechanism in NaUCC-4 cells. Low doses of exogenous EGF stimulated 3H-thymidine incorporation, and monoclonal antibodies against EGF or EGFR dose-dependently inhibited 3H-thymidine incorporation. On the other hand, these antibodies did not significantly affect hCG production. These results suggested that EGF might function in an autocrine manner to stimulate proliferation rather than differentiation of NaUCC-4 choriocarcinoma cells.

Human bone marrow mononuclear cells (BMMNCs) and enriched CD34 positive (CD34+) cells were transduced with adenovirus vectors encoding Escherichia coli beta-galactosidase gene. Tranductions were carried out by 24-hour coincubation with adenovirus vectors at different multiplicities of infections (moi). Efficacy of gene transfer into BM cells and expression of the gene product (ie, beta-galactosidase) were studied using X-Gal histochemical staining and flow cytometric analysis. X-Gal staining demonstrated that the percentage of positive cells at mois of 5 to 500 was 3.4% to 34.5% for BMMNCs and 6.0% to 20.0% for enriched CD34+ cells. Similar results (1.5% to 35.7% for BMMNCs and 5.4% to 24.2% for enriched CD34+ cells) were obtained with flow cytometric analysis using fluorescein di-beta-D-galactopyranoside (FDG). Multicolor flow cytometry analysis, which included FDG, demonstrated that BM progenitors (CD34+ or CD34+CD38-), T cells (CD2+), B cells (CD19+), natural killer cells (CD56+), granulocytes, and monocytes all expressed the adenovirus transgene. To ascertain the effects of adenovirus vectors on normal BM progenitors, the numbers of colony forming unit-granulocyte/macrophage (CFU-GM), burst-forming unit-erythrocyte (BFU-E), and high-proliferative potential-colony-forming cells (HPP-CFC) after 24-hour coincubation with adenovirus vectors were determined. When BMMNCs or enriched CD34+ cells were incubated with adenovirus vectors at mois of 5 and 50, no significant differences in the numbers of CFU-GM, BFU-E, and HPP-CFC were observed compared with the uninfected control cells. However, the numbers of CFU-GM were significantly (P < .01) decreased when BMMNCs or enriched CD34+ cells were incubated with adenovirus vectors at a moi of 500, compared with the uninfected control cells. The adenovirus infected cells, purified by cell sorting for FDG expression, were capable of growing in culture and gave rise to various colonies (ie, CFU-GM, BFU-E, and HPP-CFC). These data indicate that recombinant adenovirus vectors can be used to transfer genes to human BM hematopoietic cells with expression of the exogenous gene at a high transduction efficiency.

The effect of human granulocyte colony stimulating factor (hG-CSF) was investigated on the proliferation of choriocarcinoma cells in vitro. No growth-stimulating effect was observed. Then, 22 patients with invasive mole and 9 patients with choriocarcinoma who received combination chemotherapy were treated with hG-CSF when white blood cell counts decreased below 2,000/mm3. The duration of neutropenia was reduced significantly by 3-4 days with the use of hG-CSF. No side effects were observed. These results indicate safety and usefulness of hG-CSF as an adjunct to chemotherapy in the treatment of gestational trophoblastic disease.