Galactosyltransferase associated with tumor in patients with ovarian cancer: factors involved in elevation of serum galactosyltransferase

The serum level of beta1,4-galactosyltransferase (beta1,4-GalT) is increased in both malignancy and benign diseases. Galactosyltransferase associated with tumor (GAT) is one of the soluble forms of beta1,4-GalT, and is a marker of ovarian cancer with a high specificity. GAT and normal soluble beta1,4-GalT are both derived from the same membrane-bound form of the enzyme. This study investigated the mechanism of GAT elevation in patients with ovarian cancer. The serum levels of GAT and normal beta1,4-GalT were measured using specific monoclonal antibodies. In addition, nude mice bearing human ovarian cancer were used to assess the kinetics of tumor-derived enzymes. GAT and normal beta1,4-GalT were both detected in ovarian cancer patients, but only GAT reflected the tumor status. In tumor-bearing nude mice, both soluble forms of beta1,4-GalT were released from tumor cells, but the half-life of GAT was far shorter than that of normal beta1,4-GalT. Addition of serum from healthy women to colostrum (which has a high GAT content) reduced the GAT level, while adding patient serum caused a significantly smaller reduction of GAT. Addition of the serum from mouse which includes no human beta1,4-GalT to colostrum also reduced the GAT level with no significant change of total soluble beta1,4-GalT. These findings indicate that human serum contains certain factors that decrease the GAT level, but these factors are inhibited in ovarian cancer patients so that a high GAT level persists. It seems that the decrease of GAT occurs as a result of conversion into normal beta1,4-GalT.     

Expression of serine proteinase inhibitor PP5/TFPI-2/MSPI decreases the invasive potential of human choriocarcinoma cells in vitro and in vivo

OBJECTIVE: PP5/TFPI-2/MSPI is a Kunitz-type serine proteinase inhibitor with broad inhibitory spectra, abundantly produced by placenta and detected in the blood of pregnant women. Expression of PP5/TFPI-2/MSPI is exclusively detected in syncytiotrophoblasts of placenta, but is barely detectable in choriocarcinoma cells, a trophoblast-derived malignant tumor. Chromosome 7, in which the PP5/TFPI-2/MSPI gene is localized, is frequently lost in various types of tumors. We attempted to elucidate the relation between PP5/TFPI-2/MSPI expression and the malignant properties of choriocarcinoma cells. METHODS: Human choriocarcinoma cells, JAR, were transfected with either a human PP5/TFPI-2/MSPI expression vector or an empty vector, and stable clones were obtained. Messenger RNA expression, protein secretion/localization, growth rate, and plating efficiency were evaluated. In vitro migration and invasive activity were determined by transwell chamber experiments. In vivo tumor growth was evaluated by the subcutaneous injection of cells to nude mice and followed by histological examination. RESULTS: Expression of mRNA and protein of PP5/TFPI-2/MSPI were confirmed, and a high producing clone and a low producing clone were chosen for further analysis. The majority of secreted PP5/TFPI-2/MSPI protein was revealed to associate with the extracellular matrix. Expression of PP5/TFPI-2/MSPI did not affect the growth and migration of the tumor cells, but enhanced their plating efficiency. Its expression significantly inhibited invasion through the Matrigel. Invasive growth into the subcutaneous muscle layer was not evident in the nude mouse tumors of the PP5/TFPI-2/MSPI-expressing cells. CONCLUSION: PP5/TFPI-2/MSPI-expressing choriocarcinoma cells showed suppressed potential of invasion in vitro and in vivo. It is suggested that loss or suppression of PP5/TFPI-2/MSPI expression may result in the acquisition of invasiveness in choriocarcinoma cells.     

