An epidemiological survey of echinococcosis in intermediate and definitive hosts in Qinghai Province, China.

In order to understand the epidemiological status of alveolar and cystic echinococcosis in intermediate and definitive hosts in Qinghai Province, China, during the period 2007-2011, we investigated the infection in humans and animals, including yaks, Tibetan sheep, Tibetan dogs, and wild foxes distributed in different counties around the province. Sera from local residents were examined using a rapid serodiagnostic kit to detect specific antibodies against Echinococcus. Seropositive samples were confirmed with B-scan ultrasonography and X-ray examinations. Yaks and Tibetan sheep were checked at slaughterhouses, and cysts and suspicious lesions were collected for analysis. A rapid diagnostic strip was used to detect Echinococcus adults in Tibetan dogs. Positive dogs were dewormed and the parasites collected. Wild foxes were trapped and necropsies performed with particular attention to the intestine. Forty-eight of 735 (6.4%) humans tested were positive and 475 of 854 (55.6%) Tibetan sheep and 85 of 352 (24.15%) yaks were infected with Echinococcus. Across different counties, 214 of 948 (22.57%) Tibetan dogs were positive, and five of 36 (13.9%) wild foxes were infected with Echinococcus. Molecular studies showed that all the infections detected in humans, domestic yaks, and Tibetan sheep were the G1 genotype (E. granulosus), whereas the parasites from Tibetan foxes and Tibetan dogs were E. shiquicus and E. multilocularis, respectively. In conclusion, Echinococcosis is hyperendemic in Qinghai Province in both its intermediate and definitive hosts and the G1 genotype of cystic Echinococcus is the dominant strain.

A phase I/II study of cancer peptide vaccine S-288310 in patients with advanced urothelial carcinoma of the bladder.

Background: S-288310, a cancer peptide vaccine composed of two HLA-A*24:02-restricted peptides derived from two oncoantigens, DEP domain-containing 1 (DEPDC1) and M-phase phosphoprotein 1 (MPHOSPH1), was investigated in urothelial carcinoma (UC) of the bladder. Patients and methods: Thirty eight HLA-A*24:02-positive patients with progressive UC were enrolled in this study. In the phase I part of the study, three patients each were treated with S-288310 at 1 mg or 2 mg/peptide subcutaneously once a week to evaluate safety and tolerability. In the phase II, 32 patients were randomized to receive either 1 mg or 2 mg to evaluate the difference in cytotoxic T lymphocytes (CTL) induction and safety. Results: S-288310 was safe and well tolerated in the phase I. Of 27 patients evaluable for immune responses in the phase II, there was no difference in CTL induction rate between the 1 mg (100%) and 2 mg (80.0%) groups. Of 32 patients receiving S-288310 in the phase II, the most frequent drug-related AE was the injection site reaction that was observed in 29 patients (90.6%), but none of the patients discontinued administration due to these reactions and no dose relationship in the frequency and severity was observed. The objective response rate of the 32 patients was 6.3% and the disease control rate was 56.3%. The median overall survival (OS) rates for patients vaccinated with S-288310 after one regimen of chemotherapy, 2 regimens, or 3 or more were 14.4, 9.1 and 3.7 months, respectively, and 32.2% of patients post first-line treatment were alive at 2 years. OS of patients who showed CTL induction to both peptides was longer than that of those with CTL induction to no or one peptide. Conclusion: S-288310 was well-tolerated and effectively induced peptide-specific CTLs, which were correlated with longer survival for patients with UC of the bladder. Trial registration ID: JapicCTI-090980.

Impact of superselective transarterial infusion therapy of high-dose cisplatin on maxillary cancer with orbital invasion.

