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It has been recognized in mammals that after pancreatic duct obstruction, acinar cells progressively disappear and pancreatic islets are preserved. Previous studies by electron microscopy have suggested the involvement of apoptosis in acinar cell deletion. In the present study, we employed molecular biological methods and investigated whether acinar cell deletion is due to apoptosis. In male Sprague-Dawley rats, pancreatic duct ligation was performed through a left paramedian incision. Pancreatic tissue was studied at each of the following intervals after ligation: 3, 6, 12, 18, and 24 h and 2, 3, 5, and 7 days. DNA fragmentation was determined by in situ labeling of DNA strand breaks on tissue sections [fluorescein-labeled terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling method (TUNEL)] and by electrophoretic detection of the fragments of extracted DNA. Tissue sections were also examined by hematoxylin/eosin staining and immunohistochemical staining of insulin. Pancreatic duct ligation induced acinar cell deletion by day 5. Pancreatic tissue from control rats demonstrated no TUNEL-positive nuclei. In contrast, acinar cells from rats 12 h to 5 days after duct ligation showed TUNEL-positive nuclei. The number of TUNEL-positive nuclei was maximum 2 days after duct ligation. Electrophoresis showed DNA fragmentation after duct ligation. Control rats showed a genomic DNA pattern. Islets were preserved throughout the experimental period in duct-ligated rats and control rats. The results suggest that apoptosis may be the dominant form of acinar cell death in the rat pancreatic duct ligation model.

Exocrine pancreatic function in the early period after pancreatoduodenectomy was investigated. The effects of preoperative pancreatic duct obstruction on exocrine pancreatic function were also investigated. The volume of pancreatic juice and its amylase activity were investigated in 39 patients who underwent pancreatoduodenectomy (including pylorus-preserving pancreatoduodenectomy). The N-benzoyl-L-tyrosyl-p-aminobenzoic acid (BT-PABA) test was performed on 23 of 39 patients about 40 days after pancreatoduodenectomy. The exocrine pancreatic function was inhibited three to eight days after pancreatoduodenectomy (amylase activity: 23,700 +/- 4300 IU/day), and recovered on days 9-15 (48,000 +/- 8400 IU/day) in patients with a normal main pancreatic duct. In patients with pancreatic duct obstruction, the exocrine pancreatic function was almost eliminated (amylase activity: 440 +/- 260 IU/day) and BT-PABA test results were low (45 +/- 17%). In patients with narrowed pancreatic duct, amylase secretion was significantly inhibited even in patients with a normal number of acinar cells. There was a good positive correlation (Spearman's rank correlation coefficient, rs = 0.715, P0.01) between amylase secretion and BT-PABA test. Amylase secretion more than 10,000 IU/day is essential for a normal BT-PABA test and normal digestive function. The inhibited digestive function in patients with pancreatic duct obstruction may be due to the decreased number of acinar cells and the inhibition of exocrine pancreatic function.

65 Background: Combination chemotherapy involving S-1 and irinotecan (IRI-S) has failed to demonstrate a survival benefit over S-1 alone in metastatic gastric cancer. However, the tumor response rate was significantly higher with I-RIS. Therefore, we evaluated the effect of I-RIS as neoadjuvant chemotherapy for locally advanced gastric cancer. Methods: Patients with locally advanced gastric adenocarcinoma, T3-4, N0-3, M0, for whom curative surgery was planned after neoadjuvant chemotherapy, PS 0-1, with adequate organ function were enrolled in this study. Patients received irinotecan 80 mg/m2 on days 1 and 15 and oral S-1 80 mg/m2/day on days 1 to 21. Treatment was repeated every 28 days for 2 courses. Patients then underwent gastrectomy with lymphadenectomy. After surgery, patients resumed treatment with S-1 alone for 1 year. Results: Of the 39 patients enrolled, 37 were eligible. Two cycles of chemotherapy were completed in 34 patients, and surgery was performed in 33 patients. Of 27 RECIST-evaluable patients, 16 (59%) had a partial response and 9 (33%) had stable disease. Major grade 3 toxicities were neutropenia in 6, anorexia in 4, nausea in 3, diarrhea in 2, and fatigue in 2. Resection was performed in 32 (86%) patients and R0 resection was possible in 20 (54%) patients. The reason for R1/R2 were cy+ in 6, M1(LYM) in 5, M1(PER) in 4, M1(HEP) in 1 and PM+ in 2. Postoperative complications were observed in 13 (39%) patients. There were no treatment-related deaths. Pathological response was observed in 13 of 32 patients (41%); 2 patients had pathological CR. Median survival time was 15.9M and median progression-free survival (PFS) was 5.9M. Overall survival and PFS were significantly better in patients underwent R0 resection (p < 0.0001). Neither objective tumor response nor pathologic response predicted the survival. Conclusions: These results show that neoadjuvant S-1 and irinotecan combination chemotherapy was active and feasible for treating locally advanced gastric cancer. R0 resection is essential to achieve long-term survival. Therefore, careful diagnosis with staging laparoscopy before surgery is mandatory to avoid non-curative operation. Clinical trial information: NCT00134095.

