Effects of intrathecal and i.c.v. administration of neuropeptide W-23 and neuropeptide B on the mechanical allodynia induced by partial sciatic nerve ligation in rats.

Neuropeptide W-23 and neuropeptide B are each an endogenous ligand of GPR7. GPR7 mRNA has been detected in regions of the cortex, the hippocampus, the hypothalamus and the spinal cord in the rat. GPR7 receptor has structural features in common with both opioid and somatostatin receptors. In the present study, the effects of intrathecal and i.c.v. application of neuropeptide W-23 and neuropeptide B on the level of mechanical allodynia induced by partial sciatic nerve ligation were tested in rats. The level of mechanical allodynia was measured using von Frey filaments. Intrathecal injection of either neuropeptide W-23 or neuropeptide B attenuated the level of mechanical allodynia in a dose dependent manner at a dose between 0.1 and 10 microg, but i.c.v. injection of either neuropeptide W-23 or neuropeptide B had no effect on the level of mechanical allodynia at a dose between 3 and 30 microg. The effect of intrathecal administration of either 10 microg of neuropeptide W-23 or 10 microg of neuropeptide B was not antagonized by i.p. injection of 1 mg/kg of naloxone. Immunohistochemical examination revealed that neuropeptide W-23 was expressed mainly in the small- to medium-sized neuronal profiles in the dorsal root ganglion and that partial sciatic nerve injury decreased the percentage of neuropeptide W-23-like immunoreactivity positive neuronal profiles that were labeled by IB4. These data suggest that neuropeptide W-23 is involved in the nociceptive transmission in spinal cord and that both spinally-applied neuropeptide W-23 and spinally-applied neuropeptide B produce anti-allodynic effects in the partial sciatic nerve ligation model in rat.

Anticancer efficacy in vivo and in vitro, synergy with 5-fluorouracil, and safety of recombinant methioninase.

The elevated exogenous-methionine dependency of tumors for growth has been observed in all major cancer cell types. We have previously cloned a methioninase (rMETase) from Pseudomonas putida to deplete methionine. Growth inhibition followed by apoptotic cell death was induced by treatment of tumor cells with rMETase in vitro. A single i.p. injection of 300 units of rMETase can lower the serum methionine level in the mice from 70 microM to less than 1 microM within 2 h and maintain this depleted level for 8 h. Repeated dosing of rMETase of tumor-bearing mice could be administered without acute immune-hypersensitivity. rMETase treatment demonstrated growth inhibitory activity against human tumors in nude mice, including those which were multiple drug-resistant. No body weight loss or hematotoxicity, except a slight anemia, was found throughout the therapy. The combined treatment of the Lewis lung carcinoma with a fixed rMETase dose and increasing doses of 5-fluorouracil (5-FU) resulted in a dose-dependent enhanced antitumor efficacy for survival as well as tumor growth inhibition. Thus, methionine depletion by rMETase potentiates the antitumor efficacy of 5-FU. The data presented in this report thus indicate that rMETase is active alone, is synergistic in combination with 5-FU, and has negligible toxicity suggesting a novel clinical approach for effective cancer therapy.

Favorable survival after allogeneic stem cell transplantation with reduced-intensity conditioning regimens for relapsed/refractory follicular lymphoma.

Allogeneic stem cell transplantation (allo-SCT) is a curative option for patients with relapsed follicular lymphoma (FL). Prospective studies of reduced-intensity conditioning (RIC) have revealed that chemosensitivity at allo-SCT is the most reliable predictor of outcome; however, limited data are available for progressive/refractory disease. We report here a retrospective analysis of RIC allo-SCT for patients with FL. The purpose of this study was to elucidate the role of allo-SCT for patients with relapsed/refractory FL. We analyzed 46 patients-11 (24%) transplanted in CR, 6 (13%) transplanted in PR and 29 (63%) with progressive/refractory disease. The estimated 5-year overall survival rate was 71.6% (95% confidence interval (CI), 51.5-84.5%). According to the disease status at transplantation, the 5-year survival rate was 80.7% (95% CI, 37.7-95.4%) in the patients with CR or PR and 66.1% (95% CI, 41.5-82.3%) in those with progressive/refractory disease (P=0.29). There were no differences in relapse/progression and non-relapse mortality between the patients with chemosensitive disease and progressive/refractory disease. Allo-SCT may be a valuable treatment option, even for patients with progressive/refractory FL.