BACKGROUND AND PURPOSE: We have been performing the superselective transarterial infusion of high-dose cisplatin for advanced maxillary cancer since 1998 and the local control rate, disease free survival rate, and organ preservation have improved markedly compared with our former therapy. This study evaluates the effectiveness of superselective transarterial infusion therapy by using high-dose cisplatin on maxillary cancer with orbital invasion. MATERIALS AND METHODS: We treated 23 patients with maxillary cancer by using superselective transarterial infusion therapy with high-dose cisplatin and concomitant radiation therapy for 10 years. Of all patients, 15 showed orbital invasion, with 11 of these tumors fed by both internal maxillary and ophthalmic arteries. In all patients, we performed superselective transarterial infusion therapy via the internal maxillary artery and/or the other feeding branches from the external carotid artery. After the operation, we determined whether a pCR had occurred by checking for the presence of viable cells. In addition, we calculated the overall survival rate, preservation rate of the eyeball, and disease-free survival rate. RESULTS: For all 23 patients, pCR and overall survival rates were 95.7% and 78.4%, respectively. To date, 2 of these patients died of lung metastasis without local recurrence. For the 15 patients with orbital invasion, the respective pCR and disease-free survival rates were 93.3% and 87.5%. Eyeballs were preserved in all patients, and local recurrence occurred in only 1 patient, at the inferior wall of the maxillary sinus (not in the orbit). CONCLUSIONS: Superselective transarterial infusion therapy with high-dose cisplatin remarkably improved the local control rate and disease-free survival rate of maxillary cancer. Even in patients with orbital invasion, a high local control rate was achieved, with preservation of the eyeball, through infusion only into branches of the external carotid artery.

Diurnal variation and age-related changes of bone turnover markers in female Göttingen minipigs.

We investigated diurnal variation and age-related changes in bone turnover markers in female Gottingen minipigs. Ten females, 6-9 months of age, were used for confirmation of diurnal variation. Blood was collected at 3 h intervals for 24 h, and bone-specific alkaline phosphatase and intact osteocalcin (OC) levels were determined by enzyme immunoassay and radioimmunoassay, respectively. Urine was collected at 3 h intervals for 24 h using a tray attached to the bottom of the cage. The levels of N-terminal telopeptide of type I collagen (NTX) were determined by enzyme immunoassay. Pyridinoline and deoxypyridinoline were measured by high performance liquid chromatography. OC and NTX exhibited diurnal variation (Kruskal-Wallis test, P < 0.05), with the highest and lowest levels at 18:00 h (76.7 +/- 26.2 ng/ml) and 06:00 h (44.3 +/- 10.3 ng/ml), and at 03:00-05:59 h (550.4 +/- 82.4 nmol/micromol Cr) and 12:00-14:59 h (297.8 +/- 152.5 nmol/micromol Cr), respectively. In the study of age-related changes, blood and urine samples from 66 females (age range, 3-76 months) were examined to determine the bone turnover markers. All markers showed high correlations with age (0.569 < R(2) < 0.818). High levels of bone turnover markers were observed in young animals, decreasing with age (Kruskal-Wallis test, P < 0.01). The diurnal variation and age-related changes revealed in the present study will be useful in studies of bone diseases using female Gottingen minipigs.

Effect of high intensity training on anaerobic capacity of middle gluteal muscle in Thoroughbred horses.

We hypothesize that high intensity training for Thoroughbred horses that have been subjected to conventional training could further improve the metabolic properties of the middle gluteal muscle. Nine well-trained horses were subjected to high intensity (80-100% Vdot;O(2)max, 5 minx2) training for 12 weeks. Biopsy samples were obtained from the muscle before and after 4 and 12 weeks of training. Three of the 9 horses did not complete the training programme. In the remaining 6 horses, activities of succinic dehydrogenase (SDH), phosphofructokinase (PFK) and 3-hydroxy acyl CoA dehydrogenase (HAD), and the composition of myosin heavy chain isoforms were analyzed by biochemical techniques. After 12 weeks of training, a significant increase was found in PFK activity but not in the SDH and HAD activities. There were no significant changes in the composition of myosin heavy chain isoforms. The high intensity training in this study was effective at increasing glycolytic enzyme activity, indicating the possibility to improve anaerobic capacity, which potentially could contribute greatly to performance in Thoroughbred horses. This study also highlighted a fact that high intensity training should be given with the great care to prevent the skeletal muscle injuries.