Destruxins,cyclodepsipeptides, block the formation of actin rings and prominent clear zones and ruffled borders in osteoclasts

Bone-resorbing osteoclasts exhibit polarized morphological structures such as actin rings, clear zones, and ruffled borders. To gain insight into the mechanism of bone-resorbing activity of osteoclast and to discover new types of anti-resorptive agents, we have screened for natural compounds that inhibit the bone-resorbing activity of osteoclast-like multinucleated cells (OCLs). Destruxin B (DestB) and E (DestE), cyclodepsipeptides, were found to inhibit pit formation without affecting osteoclast differentiation and survival. Destruxins reversibly induced morphological changes in OCLs in a dose-dependent manner (DestB, 0.2-1 microM; DestE, 0.01-0.05 microM) and inhibited pit formation. Destruxin-induced morphological changes were accompanied by disruption of the actin rings in OCLs. The formation of actin rings in OCLs after adhesion was also inhibited by destruxins. Electron microscopical analysis revealed that destruxin-treated OCLs on dentine slices have no prominent clear zones and ruffled borders. The effective concentrations of destruxins on the morphological changes were almost the same as those that inhibited bone resorption in organ culture system. These results suggest that the anti-resorptive effects of destruxins result from induction of a disorder of the morphological structures in polarized OCLs.     

Involvement of vacuolar H+ -ATPase in incorporation of risedronate into osteoclasts

Although osteoclasts incorporate bisphosphonates during bone resorption, the mechanism of this incorporation by osteoclasts is not known. We previously reported that bisphosphonates disrupt the actin rings (clear zones) formed in normal osteoclasts, but did not disrupt actin rings in osteoclasts derived from osteosclerotic oc/oc mice, which have a defect in the gene encoding vacuolar H(+)-ATPase (V-ATPase). The present study showed that V-ATPase is directly involved in the incorporation of risedronate, a nitrogen containing bisphosphonate, into osteoclasts. Treatment of osteoclasts with risedronate disrupted actin rings and inhibited pit formation by osteoclasts on dentine slices. Bafilomycin A(1), a V-ATPase inhibitor, inhibited the pit-forming activity of osteoclasts but did not disrupt actin rings. Risedronate failed to disrupt actin rings in the presence of bafilomycin A(1). E-64, a lysosomal cysteine proteinase inhibitor, showed no inhibitory effect on the demineralization of dentine by osteoclasts but inhibited the digestion of dentine matrix proteins without disrupting actin rings. Risedronate disrupted actin rings even in the presence of E-64. Treatment of osteoclasts placed on plastic plates with risedronate also disrupted actin rings. Bafilomycin A(1) but not E64 prevented the disruption of actin rings in osteoclasts treated with risedronate on plastic plates. Inhibition of V-ATPase with bafilomycin A(1) also prevented disruption of actin rings by etidronate, a non-nitrogen-containing bisphosphonate. These results suggest that V-ATPase induced acidification beneath the ruffled borders of osteoclasts and subsequent bone demineralization triggers the incorporation of both nitrogen-containing and non-nitrogen-containing bisphosphonates into osteoclasts.     

Effects of coffee cherry, the residue left after removal of the beans from the coffee fruit, on mammary glands, automatic behavior and related parameters in mice

To clarify the mechanisms of the anti-mammary tumor activity of coffee cherry (CC), the residue left after the removal of beans from the fruit, the effects in SHN mice of CC on plasma and urine component levels, mammary gland growth, spontaneous motor activity and several related parameters were examined. Hot water extract of CC was given to 2-month-old mice in drinking water (0.5%) for 60 days. The treatment prevented the elevation of plasma and urine levels of alanin amino-transferase and asparate aminotransferase, indicating that CC can protect against metabolic abnormality, which is a cause of the high mammary tumor susceptibility of SHN mice. It also resulted in an inhibition of the formation of precancerous mammary hyperplastic alveolar nodules. Neither food and water intake nor spontaneous motor activity was affected by CC. The findings provide novel information on the mechanism of the protective effect of CC on mammary tumorigenesis and confirm the usefulness of CC as a safe chemopreventive agent of mammary and other types of tumors.     

Doxifluridine decreases serum levels of interleukin-6 in a cancer cachexia model]

Cancer cachexia is a common paraneoplastic syndrome in patients with advanced malignancies. However, the mechanisms of the development of cancer cachexia remain to be elucidated. Interleukin (IL)-6 is known to be involved in the development of cancer cachexia. The KPL-4 human breast cancer cell line, which was recently established in our laboratory, secretes IL-6 into the culture medium. Oral administration of doxifluridine (5'-DFUR, 60 mg/kg or 120 mg/kg) significantly inhibited the growth of KPL-4 tumors, reduced the tissue levels of IL-6, and alleviated body weight loss. Serum IL-6 levels were also lowered by 5'-DFUR in nude mice bearing KPL-4 tumors. Additionally, it is suggested that tumor necrosis factor (TNF)-alpha is involved in the cachexia induced by KPL-4 tumors. We suggest that 5'-DFUR suppresses cancer cachexia by lowering IL-6 levels in the tumor tissues and serum.     