Quick quantitative analysis of gene dosages associated with prognosis in neuroblastoma.

The amplification of the N-myc gene and a gain of the chromosome 17q arm correlate with an unfavorable outcome in patients with neuroblastoma. In this study, we determined the gene dosage of the N-myc gene (located at 2p24) and Survivin gene (located at 17q25) using the p53 gene (located at 17p13) as the internal control gene by the TaqMan polymerase chain reaction (PCR)-based gene dosage analysis in 25 neuroblastoma samples. Based on the assumption that the gene dosages of each gene of a normal individual lymphocytes are 1.0, 11 of the 25 cases with a corrected gene dosage of N-myc (N-myc/p53) of more than 2.0 had a more unfavorable prognosis than the 14 cases with a N-myc gene dosage of less than 2.0 (5-year survival rate: 18 vs. 71%, P<0.01). Ten of 25 cases with a corrected Survivin gene dosage (Survivin/p53) of more than 2.0 had a more unfavorable prognosis than the 15 cases with a Survivin gene dosage of less than 2.0 (5-year survival rate: 10 vs. 67%, P<0.01). This quantitative PCR system is considered to be useful for quickly and accurately evaluating the degree of malignancy of neuroblastoma in order to select the optimal treatment.

A microsatellite instability analysis in neuroblastoma based on a high resolution fluorescent microsatellite analysis.

It has recently been reported that mismatch repair enzymes, which are one type of DNA repair enzymes, are the causative genes for a major group of hereditary non-polyposis colon cancers (HNPCC). Abnormalities in the mismatch repair system can be monitored by observing instability at the microsatellite loci (MSI) in cancer cells. MSI has been reported not only in tumors associated with hereditary non-polyposis colorectal cancer but also in sporadic forms of various tumors. No correlation between pediatric malignant tumors and the mismatch repair system has yet been reported. In the present study, we examined the frequency of MSI in 21 neuroblastomas, which are the most common solid tumors in childhood, using a high resolution fluorescent microsatellite analysis. MSI on five microsatellite loci was detected in none of the 21 samples. Other mechanisms independent of mismatch repair deficiency may thus play a role in both tumorigenesis and the development of neuroblastoma.

Three putrescine bisamides from the leaves of Aglaia grandis.

Three putrescine (i.e. 1,4-butanediamine) bisamides were isolated from the leaves of Aglaia grandis. Their structures were elucidated by interpretation of spectral data.

Evoked potential abnormalities in postoperative patients with biliary atresia.

The somatosensory evoked potentials from the lower extremities were measured postoperatively in 15 patients with biliary atresia to investigate whether they were free of neurologic dysfunction. Because long-standing cholestasis causes progressive neuropathy due to malabsorption of vitamin E, the serum vitamin E, D, and A levels were also examined to evaluate the fat-soluble vitamin status. The cerebral evoked potentials to posterior tibial nerve stimulation were recorded in all 15 patients as well as in 45 controls, but spinal evoked potentials examined at the level of the cauda equina could not be recorded in five patients more than 8 years of age with long-standing icterus. The remaining 10 patients exhibited spinal evoked potentials as observed in the controls, but the mean neural conduction velocity at the cauda equina was significantly lower than that of the 45 controls (42.0 +/- 5.1 m/s vs 52.3 +/- 6.8 m/s, P = .0002). The serum vitamin E, D, and A levels were within the normal range in 13, 9, and 1 of 15 patients, respectively. These results suggest that the patients with long-term follow-up are still at risk of developing neural disturbances even with normal serum vitamin E status.

(2R-trans)-2-butyl-5-heptylpyrrolidine as a potent sigma receptor ligand produced by Streptomyces longispororuber.

A potent sigma (sigma) receptor ligand was isolated from the culture broth of Streptomyces longispororuber #525. The active compound was identified to be (2R-trans)-2-butyl-5-heptylpyrrolidine by spectroscopic and chemical studies. The compound exhibited high affinity and selectivity for sigma receptors. The IC50 values toward sigma1, sigma2 and dopamine D2 receptors were 2.0, 22.7 and more than 40,000 nM, respectively. Its (2S-trans)- and (+/-)-cis-isomers, both synthesized, were also found to be high affinity sigma ligands.