The short-chain pseudopeptide, [D-Phe6, Leu13 psi (CH2NH)Leu14]bombesin(6-14) (RDI), is reported to be a potent antagonist of bombesin, and development of this type of compound has greatly contributed to the investigation of biological actions of bombesin and its related peptides. We recently synthesized (E)-alkene bombesin isostere by replacing the peptide bond with an (E)-double bond: [D-Phe6, Leu13 psi [(E)CH = CH]Leu14] bombesin(6-14) (EABI). The present study examined the effect of EABI on amylase release from rat pancreatic acini. EABI showed no agonistic activity at concentrations up to 1 microM, and RBI showed slight agonistic activity at concentrations > 10 nM. EABI caused a dose-dependent inhibition of amylase release stimulated by 0.1 nM bombesin, with an IC50 of 6.7 +/- 1.7 nM, and induced almost-complete inhibition at 0.3 microM. RDI caused a dose-dependent inhibition of amylase release, with an IC50 of 68.7 +/- 16 nM. EABI caused a parallel and rightward shift of the entire dose-response curve of bombesin-stimulated amylase release, and the degree of the shift was dependent on the concentrations of EABI. EABI (100 nM) and RDI (100 nM) inhibited amylase releases stimulated by gastrin-releasing peptide (1 nM) and neuromedin-C (1 nM). In contrast, amylase release stimulated by cholecystokinin octapeptide (0.1 nM), carbachol (10 microM), vasoactive intestinal peptide (1 nM), and gastrin-17 (10 nM) was not inhibited by EABI and RDI. The results indicate that EABI is a potent and specific bombesin receptor antagonist.(ABSTRACT TRUNCATED AT 250 WORDS)

Summary We have examined the effect of synthetic human cholecystokinin (CCK-33 and CCK-8) on pancreatic blood flow and protein output in anesthetized dogs. Human CCK-33 and CCK-8 increased pancreatic blood flow and protein output in a dose-related manner. There were no significant differences in increasing pancreatic blood flow between human CCK-33 and CCK-8, and increases in blood flow were closely related to the increase of the pancreatic enzyme secretion. L-364,718 (20 nmollkg) caused a potent inhibition of CCKstimulated pancreatic blood flow as well as protein output. The degree of inhibition by L-364,718 was dependent on the amount of CCK infused. This study demonstrates that increasing effect on pancreatic blood flow may be one of the biological actions of CCK mediated via CCK receptor. The CCK-33, one of longer molecular forms of CCK, is an important biological stimulator of pancreatic blood flow as well as of exocrine pancreatic secretion.

We examined in male Sprague-Dawley rats the effects of nicotine at doses of 50 (0.31 mM) and 200 mg/L (1.23 mM) given for a period of 16 weeks on body weight gain, food and fluid intake, plasma CCK, glucose and insulin levels, amylase secretory responses of isolated pancreatic acinar cells to CCK-8 and carbachol, and histopathology (gross and light microscopy) of stomach and pancreas. These parameters were re-examined further in animals treated with nicotine at doses of 200 mg/L (1.23 mM) for 12 weeks and given tap water for an additional 4 weeks to evaluate the effects of nicotine withdrawal. Metabolic data suggest that decreases in body weight gain, food and fluid intake, and plasma levels of glucose and insulin by nicotine are dose dependent. Endocrinological studies showed that the plasma levels of CCK were significantly increased with nicotine but the amylase secretory response of pancreatic acinar cells was inhibited in response to CCK-8 and carbachol. Histopathologic data revealed that treatment of animals with a high dose of nicotine enhanced the appearance of numerous vacuoles in the pancreatic acinar cell cytoplasm. When the pancreatic acinar cell morphology was closely examined, it showed evidence of pyknotic nuclei and fusion of vacuoles. Prominent loss of gastric mucosal surface was found in nicotine-treated animals with gross microscopic evidence of bleeding ulcers. All of the metabolic parameters except body weight gain were reversed upon nicotine withdrawal. In addition, plasma CCK levels and pancreatic enzyme secretion were reversed upon nicotine withdrawal.(ABSTRACT TRUNCATED AT 250 WORDS)