Metabolic dependency of ionophore A23187-induced contraction of ileal longitudinal smooth muscle.

1. Tonic contraction in response to K+ is well known to be highly dependent on aerobic metabolism in ileal muscle. The ionophore A23187 (10(-5) M) induced an initial contraction and successive rhythmic contraction in ileal muscle, but did not induce tonic contraction. This study, therefore, was performed to investigate the metabolic dependency during contraction induced by A23187. 2. Under hypoxic conditions, A23187 (10(-5) M) induced an initial contraction accompanied by an increase in lactate release. However, it induced only small rhythmic contractions and decreases in ATP, phosphocreatinine (PCr) and glycogen contents. 3. In glucose-free medium, A23187 (10(-5) M) induced an initial contraction and concomitant significant decrease in the ATP and glycogen contents. However, it did not induce successive rhythmic contractions. In 'glycogen-depleted muscle' stimulated repeatedly with 60 mM K+ in glucose-free medium, 60 mM K+ induced a phasic contraction without tonic contraction. A23187 induced no contraction under these conditions. 4. These results suggested that the initial contraction induced by A23187 was dependent on endogenous glycogen, while successive rhythmic contractions were dependent on aerobic metabolism after supply of external glucose to the ileal muscle.

Immunohistochemical study of the neuropeptides in the stellate ganglion of the water buffalo.

The localization of some neuropeptides including neuropeptide Y (NPY), substance P (SP), calcitonin gene related peptide (CGRP), vasoactive intestinal peptide (VIP), galanin (Gal), methionine enkephalin (M-ENK), tyrosine hydroxylase (TH) immunoreactivity was studied in the stellate ganglion (SG) of water buffalo. NPY, SP, Gal and TH immunoreactivities were present in almost all of the ganglion cells. NPY, SP, Gal, SP, CGRP, VIP and M-ENK immunoreactive nerve fibers were also seen in the SG. The localization and pattern of distribution of these peptides in the water buffalo stellate ganglion were compared with those in stellate ganglia of other mammalian species.

The major low molecular weight apolipoprotein from normal and hyperlipidemia atherosclerosis-prone (LAP) Japanese quail.

The low molecular weight (LMW) apolipoprotein of apo C plays an important role in the metabolism of triglyceride-rich lipoproteins. This study aimed at a characterization of the major LMW apolipoproteins from normal quail strain, and also from LAP (hyperlipidemia atherosclerosis-prone) strain to identify its genetic disorder. The major LMW apoprotein cDNA clone from normal quail comprised of approximately 500 bp, and encoded polypeptide of 78 amino acid residues containing 57 amino acids as a mature apolipoprotein. Although the quail LMW apoprotein showed a low homology to either apo C-I, C-II, or C-III of other animals, it retained a well-developed amphipathic alpha-helix structure. There was no difference in the deduced primary structure of the quail LMW apoprotein between LAP and normal strain. An analysis of the mRNA expression showed that the quail LMW apoprotein was only expressed in the liver of both LAP and normal Japanese quail. No difference was noted in the hepatic expression of the quail LMW apoprotein mRNA between normal and LAP strains with neither normal nor atherogenic dietary conditions. The structure and expression of the major LMW apoprotein thus had no relevance to higher susceptibility of LAP strain to the experimental atherosclerosis.

Effect of treatment time on calcium antagonism by cadmium ions in a guinea-pig taenia coli.

1. When pretreated for 1 min, with Cd2+ at low concentrations (0.001-0.01 mM), there was a parallel rightward shift of Ca2+ concentration-curves in guinea-pig taenia coli in K+-depolarized Ca2+-free medium. However, when pretreated for 30 min, Cd2+ reduced the maximal Ca2+ response size. 2. The application of 0.01 mM Cd2+ for 1, 5 and 30 min in Ca2+-free, K+-medium reduced to the same degree the Ca uptake after addition of 3 mM Ca2+. The inhibitory action on the tension by Cd2+ however, became greater as the pretreatment time with Cd2+ increased. 3. Within 5 min of Cd2+ (0.01 mM) treatment, Cd2+ chiefly bound to the cell membrane, however, with a longer duration (30 min), Cd2+ entered the cytoplasm where EDTA could not reach. 4. Cd2+ above 0.0005 mM reduced dose-dependently the respiration of isolated mitochondria. 5. These results suggest that with short duration exposure (1-5 min) of taenia coli cells to Cd2+, the interference with Ca2+ entry through voltage-dependent Ca2+ channels is predominant but for longer exposure times, intracellular actions of Cd2+ contribute to its inhibitory effects.