Clinical implications of minimal disease in the bone marrow and peripheral blood in neuroblastoma.

PURPOSE: In patients with neuroblastoma (NB), minimal disease (MD) in bone marrow (BM) and peripheral blood (PB) is thought to play an important role in metastasis. The current study was designed to evaluate the clinical implications of the detection of MD in NB at the initial diagnosis. METHODS: Expression of the neuroendocrine protein gene product 9.5 (PGP9.5) and tyrosine hydroxylase (TH) mRNA in BM and PB obtained from 18 patients with NB was investigated by reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: MD was detected in the BM obtained from 4 of 14 localized NB patients and also in the PB from 2. However, it was found also in both the BM and PB obtained from all 4 patients with metastatic NB. Two of the 4 MD-positive patients with localized NB had metastatic recurrence after a complete tumor excision. They also had unfavorable biological prognostic factors compared with the other 2 who did not have recurrent disease. CONCLUSION: MD detected by RT-PCR in the BM and the PB of patients with NB thus suggests a risk for metastatic disease, which in association with other unfavorable biological features may indicate a poor prognosis.

Analysis of the prognostic factors relating to better clinical outcome in ganglioneuroblastoma.

PURPOSE: To elucidate the precise reason for the better prognosis in ganglioneuroblastoma (GNB) than in neuroblastoma (NB), the prognostic factors (age at diagnosis, stage at diagnosis, primary site, N-myc amplification, Shimada classification, and ploidy) were analyzed. METHODS: A retrospective analysis of the 57 neuroblastoma cases (20 GNB cases and 37 NB cases), that had not been detected by mass screening over the past 19 years at the authors' institute, was carried out. RESULTS: A Kaplan-Meier analysis of the 5-year survival rates were 67.2% and 35.1% in cases of GNB and NB, respectively, and these rates were significantly higher in GNB (logrank test; P = .04631). No significant differences were seen between GNB and NB regarding the rate in patients 1 year of age or older (95.0% v 78.4%; P = .1005), the rate of advanced cases (60.0% v 78.4%, P = .1406), the rate of an unfavorable histology (Shimada classification, 54.5% v 68.4%, P = .4473), or the diploid pattern rate (75.0% v 76.9%, P = .9200). However, N-myc amplification was found exclusively in NB (amplified cases per examined cases; 15/31), and all cases with N-myc amplification died. When the 5-year survival rate was evaluated in the patient without N-myc amplification between GNB and NB, no significant difference was observed (logrank test; P = .8568). CONCLUSION: The better prognosis in patients with GNB was thus thought to be exclusively related to an absence of N-myc amplification.

Influence of age on the presentation and outcome of choledochal cyst.

PURPOSE: The aim of this study was to identify the most appropriate timing for surgery in newborns with choledochal cysts. METHODS: The clinical and histological data of 8 newborn cases of choledochal cysts (newborn group) were compared with 45 cases that were diagnosed later than the newborn period (late group). RESULTS: The mean and standard deviation of age at diagnosis was 0.4+/-0.4 months and 43.4+/-37.8 months in the newborn and late group, respectively. The age at operation was significantly earlier in the newborn group than in the late group (4.9+/-4.5 months v54.7+/-47.0 months). Although no significant difference in the time that elapsed between the operation and the onset of symptoms between the newborn and late groups (4.5+/-4.7 months v11.3+/-21.1 months), the serum bilirubin level (4.6+/-3.8 mg/dL v1.8+/-3.4 mg/dL) and the grade of liver fibrosis (2.0+/-0.8 v1.1+/-0.8) were significantly higher in the newborn group than in the late group. CONCLUSION: The newborn group should be considered as a special group of patients with different clinical course and pathology than those of the late group, which necessitate the early surgical intervention to prevent progression of liver fibrosis.

Highly sensitive analysis for N-myc amplification in neuroblastoma based on fluorescence in situ hybridization.