The receptor binding of CCK analogues was determined in terms of the inhibition of [125I]CCK binding in isolated rat pancreatic acini. The inhibition curve produced by CCK-8 showed the same feature as that produced by synthetic human CCK-33. The relative potency values of CCK analogues to half-maximally inhibit specific CCK binding were calculated; CCK-8 was equal to human CCK-33, 3-fold stronger than natural porcine CCK-33 and 39, and 700-fold stronger than the unsulphated form of synthetic human CCK-33. Our data suggest that CCK-33, one of the longer molecular forms of CCK, is as important as CCK-8 in the mechanism of physiological actions of CCK.

The effect of intravenous administration of human epidermal growth factor on the splanchnic blood flows was examined in anesthetized dogs, using an ultrasonic transit-time volume flow meter. Human epidermal growth factor (0.1, 0.5 and 1 microgram/kg) significantly increased blood flows in the portal vein (36.9 +/- 7.4% at 1 microgram/kg) and the superior mesenteric artery (49.0 +/- 16.8% at 1 microgram/kg). Systemic blood pressure monitored simultaneously was significantly decreased (8.4 +/- 1.2% at 1 microgram/kg). This study is the first to demonstrate that intravenous administration of epidermal growth factor increases the portal venous blood flow.

Clinical trial of intraoperative radiation and external-beam radiation combined with twice-weekly gemcitabine for the treatment of locally advanced pancreatic cancer

(BACKGROUND) Agonistic anti-Fas antibody (CH-11) has been reported to have little growth inhibiting potency to almost all pancreatic cancer cell lines (PCCs). DcR3 is antagonistic soluble receptor against Fas ligand, which belongs to a new TNF superfamily. Genomic amplification and mRNA expression of DcR3 have been reported in colon and lung cancers. However no information is available of the DcR3 status in PCCs. Here in the study we examined the effect of CH-11 in the presence of IFN,yand DcR3 genomic amplification and mRNA expression in PCCs. (METHORD) 1) PCCs (AsPC-1, BxPC-3, Capan-2, CFPAC-1, HPAC, MIA PaCa-2) were incubated with various concentrations of CH-11 after pretreatment with IFN~. Attached ceils were harvested and counted using Coulter Counter. 2) Genomic DNA was extracted from PCCs and quantitative PCR was performed using TaqMan system. The ratio of DNA copy number of DcR3 to beta actin was calculated. 3) mRNA was isolated from subconfluent PCCs and converted to cDNA. Quantitications of mRNA of DcR3 and beta actin were carried out with TaqMan RT-PCR. (RESULT) 1) CH-11 suppressed the growth of PCCs except for AsPC-1 to various extents when pretreated with IFN-/. 2) BxPC-3 and HPAC showed DcR3 genomic amplification (2.6 + 0.4, 2.9 + 0.4 times). Genomic amplification of DcR3 and the sensitivity to CH-11 were not correlated in PCCs. 3) Capan-2 and CFPAC-1 strongly expressed DcR3 mRNA. DcR3 mRNA expression and the sensitivity of CH-11 were not correlated in PCCs. Furthermore genomic amplification and mRNA expression of DcR3 were not correlated each other in PCCs. (CONCLUSION) Agonistic anti-Fas antibody inhibited the growth of pancreatic cancer cell lines in the presence of IFN~. Several pancreatic cancer cell lines strongly expressed DcR3 mRNA, while DcR3 genomic amplification and mRNA expression are not related to the sensitivity to CH-11 in pancreatic cancer cells. The expression of DcR3 is not involved in the inhibitory effect of CH-11 on pancreatic cancer cell growth.

The dotetracontapeptide corresponding to the revised amino acid sequence of porcine GIP (glucose-dependent insulinotropic polypeptide; originally named gastric inhibitory polypeptide) was synthesized by assembling eight peptide fragments of established purity, followed by thioanisole-mediated deprotection with trifluoromethanesulfonic acid in trifluoroacetic acid. The cycloheptyl esters of aspartic acid and glutamic acid were employed to suppress base-catalyzed side reactions. When tested in dogs, the synthetic peptide produced a significant increase of immunoreactive insulin in blood under background infusion of glucose.