Epitope mapping of monoclonal antibodies against N-domain of carcinoembryonic antigen.

Monoclonal antibodies (MoAbs) against N-domain of carcinoembryonic antigen (CEA), C249, K348, K1338, and K1444, that inhibit CEA-mediated cell adhesion, did not crossreact with nonspecific cross-reacting antigen (NCA). To determine amino acid sequences involved in the adhesion, epitopes of the MoAbs were mapped with recombinant NCAs carrying CEA-NCA chimeric N-domain. The data showed that the epitopes of C249, K1338, K1444 are located within the regions 1-32, 1-32, and 33-59 of CEA, respectively, and that two discrete regions 1-32 and 60-93 may be related to the epitope of K348. Comparison of amino acid sequences between CEA and NCA suggested that four residues (21, 27-29), eight residues (21, 27-29, 66, 78, 79, 89), and three residues (43, 44, 46) are important for recognition by C249 (or K1338), K348, and K1444, respectively. These residues seem to participate in the cell adhesion mediated by CEA.

Humanization of a mouse neutralizing monoclonal antibody against tumor necrosis factor-alpha (TNF-alpha).

An anti-human tumor necrosis factor-alpha (TNF-alpha) monoclonal antibody, designated as 3B10, inhibits the biological activity of human TNF-alpha. In the present study, we constructed humanized version of the antibody by grafting its complementarity-determining regions (CDRs) onto a human antibody, HBS-1. Using a molecular model of mouse 3B10, framework residues affecting the CDR conformation were identified. Thus, these residues were also introduced into the framework together with the CDRs in a stepwise manner, depending on the degree of the possible importance of the residues. As a result, one humanized version (h3B10-9) which possesses nine mouse framework residues showed the same binding activity as that of the chimeric version. This humanized anti-TNF-alpha antibody is expected to be less immunogenic and thus more suitable for possible clinical use.

Construction and expression of a mouse-human chimeric antibody against human tumor necrosis factor-alpha.

A mouse anti-human tumor necrosis factor-alpha (TNF-alpha) monoclonal antibody (MoAb), designated as 3B10, has previously been produced and characterized by our laboratory. We report here the construction and the expression of mouse-human chimeric antibody derived from the MoAb. cDNAs encoding variable regions of heavy and light chains were prepared from 3B10 cells by polymerase chain reaction, and introduced to mammalian expression vectors containing cDNA for human gamma1 and kappa constant regions, respectively. Cotransfection of the vectors into CHO cells resulted in production of antibody reacting with human TNF-alpha. In SDS-PAGE analysis, the chimeric antibody, c3B10, migrated at 170 kDa under a nonreducing condition, whereas two bands with 58 and 28 kDa appeared following treatment with 2-mercaptoethanol. Both c3B10 and mouse 3B10 neutralized the cytotoxic activity of human TNF-alpha to the same level, indicating that c3B10 holds the binding activity of its original MoAb. These findings suggest that the introduced genes for chimeric heavy and light chains are transcribed and translated to produce the chimeric heavy and light chain peptides, and that the peptides are assembled to form native IgG molecule. The chimeric anti-TNF-alpha antibody described in this study is expected to be less immunogenic and thus more suitable for possible clinical use.

Effects of aluminum ions on K(+)-induced contraction in ileal longitudinal smooth muscle.

