Yoshihiro Ishiguro

Biological and biochemical properties of Ret with kinase domain mutations identified in multiple endocrine neoplasia type 2B and familial medullary thyroid carcinoma.

Several mutations were identified in the kinase domain of the RET proto-oncogene in patients with multiple endocrine neoplasia (MEN) 2B, familial medullary thyroid carcinoma (FMTC) or sporadic medullary thyroid carcinoma. We introduced seven mutations (glutamic acid 768→aspartic acid (E768D), valine 804→leucine (V804L), alanine 883→phenylalanine (A883F), serine 891→alanine (S891A), methionine 918 →threonine (M918T), alanine 919→proline (A919P) and E768D/A919P) into the short and long isoforms of RET cDNA and transfected the mutant cDNAs into NIH3T3 cells. The transforming activity of the long isoform of Ret with each mutation was much higher that that of its short isoform. Based on the levels of the transforming activity, these mutant RET genes were classified into two groups; a group with high transforming activity (A883F, M918T and E768D/A919P) and a group with low transforming activity (E768D, V804L, S891A and A919P) (designated high group and low group). Interestingly, the level of transforming activity correlated with clinical phenotypes; high group Ret with the A883F or M918T mutation and low group Ret with the E768D, V804L or S891A mutation were associated with the development of MEN 2B and FMTC, respectively. In addition, we found that substitution of phenylalanine for tyrosine 905 present in the kinase domain abolished both transforming and autophosphorylation activities of low group Ret whereas it did not affect the activities of high group Ret.

Contrast-enhanced endoscopic ultrasonography in gallbladder diseases

BACKGROUND The purpose of this study was to investigate the usefulness of contrast-enhanced endoscopic ultrasonography in gallbladder diseases. METHODS Subjects were 38 patients including 12 with adenocarcinoma, 2 adenosquamous carcinoma, 6 cholesterol polyp, 10 cholecystitis, and 8 adenomyomatosis. After endoscopic ultrasonography, sonicated albumin was intravenously injected and the enhanced effect on images of these lesions was determined. For malignancies we compared diagnostic accuracy (T factor, TNM classification) of endoscopic ultrasonography and contrast-enhanced endoscopic ultrasonography. Vascularity as shown by contrast-enhanced endoscopic ultrasonography and angiograms was compared. RESULTS Enhancement was observed in 11 patients with adenocarcinoma but not of those with adenosquamous carcinoma or cholesterol polyp. Angiography provided hypervascular images for all cases of adenocarcinoma, but all other lesions were hypovascular. In one case of adenocarcinoma, 3 cases of adenomyomatosis, and 8 cases of cholecystitis, there was a discrepancy between contrast-enhanced endoscopic ultrasonography images and angiograms with regard to vascularity. The accuracy of depth of tumor invasion for endoscopic ultrasonography was 78.6% (11 of 14) versus 92.9% (13 of 14) for contrast-enhanced endoscopic ultrasonography. CONCLUSION Contrast-enhanced endoscopic ultrasonography is useful in the diagnosis of gallbladder lesions.

The Role of Amino Acids Surrounding Tyrosine 1062 in Ret in Specific Binding of the Shc Phosphotyrosine- Binding Domain*

We investigated the role of the I-E-N-K-L (amino acids 1057‐1061) sequence amino-terminal to Tyr 1062 in Ret for binding of the Shc phosphotyrosine-binding (PTB) domain. Substitution of Ser for Ile 1057 (I1057S), Ala for Asn 1059 (N1059A), or Pro for Leu 1061 (L1061P) in this sequence significantly decreased the transforming activity of Ret with the multiple endocrine neoplasm type 2A (MEN2A) mutation as well as the binding affinity of Shc to it in vivo and in vitro, indicating that these three amino acids play a role in Shc binding. In addition, as the RET protooncogene is translated as three isoforms of 1114 amino acids (Ret 51), 1106 amino acids (Ret 43), and 1072 amino acids (Ret 9) that differ from one another in the sequence carboxyl-terminal to Tyr 1062 , we examined whether these sequence differences influence the binding affinity of Shc to Ret. As a result, we found that the transforming activity of Ret 43 isoform with the MEN2A mutation and the binding affinity of Shc to it were very low, although the consensus sequence for the binding of the Shc PTB domain is conserved in the Ret 43 isoform. This finding suggested that the sequence carboxyl-terminal to Tyr 1062 in Ret could also influence the binding affinity to Shc. (Endocrinology 140: 3992‐3998, 1999)

An evaluation of three-dimensional ultrasonography for the measurement of gallbladder volume.