教室で維持している慢性膵胃痩犬による意識下実験の成果について述べるとともに, 新しく開発した慢性胆膵胃瘻犬の作製維持の方法とその意義について述べた. 慢性膵胃痩犬は, (1) 空腹時, (2) 十二指腸内刺激前後, (3) 外因性刺激前後, (4) 食餌負荷前後の膵外分泌動態の観察検討に有用であった. 慢性膵胃瘻犬作製維持の技術, 経験から慢性胆膵胃瘻犬を開発した. 慢性胆膵胃瘻犬の作製後2ヵ月生存率は71.4%であり, また6ヵ月以上の長期生存を得た. 十二指腸内オレイン酸刺激実験により慢性胆膵胃痩犬が生理的な反応系を有することが確認され, 本モデルが今後肝胆膵胃臓器相関の研究に有用であると考えられた.

Eight peptide fragments were prepared by known amide-forming reactions as building blocks for the solution synthesis of the dotetracontapeptide corresponding to the entire amino acid sequence of human intestinal GIP (gastric inhibitory polypeptide or glucose-dependent insulinotropic polypeptide). Besides Lys(Z), Trp(Mts) and Gln-OBzl, two new amino acid derivatives, Asp(OChp) and Glu(OChp) [Mts=mesitylenesulfonyl, Chp=cycloheptyl], were employed to suppress various side reactions. These fragments were successively assembled by the azide procedure to minimize racemization and all protecting groups employed were removed from the protected GIP by using 1M trifluoromethanesulfonic acid-thioanisole in trifluoroacetic acid.Synthetic GIP exhibited a significant glucose-dependent insulinotropic activity in dogs, but failed to produce any notable anti-gastric activity in rats.

The effects of nicotine on the pH of acinar suspension, amylase release and on amylase response stimulated by carbachol were examined in isolated rat pancreatic acini. Additions of nicotine at concentrations ranging from 10 microM to 30 mM caused dose-dependent increases in pH of acinar suspension with simultaneous amylase release (p less than 0.05). There was no increase in amylase release when acinar cells were incubated with nicotine adjusted to pH 7.40. Carbachol alone released amylase whereas nicotine (pH 7.40) at a concentration of 10 mM caused a significant and nonparallel inhibition of amylase release in response to graded doses of carbachol. At concentrations ranges between 3 microM and 10 mM, nicotine at pH 7.40 inhibited amylase release stimulated by 1 microM carbachol, with a half maximal inhibition at 0.8 +/- 0.2 mM. These results indicate that in isolated rat pancreatic acini nicotine at pH 7.40 has no effect on basal nonstimulated amylase release but it inhibits carbachol-stimulated amylase response in a noncompetitive manner. These observations may have direct implications in underlying mechanism of pancreatic disorders.

特発性S状結腸穿孔は比較的稀であり, 本邦ではわれわれの2例を含めて50例を数えるのみであった.これらの多くは糞便性腹膜炎なるが故に, より重篤な臨床経過を示し, 予後不良なエンドトキシンショックへと移行する危険性が高いと思われる.われわれはこれら症例について, 穿孔の誘因, 入院時臨床所見, 手術々式とその予後等, 統計的に考察を加えたところ, 36%に白血球数の減少が認められ, しかもこれらの症例の場合には死亡率が50%にも達していた.このような臨床検査所見上の本症の特異性を把握しておくことが早期診に, ひいては治療成績の向上にもつながると思われる.

The effect of L-364,718, a cholecystokinin (CCK) receptor antagonist, on exocrine pancreatic secretion, gastric secretion, and plasma levels of gastrointestinal (GI) peptides stimulated by gastrin-releasing peptide (GRP) was examined in five conscious dogs. Intravenous infusion of graded doses of synthetic porcine GRP (18, 36, and 178 pmol/kg/h) caused significant and dose-dependent increases in pancreatic and gastric juice secretion and in plasma levels of pancreatic polypeptide (PP), CCK, and gastrin. Intravenous injection of L-364,718 (20 nmol/kg) significantly inhibited GRP-stimulated pancreatic outputs of juice volume, protein, and amylase and plasma PP release. L-364,718, however, did not affect gastric juice volume and plasma levels of CCK and gastrin. The results suggest that endogenously released CCK is, at least in part, responsible for GRP-stimulated pancreatic protein and enzyme secretions and PP release in dogs. The results further suggest that GRP-stimulated pancreatic secretion might be, in part, a direct response of GRP to exocrine pancreas.