The mechanism of the inhibition of K(+)-induced contraction caused by aluminum ions (Al3+) was analyzed in ileal longitudinal muscle and taenia coli of guinea-pig. Al3+ (2-5 mM) inhibited dose-dependently the high-K+ (60 mM, hypertonic)-induced contraction in ileal muscle. Al3+ reduced the size of the maximal response to Ca2+ without shifting the concentration-response curves to Ca2+ in taenia coli. Al3+ caused a significant decrease in Ca uptake, measured by the lanthanum method, during the K(+)-induced ileal first phase of the response. Following treatment with 5 mM Al3+ for 0.5-20 min, the ileal K(+)-induced contraction returned to a greater extent, after washing with medium containing the chelator deferoxamine, compared to normal medium. However, with increasing duration, > 30 min of Al3+ treatment, the K(+)-induced contraction did not recover, despite washing with deferoxamine. The aluminum in the cells was even more eliminated by washing with deferoxamine when the duration of Al3+ treatment was < 20 min. The present findings indicate that the inhibitory action by Al3+ on K(+)-induced contraction at the shorter treatment (< 20 min) may result from the interference of calcium permeability at the cell membrane in ileal muscle. However, with increasing duration of Al3+ treatment, Al3+ ions accumulate in intracellular compartments where deferoxamine cannot reach, and it may exert an inhibitory action on internal sites in ileal cells.

Genotyping of the CYP1A1 and GSTM1 genes in esophageal carcinoma patients with special reference to smoking.

BACKGROUND: Aromatic hydrocarbons, including benzol[a]pyrene, in tobacco smoke first require metabolic activation by phase I enzymes, cytochrome P450s (CYP450s), and then are subjected to detoxification by phase II enzymes, the glutathione-S-transferases. A high risk lung carcinoma group has been reported to have specific polymorphisms of the cytochrome P450 (CYP1A1) gene and the glutathione-S-transferase (GSTM1) gene. In this study, the authors investigated whether such genotypes were also risk factors for esophageal carcinoma. METHODS: Subjects were comprised of 89 esophageal carcinoma patients and 137 noncancer controls. Forty-nine of the patients and 60 of the control subjects were smokers. Genotypic studies of both CYP1A1 and GSTM1 were performed in the cancer tissues of all 89 patients. Genotypes of peripheral blood leukocytes taken from the control subjects were also determined. Genotypes of the CYP1A1 and GSTM1 genes were determined by the polymerase chain reaction. RESULTS: Patients who were heavy smokers with the genotypes Val/Val (V/V) for CYPIAI and the combined genotype of V/V for CYP1A1 and GSTM1- were a statistically high risk group compared with control subjects (P < 0.01, chi-square = 10.6 vs. P < 0.01, chi-square = 11.0). The association of V/V for CYPIAI with a smoking index > or = 600 in esophageal carcinoma patients was estimated at 6.63 (95% confidence interval [CI], 1.86-23.7). The association of combined genotypes of V/V of CYP1A1 and GSTM1 with a smoking index > or = 600 in esophageal carcinoma patients was estimated at 12.7 (95% CI, 1.97-81.8) CONCLUSIONS: Specific genotypes of the CYP1A1 and GSTM1- genes are related to the incidence of esophageal carcinoma, especially in heavy smokers.

Changes of response to caffeine in Ca(2+)-deficient medium in guinea-pig tenia coli.

1. After the washing out of the first caffeine, the second 25 mM caffeine-induced tension was about 50% of the first response in tenia coli. Cyclopiazonic acid (CPA), a specific sarcoplasmic reticulum Ca(2+)-ATPase blocker, almost inhibited the contraction to the next application of caffeine after the washing out caffeine in the presence of CPA. The peak tension to caffeine increased after the pretreatment with 10 mM K+ for 1 min. After pretreatment with K+ in the presence of 10(-7) M nifedipine (only phasic response), the tension to caffeine also increased. 2. The response to caffeine was rapidly reduced after incubation in a Ca(2+)-free medium and was abolished after 6 min of incubation. However, the response to caffeine in Ca(2+)-free medium 30 sec. after pretreatment with 10 mM K+ in normal Ca medium was larger. 3. These results suggest that, when the cell membrane is in the resting state, Ca2+ enters the cytoplasm by a leaky pathway and enters storage sites through Ca(2+)-ATPase. Furthermore, when the cell membrane is depolarized for a short period (less than 1 min), Ca2+ entry occurs, leading to refilling of the Ca2+ storage sites utilized by caffeine. The retention of Ca2+ in the storage sites in the Ca(2+)-free condition in tenia coli was less relative to the position in vascular smooth muscle.