BACKGROUND/PURPOSE: The N-myc amplification status in neuroblastoma has been evaluated previously for the whole tumor by the Southern blot method. The aim of this study is to evaluate the effectiveness of the fluorescence in situ hybridization (FISH) method to analyze N-myc amplification in neuroblastoma and compare the findings with those using the Southern blot method. METHODS: In 26 neuroblastoma primary tumors and metastatic lesions, the N-myc amplification status was evaluated by both the Southern blot method and FISH method. RESULTS: Of the 22 samples with no N-myc amplification using Southern blot, no cells with N-myc amplification using FISH were present in 21 of the samples. However, one metastatic liver lesion showed 16% of the nuclei to display more than 10 copies of N-myc based on FISH analysis. In the 4 remaining samples with N-myc amplification using the Southern blot method (17 copies, 15 copies, 6 copies, and 3 copies), the rates of cells with more than 10 copies of N-myc based on a FISH analysis were 79%, 68%, 94%, and 9%, respectively. CONCLUSIONS: The FISH method can detect more accurately N-myc amplification than the Southern blot method either when the rate of cells with N-myc amplification is low or intratumor heterogeneity is present.

Bile ductular proliferation as a prognostic factor in biliary atresia: an immunohistochemical assessment.

PURPOSE: The correlation between the histological findings of the intrahepatic biliary epithelium and postoperative bile drainage in biliary atresia (BA) was investigated. METHODS: The patients with BA were classified into 2 groups, consisting of a good bile drainage group (GBD, n = 14, mean age at initial operation, 57.6+/-18.0 days) and a poor bile drainage group (PBD, n = 11, mean age at initial operation, 86.9+/-42.7 days). Liver specimens from an initial Kasai's operation were examined by immunostaining using anticytokeratin 7 (CK7) antibody and anti-MIB-1 antibody. The number of CK7-positive cells in the bile ductules was microscopically calculated within the 40-microm-thick interstitium along the limiting plate (LP), and the CK7-positive cell number per unit length of the LP was estimated. In addition, the MIB-1 index in bile ductules also was determined. RESULTS: The number of CK7-positive cells in PBD was significantly higher than that in GBD (167.6+/-45.6 v 117.8+/-32.4/ mm, P<.05). However, the MIB-1 index in biliary cells did not differ between the 2 groups. CONCLUSION: An increased number of intrahepatic bile duct epithelial cells in liver specimens at the initial operation may be a poor prognostic factor in BA and appears to depend on the duration of bile stasis rather than the degree of bile stasis.

Long-term results and clinical problems after portoenterostomy in patients with biliary atresia.

From January 1962 through December 1986, 71 cases of biliary atresia were operated on. Nine of these patients underwent only an exploratory laparotomy. A hepatic portoenterostomy (Kasai's operation) was performed on 62 infants with biliary atresia. Of these 62 children, 18 survived more than 10 years. However, 4 of the 18 died shortly thereafter because of persistent or recurrent jaundice and hepatic failure. Fourteen patients, however, are still alive more than ten years after portoenterostomy, today the actual 10-year survival rate is thus 22.5%. Six of them have no severe complications and their serum total bilirubin is less than 1.0 mg/dl. However, 8 patients, whose serum total bilirubin is more than 1.0 mg/dl, demonstrated numerous clinical problems such as esophageal varices, hypersplenism and recurrent jaundice after cholangitis. One of the patients in the latter group eventually underwent a liver transplantation because of the recurrent jaundice and hepatic insufficiency. In the treatment of biliary atresia, a close and careful follow-up is therefore essential especially for patients whose serum total bilirubin is more than 1.0 mg/dl.

Lung cancer in a child with a substantial family history of cancer.

The occurrence of primary lung cancer is rare in childhood. The case of an 11-year-old boy with primary lung cancer is presented in this report. He had a substantial family history of cancer. His chief complaint was coughing with right chest pain. A chest radiograph showed a coin lesion in the right lower lung. A right lower lobectomy revealed a squamous cell carcinoma (stage IIIA at Japanese TNM classification). Systemic chemotherapy using cisplatin, vindesine, THP-adriamycin and cyclophosphamide was performed. Six months after surgery, a recurrent tumor occurred. An analysis of the familial cancer related genes (p53 gene and mismatch repair gene) showed no abnormality.