Complications related to hepatic arterial infusion chemotherapy for liver metastasis from colorectal cancer

We evaluated the complications of hepatic arterial infusion (HAI) chemotherapy in patients (pts) with hepatic metastasis from colorectal cancer. The subjects consisted of 61 pts with hepatic metastasis from colorectal cancer, who were treated by combined chemotherapy with 5-FU and CDDP weekly or continuously. Indwelling route of catheter: 30 via gastroduodenal artery (GDA) at the time of laparotomy ('LP'), 21 via femoral artery (FA) and catheter tip in PHA ('PHA'), 10 via FA and catheter tip is inserted with steel coil into the GDA ('GDA-coil'). Complications resulting in interruption of therapy occurred in 19 pts (31%), and the 'GDA-coil' method had a lower rate of complication than others. There was no difference in the incidence rate of complications between the two chemotherapy regimens. The complications of this therapy were: 8 (13%) cases of hepatic arterial occlusion, 3 (5%) cases of duodenal ulcer, 4 (7%) cases of catheter tip dislocation, 2 (3%) cases of catheter tip dislocation to the duodenal bulb, and 1 (2%) case of liver abscess. Hepatic arterial occlusion occurred frequently in LP. Up to 67% of patients with duodenal ulcer had hepatic arterial occlusion at the same time. All pts with catheter tip dislocation were 'PHA', and all pts with catheter tip dislocation to the duodenal bulb were 'LP'. In conclusion: 1. The best indwelling route for the catheter is by the 'GDA-coil' method. 2. To diagnose complications soon, regular CTA or DSA is necessary.     

Differences in apoptosis induced by anticancer drugs in sublines (SKG-3a, SKG-3b) from a human uterine cervical epidermoid carcinoma.

SKG-3a and SKG-3b are two distinct human uterine cervical epidermoid carcinoma cell lines derived from a single donor. We studied these two closely related cell lines from the standpoint of drug susceptibility. The growth inhibitory effects of cisplatin (CDDP), doxorubicin (ADM), etoposide (VP-16), and paclitaxel (taxol) on SKG-3a and SKG-3b cells assessed by crystalviolet dye uptake assay were almost the same. SKG-3b cells treated with CDDP, ADM, VP-16, and taxol showed the apoptotic cell death, whereas apoptosis in SKG-3a cells was not induced by these anticancer drugs. Caspase-3 activity was increased only in the SKG-3b cell lysate after treatment with CDDP, ADM, and VP-16 but was not found in the SKG-3a cell lysate. These results indicate that despite growth inhibitory effects of anticancer drugs being almost the same, there may be differences in the common signaling pathways involved in the apoptotic process between SKG-3a and SKG-3b obtained from the same tumor.     

Relation of hepatic protein synthesis and hepatic functional mass in obstructive jaundiced rats

In obstructive jaundiced rat, the change of hepatic functional mass assessed by [14C]-aminopyrine breath test (ABT) and galactose tolerance test (GaTT) and hepatic protein synthesis measured by [14C]-leucine incorporation into hepatic protein fraction were investigated. Bile duct ligation (BDL) for 5 and 14 days was followed by choledocho-duodenal fistula as the relief of obstruction. Hepatic functional mass measured by ABT and GaTT revealed a remarkable decrease at 5 and 14 days after BDL without differences in grades. These depressed values returned to the preoperative ones in 10 to 20 days after the relief of obstruction. On the contrary, hepatic protein synthesis was reciprocally enhanced after BDL. After the relief of obstruction the enhancement of hepatic protein synthesis was prolonged and then returned to the normal level in 20 days. These data suggested that in obstructive jaundice hepatic protein synthesis was stimulated by several stress and continued to be enhanced even after the relief of obstruction. These enhancement of hepatic protein synthesis would induce to decrease hepatic functional mass.