Na(+)-independent glucose utilization during Mn(2+)-induced contraction in ileal longitudinal smooth muscle.

In Ca(2+)- and Na(+)-deficient, isotonic 126 mM K+ medium, addition of 5 mM Mn2+ caused a tension about 2.5 x greater than the tonic response induced by 126 mM K+ medium (Ca2+ 2.5 mM, Na+ 0 mM) in ileal muscle. When glycogen was depleted by incubation in a glucose-free, hypertonic 60 mM K+ medium, addition of 5 mM Mn2+ induced only a very weak tension in Ca(2+)-free, isotonic 126 K+ medium. Phlorizin (10(-3) M), a blocker of Na(+)-coupled glucose cotransporter and ouabain (9 x 10(-5) M), an inhibitor of Na+, K(+)-ATPase, failed to inhibit the tension elicited by 5 mM Mn2+ in a Ca(2+)- and Na(+)-deficient, isotonic 126 mM K+ medium. Mn2+ was accumulated in the intracellular compartment in a Ca(2+)- and Na(+)-deficient, isotonic 126 mM K+ medium. The tissue ATP concentration was significantly reduced in a Na(+)-deficient 126 mM K+ medium. However, it recovered almost completely when 5 mM Mn2+ was added to the isotonic 126 mM K+ medium. These results suggest that the Mn(2+)-induced contraction in depolarized ileal longitudinal muscle in Na(+)-deficient medium may be maintained by a glucose transport which is not dependent on Na+ and insensitive to phlorizin.

Involvement of protein kinase A in the subjective nocturnal rise of melatonin release by chick pineal cells in constant darkness.

Melatonin release by chick cultured pineal cells increases during the dark periods and decreases during the light periods under light-dark cycles, and this rhythmic secretion is maintained under constant conditions with a period of almost 24 hr. The mechanisms by which the circadian oscillator drives the melatonin rhythm under constant conditions have not been elucidated enough. We examined the possibility that cyclic AMP-dependent protein kinase A is involved in the subjective nocturnal increase in melatonin release by chick pineal cells cultured under constant darkness. The subjective nocturnal increase of melatonin release was suppressed dose dependently by H8 (protein kinase inhibitor) and H89 (specific protein kinase A inhibitor), but not by calphostin C (specific protein kinase C inhibitor) in static cell cultures. In a cell perfusion experiment, 9 hr pulses of H8 and H89 starting at ZT 9 (CT 11.2) hr suppressed the subjective nocturnal increase in melatonin rhythm in dose-dependent manner without causing a phase shift. An intracellular Ca2+ chelator, O,O'-bis(2-aminophenoxy)ethyleneglycol-N,N,N',N'-tetraacetic acid tetraacetoxymethyl ester (BAPTA-AM), and extracellular Ca2+ chelators, O,O'-bis(2-aminophenoxy)ethyleneglycol-N,N,N',N'-tetraacetic acid tetrapotassium salt hydrate (BAPTA) and ethyleneglycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), suppressed both the subjective nocturnal increases in melatonin release and cAMP levels dose dependently. This direct evidence strongly supports the hypothesis that cAMP-dependent protein kinase A may be involved in the subjective nocturnal increase in melatonin release by chick pineal cells and that intracellular Ca2+ plays an important role in pineal adenylate cyclase activation.

[The long-term effect and outcome of preoperative chemotherapy combined with radiation therapy for bladder cancer].