This study was conducted to elucidate plasma cholecystokinin (CCK) and pancreatic polypeptide (PP) response after pancreatoduodenectomy and to compare response of CCK and PP in patients who had pancreatoduodenectomy with Billroth I and Billroth II type of reconstruction. Basal levels of plasma CCK were significantly lower in patients who had pancreatoduodenectomy (9.6 +/- 0.8 pmol/L) than in the control (preoperative patients: 14.6 +/- 2.0 pmol/L) probably because of the removal of the entire duodenum due to pancreatoduodenectomy, since vagotomy, which is concomitantly brought about by pancreatoduodenectomy, does not appear to interfere with release of CCK. Significant amounts of CCK (integrated CCK: 497 +/- 111 pmol-120 min/L), although less amounts than in the preoperative patients (integrated CCK: 901 +/- 167 pmol-120 min/L), were still released in response to oral fatty meal after pancreatoduodenectomy. Plasma CCK response to oral fatty meal was significantly greater in patients who had pancreatoduodenectomy with Billroth I type of reconstruction (integrated CCK: 705 +/- 153 pmol-120 min/L) than in patients who had pancreatoduodenectomy with Billroth II type of reconstruction (248 +/- 63 pmol-120 min/L). Simultaneous measurement of plasma levels of PP revealed complete abolishment of PP response by pancreatoduodenectomy. Since PP secretion can be produced by vagal stimulation, it is most likely that the decreased PP secretion is due to vagotomy rather than removal of the duodenum and pancreas. Significant amounts of CCK released after pancreatoduodenectomy, in which the main sources of release of CCK are removed, may suggest the compensatory mechanism of the remnant upper small intestine. This study also suggests the necessity of re-evaluating Billroth I type of anastomosis as a physiologic reconstruction procedure for the remnant alimentary tract after pancreatoduodenectomy.

Human GIP (gastric inhibitory polypeptide or glucose-dependent insulinotropic polypeptide) was synthesized in a conventional solution method by assembling eight peptide fragments followed by deprotection with 1 M trifluoromethanesulfonic acid-thioanisole in trifluoroacetic acid. New amino acid derivatives, Trp (Mts), Asp (OChp), and Glu (OChp) [Mts=mesitylenesulfonyl, Chp=cycloheptyl] were employed to suppress various side reactions. Synthetic GIP exhibited a significant increase of immunoreactive insulin level in dog plasma under basal ground infusion of glucose.

The effects of L364718, a new CCK receptor antagonist, on CCK-8 stimulated pancreatic secretion and PP release were examined in three conscious dogs with pancreatic fistulas. L364718 (20 nmol/kg) caused a potent inhibition of CCK-8 stimulated pancreatic protein, amylase and trypsin secretion but not of volume and bicarbonate secretion. Release of PP by CCK was also significantly suppressed by L364718. The degree of inhibition by L364718 was dependent upon the amount of CCK-8 infused. This study demonstrates that L364718 acts as a potent antagonist of CCK's action on pancreatic enzyme secretion and PP release in dogs and suggests that this agent might be a useful tool for studying the physiological role of CCK in conscious animals.

The effects of natural peptide YY on pancreatic secretion under different stimulatory conditions and on fatty acid-induced cholecystokinin release were examined in five conscious dogs with pancreatic and gastric fistulas. Intravenous infusion of natural peptide YY at 1 and 0.2 microgram/kg/hr caused 60 and 40% inhibition of secretin- and cholecystokinin-stimulated secretion, respectively, and 40-50 and 20-40% inhibition of intraduodenal oleate stimulated secretion, respectively. A significant but transient decrease in the plasma cholecystokinin level was observed in response to peptide YY under oleate stimulation. The present study demonstrates that peptide YY has a potent inhibitory effect on both exogenously and endogenously stimulated pancreatic secretion and has a mild suppressive effect on fatty acid-induced cholecystokinin release suggesting that this peptide is an important colonic inhibitor of pancreatic secretion.

This study was undertaken in male Sprague-Dawley rats to test the hypothesis that chronic ingestion of a low dose of nicotine suppresses body weight gain. The results from this study suggest that chronic nicotine ingestion induces weight loss in rats without the loss of their food intake. To determine whether the nicotine-induced body weight reductions are associated with endocrinological changes, the levels of gastrin and CCK in plasma were measured by specific radioimmunoassays and were found significantly elevated during chronic ingestion of nicotine. The data indicate that reduction of body weight mass by nicotine might be dependent on both hormonal and metabolic factors.