OBJECTIVE:Various three-dimensional ultrasonography systems have been developed. We estimated the accuracy of a three-dimensional ultrasonography system for measuring gallbladder volume and compared the results to the sum-of-cylinders method.METHODS:In an in vitro study, 10 balloons of various shapes, sized 5–68 mL were scanned by real-time ultrasonography. In an in vivo study, we evaluated the gallbladder emptying of 14 healthy male volunteers after ingestion of two raw egg yolks. In both studies, volume measurement was performed by the three-dimensional ultrasonography method and the sum-of-cylinders method.RESULTS:With the three-dimensional ultrasonography method, the mean difference between the measured volume and the true volume and the limits of agreement were smaller than those of the sum-of-cylinders method. Gallbladder volumes did not differ significantly with both ultrasound methods.CONCLUSION:The three-dimensional ultrasonography method accurately determined gallbladder volumes.

Development of small hepatocellular carcinoma 80 months after clearance of hepatitis C virus with interferon therapy

We describe a patient who had a complete and sustained response to interferon (IFN) therapy for chronic hepatitis C but developed small hepatocellular carcinoma (HCC) 80 months later. A 55-year-old Japanese man with hepatitis C virus (HCV) infection and histological features of chronic active hepatitis was treated with recombinant IFN alpha-2a, 9,000,000 U daily for 2 weeks followed by three times a week for 22 weeks. He successfully responded to IFN therapy with a normalization of serum alanine aminotransferase and continuous disappearance of serum HCV-RNA. However, 80 months after the cessation of IFN therapy, the patients alpha-fetoprotein (AFP) level became elevated for the first time and HCC, 12 mm in diameter, was detected by routine ultrasonographic screening. Laparotomy revealed a small HCC with no metastasis, and the nontumorous liver demonstrated chronic inactive hepatitis. This case indicates the need for careful follow-up using ultrasonography and AFP testing for at least 7 years after completing IFN therapy in all patients with chronic hepatitis C, even if the patients have a complete response to the therapy.

NON-ALCOHOLIC SCLEROSING PANCREATITIS WITH SJOGREN'S SYNDROME AND TUBULOINTERSTITIAL NEPHRITIS

We recently encountered a 56‐year‐old Japanese man with pancreatitis who had hyperglobulinemia, was autoantibody‐positive and responded to steroid therapy. The patient had been found to have asymptomatic proteinuria at an annual medical check‐up and had experienced a dry mouth for several years. He was diagnosed as having Sjögrens syndrome indicated by the dry mouth and positive findings from a lip biopsy and a Schirmers test. Tubulointerstitial nephritis, causing proteinuria, was verified by percutaneous renal biopsy. Antinuclear antibody was positive at the 1 : 160 titer. Serum γ‐globulin and IgG values were markedly increased, whereas complement components C3 and C4 were lowered. Abdominal computed tomography and ultrasonography demonstrated a diffusely enlarged pancreas without localized pancreatic mass or para‐aortic lymphadenopathy. Neither calcification nor cysts were detected in the pancreas. Endoscopic retrograde cholangiopancreatography revealed a diffuse narrowing of the main pancreatic duct with an irregular wall. Wedged biopsy specimens of the pancreas by an exploration of the abdomen showed prominent lymphocytic infiltrates including some plasmacytes, as well as decreased exocrine parenchyma and inter‐ and intralobular fibrosis. These findings suggested a diagnosis of autoimmune‐related pancreatitis. Steroid therapy was carried out with a marked improvement of his clinical symptoms and laboratory data.

7082 Pancreatoscopic laser lithotripsy (psll) of pancreatic duct stoned.

Introduction: We report the result of pancreatoscopic laser lithotripsy (PSLL) of pancreatic duct stones (PDS). Subjects and Methods: The subjects were 21 patients (18 male, 3 female) with pancreatitis by PDS at our department between 1993 and 1999. The laser used in this study was a flashlamp-pumped pulsed dye laser with a wavelength of 504nm. The laser energy was transmitted via a 320μm flexible quartz fiber. The pancreatoscope was a 1.67mm miniscope with an inner channel 0.55mm in diameter. This miniscope inserted with a 7Fr plastic sheath through a standard duodenoscope was used for transpapillary laser application under direct vision of PDS. The laser energy was transmitted to PDS through the fine laser fiber inserted via the channel. The laser pulses were applied at a repitation rate of 10Hz and an energy output of 30mJ. If the stones fragmented too slowly the energy was increased up to 100mJ provied precise targeting of PDS could be achieved. Extracorporeal shock wave lithotripsy (ESWL) was applied to the cases in which PDS could not be disintegrated completely by PSLL alone. According to therapeutic procedure, all patients were classified into three groups: PSLL group, combination (PSLL and ESWL) group, and ESWL group. We evaluated number and diameter of PDS, treatment period, total number of sessions, relief of pain and clearance of PDS for each group. Conclusion: PSLL is a very useful and effective technique in the treatment of symptomatic PDS.