(BACKGROUND). The object of this study is to evaluate the efficacy of preoperative chemotherapy combined with radiation therapy for bladder cancer. (METHOD). A total of 44 patients with bladder cancer were treated by preoperative chemotherapy combined with radiation therapy between October, 1981 and December, 1986. Of the 44 patients, ranging in age from 40 to 82, with an average age of 65.8, 34 were male and 10 were female. Clinical stages included 4 patients in Ta, 25 in T1, 11 in T2, and 4 in T3. Each patient was treated twice with 15 gray of radiation to the small pelvic cavity and a chemotherapy combination of adriamycin, cis-platinum, tegaful, and peplomycin. The average observation time after the therapy was 83 month, with the maximum being 146 months. (RESULTS). Complete remission was included in 5 patients, partial remission in 27, and no change in 12. Thus, the overall effective rate was 72.8%. Operations, selected by the results of the preoperative therapy, included transurethral resection on 28 patients, transurethral fulguration on 2, partial cystectomy on 4, resection of tumor on 4, and total cystectomy on 3. Operations were not performed on 2 patients and not allowed on 1 patient. The outcome during the long-term follow-up included cancer related deaths in 4 patients, and death resulting from other disorders in 9. The 5-year survival rates for superficial and invasive bladder cancer were 92.4%, and 83.9%, respectively. The 10-year survival rates for superficial and invasive bladder cancer were also 92.4% and 83.9%, respectively. The 3-year and 5-year non-recurrence rates for superficial bladder cancer were 75.8%, and 66.9% respectively, according to the Kaplan-Meier method. On the other hand, the 3-year and 5-year non-recurrence rates for invasive bladder cancer were both 73.8%. During the follow-up between 9 and 11 years, 3 upper tract tumor were diagnosed (2 ureteral cancer, and 1 renal pelvic cancer). (CONCLUSION). We concluded that preoperative chemotherapy combined with radiation therapy may be effective for the treatment of bladder cancer.

Development of regulation of melatonin release in pineal cells in chick embryo.

Melatonin release in a pineal cell culture from 13- and 14-day-old chick embryos increased during the dark phase and decreased during the light phase of a 12 h light:12 h dark cycle. When the light-dark cycle was reversed, the pattern of melatonin release in the culture also reversed. 8-Bromo cyclic-AMP stimulated melatonin release in both the light and dark phases. However, no rhythm of melatonin release was detected under constant dark (DD) conditions in a cell culture from 14-day-old chick embryos. In 18-day-old chick embryos, the pineal cell culture expressed a circadian rhythm of melatonin release under DD conditions. These results indicate that mechanisms regulating melatonin synthesis in the avian pineal gland are established during embryonic life.

Epitope mapping of monoclonal antibodies to tumor necrosis factor-alpha by synthetic peptide approach.

A monoclonal antibody (mAb) against human tumor necrosis factor-alpha (TNF-alpha), designated 3B10, neutralizes biological activity of TNF-alpha, while another anti-TNF-alpha mAb 10F10 does not. In Western blot analysis, both mAbs bound to SDS-denatured TNF-alpha, indicating that the epitopes recognized by the mAbs are sequential but not conformational. To map precisely the epitopes of the mAbs, 76 overlapping octapeptides corresponding to an entire sequence of TNF-alpha were synthesized and their abilities to react with the mAbs were examined by enzyme-linked immunosorbent assay (ELISA). 3B10 bound to only one peptide at position 81-88 of TNF-alpha, SRIAVSYQ, whereas 10F10 was reactive with three overlapping peptides, ANALLANG (33-40), ALLANGVE (35-42), and LANGVELR (37-44). These results demonstrate that the 81-88 and 37-40 regions are important for the recognition of TNF-alpha by 3B10 and by 10F10, respectively. In solid-phase ELISA, 3B10 inhibited the binding of TNF-alpha to soluble TNF receptors, sTNF-RI and sTNF-RII. In contrast, 10F10 exerted little effect on the binding. TNF-alpha was detected by sandwich-type ELISA where 3B10 alone was used for both capture and detection, suggesting that 3B10 did not interfere with the trimer formation of TNF-alpha. The results obtained in this study suggest that the 81-88 region of TNF-alpha may participate in the receptor binding and that 3B10 neutralizes the activities of TNF-alpha by blocking the region.