Hazards of hepatic portocholecystostomy in biliary atresia.

A hepatic portocholecystostomy (HPC) has been recommended to avoid postoperative cholangitis in the case of a patent distal extrahepatic bile duct (PDEBD) for the treatment of biliary atresia (BA). We investigated the efficacy and clinical problems of HPC in BA. The clinical records of eight patients with BA and PDEBD were reviewed. The diameter of the common bile duct was compared between the patients with BA and PDEBD and age-matched patients with neonatal hepatitis (NH). Five of 8 patients with PDEBD underwent HPC. One patient had to be converted to a cholecystojejunostomy because of common bile duct stenosis at 19 years of age. The other two patients underwent a reoperation by a hepatic portojejunostomy due to poor bile drainage after HPC. Another patient became jaundice-free one month after HPC, but died of sepsis due to bile leakage 3 months thereafter. The mean diameter of the common bile duct in BA with PDEBD was significantly smaller than that of NH (0.76 +/- 0.16 mm (n = 8) in BA vs. 1.90 +/- 0.39 mm (n = 11) in NH, p<0.01). HPC was thus found to be an excellent operative method for preventing postoperative cholangitis in BA, however, many clinical problems still need to be overcome for such a narrow distal duct.

[Antiangiogenic and antitumor effect of BPHA, an orally-active matrix metalloproteinase inhibitor, in in vivo murine and human tumor model].

We examined the antiangiogenic and antitumor efficacy of a newly-developed matrix metalloproteinase (MMP) inhibitor, BPHA (N-biphenyl sulfonyl-phenylalanine hydroxiamic acid). BPHA potently inhibits MMP-2, 9 and 14 but not MMP-1, 3 or 7. In contrast, (-)BPHA, an enantiomer of BPHA, was inactive against all MMP tested. Daily oral administration of BPHA in mice resulted in potent inhibition of tumor-induced angiogenesis, primary tumor growth and liver metastasis, whereas (-)BPHA did not. These results demonstrate that selective MMP inhibition is correlated with antiangiogenic and antitumor efficacy and that the selective MMP inhibitor BPHA has therapeutic potential without hematotoxic effect or loss of body weight.

The progressive degeneration of interlobular bile ducts in biliary atresia: an ultrastructural study.

The ultrastructure of the interlobular bile ducts were observed in hepatic specimens obtained at surgery in ten patients with biliary atresia, in order to investigate the progress of their degeneration over time after the onset of the disease. As an index for the passage of time, the degree of fibrosis in the portal tracts was investigated. Then the ultrastructural features in the interlobular bile ducts were contrasted with the grading of portal fibrosis Generally, the ultrastructural changes of the interlobular bile ducts were more marked in the cases with severe portal fibrosis than that observed in the cases with mild fibrosis. This correlation suggests that the degeneration of the interlobular bile ducts progressively worsens over time after the obliteration of the extrahepatic bile duct.

[Changes in pancreatic hormones in portal blood and morphological changes of islet cells of Langerhans after hepatectomy].

Hybrid adult dogs weighing 12 to 15 kg were laparotomized before, one week, and one month after partial hepatectomy: Pancreatic tissues were taken and examined by the PAP staining. Also, the animals were intravenously injected via peripheral veins with 50 percent glucose 0.5 g/kg before, one week, and one month after surgery, and their portal blood was taken before, 5, 15, 30, 45 and 60 minutes after injection, and submitted to examination of insulin, glucagon, somatostatin and blood sugar levels. We obtained these results as follows. sigma IRI which represents an increment of insulin secretion after glucose loads, showed a decrease at one week after surgery (p less than 0.05) and an increase at one month after surgery (p less than 0.05). Somatostatin levels of portal blood showed marked increase after surgery (p less than 0.05). Observation on the area of Langerhans islets revealed approximately 4.6 fold hypertrophy from the size of 486 +/- 23 microns m2 before surgery to 2236 +/- 98 microns m2 one month after surgery (p less than 0.05). Insulin secreting B cells were found to contain increased secreting granules. Somatostatin secreting D cells increased 2.8-fold in number one month after surgery, showing mitotic figures.