7015 The role of endoscopic ultrasonography for the pancreatobiliary diseases in medical checkup.

Introduction: EUS is considered to be useful modality for the pancreatobiliary(PB) disorders.We evaluated the efficiency of EUS for PB disorders. Subjects and Methods: Subjects consisted of 874 cases suspicious of PB disorders by US-performed for 82436 persons( 4/1994 - 12/1998 at Aichi Health Care Center.). Check up criteria of US were in table 1. We made EUS exam. for above listed 874 cases, and evaluated following points ; 1.True negative rate. 2. Coincidence rate between EUS diagnosis and definite diagnosis(Table-2). 3.Unchanged period from the point diagosed as benign. Every 6 month observasion by US was done, if there was any change on US, the case was re-examined by EUS, and once a year, individual prognosis inquiry was made by mail. Results: 1. 100%. Also, the cases followed by only US remained under US criteria. 2. 96.7%, 2 cases of pancreas divisum was misdiagnosed. 3.All the cases diagosed as benign remained unchanged. Conclusion: Using EUS , we could restrict to only 0.07%(60/82436) necessary to be sent to special hospital. EUS is an efficient and accurate examination of PB diseases in mass screening.

3444 Usefulness of contrast echolymphography using endoscopic ultrasonography-guided puncture.

BACKGROUND: The accuracy of EUS in the assessment of regional lymph node metastasis leaves to be desired. Although EUS-guided fine needle aspiration biopsy (EUS-FNAB). is a superior examination method, there are various problems, such as difficulty in collecting tissue specimens and collection of tissue specimens from noncancerous regions. AIM: To qualitatively improve the capability of diagnosing lymph node swelling, contrastenhanced echolymphography (CE-EL) was performed using EUS-guided puncture. SUBJECTS AND METHOD : In the present study, 8 malignant swollen lymph nodes surgically resected from patients with gastrointestinal cancers and 36 lymph nodes obtained from 36 patients in whom abdominal lymph node swelling was indicated by EUS (14 cases of metastatic lymph node swelling and 22 cases of benign lymph node swelling) were examined. Lymph nodes were punctured under real-time EUS guidance, and carbon dioxide microbubbles were topically injected to evaluate ultrasonograms before and after microbubble injection. RESULTS: CE-EL of freshly resected metastatic lymph nodes showed nonhomogeneous patterns. Resions in the lymph node demonstrating filling defects during CE-EL were pathologically correlated with resions showing neoplastic infiltration. When the results of EUS performed before CE-EL were clinically evaluated, there were no apparent differences in any echo features except the shape of lymph nodes between the malignant and benign groups (malignant lymph nodes showed rounded shapes). In contrast, CE-EL demonstrated nonhomogeneous patterns, poor distribution of contrast medium, and filling defects in 92.9 to 100% of subjects in the malignant group. However, CE-EL demonstrated uniform patterns in 68.2% of subjects in the benign group. Therefore, there were differences in CE-EL findings between the two groups. The sensitivity, specificity, rate of positivity and negativity, and accuracy of differentiation diagnosis by CEEL were 92.8%, 95.5%, 92.8%, 95.5%, and 94.4%, respectively. CONCLUSION: The diagnostic capability is expected to be improved by combining CE-EL with EUS-FNAB. Therefore, the combination of CE-EL and EUSFNAB is considered a favorable examination method for differentially diagnosing swollen lymph nodes.

Transpapillary biopsy of the gallbladder disease

BACKGROUND Endoscopic biopsy is routine in gastrointestinal disease; similarly, histopathologic diagnosis is desirable in gallbladder disease. In this study we examined the clinical usefulness and the problems associated with transpapillary gallbladder biopsy. METHODS Transpapillary gallbladder biopsy was attempted in 9 patients with gallbladder disease. After inserting a catheter sheath into the gallbladder using a guidewire via the transpapillary route without sphincterotomy, we inserted a biopsy forceps into the lumen of the sheath up to the gallbladder lumen and obtained specimens. RESULTS We could obtain sufficient specimens for histopathologic diagnosis in 8 of 9 cases (88.9%). Of the 8 successful cases, targeted specimens were obtained in 7 (87.5%). Diagnostic accuracy with respect to malignant versus benign disease was 100% (2 of 2) and 83.3% (5 of 6), respectively. There were no complications. CONCLUSION Transpapillary gallbladder biopsy is a clinically useful technique because it facilitates histopathologic diagnosis and therefore guides the choice of therapy.