Tomonori Kishino

Abnormalities of epidermal growth factor receptor in lung squamous-cell carcinomas, adenosquamous carcinomas, and large-cell carcinomas: tyrosine kinase domain mutations are not rare in tumors with an adenocarcinoma component.

Tyrosine kinase domain (TKD) gene mutations of the epidermal growth factor receptor gene (EGFR) have proven to be clinically significant in nonsmall‐cell lung cancer (NSCLC), particularly in adenocarcinoma. However, TKD mutations together with deletion mutations in the extracellular domain of EGFR (EGFRvIII) have not been fully investigated in NSCLC except for adenocarcinoma. The present study sought to gain further insight into the significance of EGFR mutations in NSCLC by focusing on nonadenocarcinoma NSCLC.

Visceral fat thickness in overweight men correlates with alterations in serum fatty acid composition.

BACKGROUND We examined relationships between visceral fat amount and alterations in serum fatty acid composition, both of which represent critical factors in the development of metabolic syndrome. METHODS Correlations were analyzed between visceral fat thickness as measured by ultrasonography and proportions of individual fatty acids in 21 normal-weight and 24 overweight Japanese men. RESULTS Significant associations were identified in overweight subjects. Visceral fat thickness displayed positive correlations to levels of palmitic acid and saturated fatty acids (r=0.475, P<0.05 and r=0.545, P<0.01, respectively); and negative correlations to levels of linoleic acid and polyunsaturated fatty acids (r=-0.513, P<0.05 and r=-0.428, P<0.05, respectively). Visceral fat thickness was also correlated with estimated desaturase activities, with positive correlations to Delta9- and Delta6-desaturase activities and negative correlations to Delta5-desaturase activity (r=0.580, P<0.01, r=0.669, P<0.01 and r=-0.559, P<0.01, respectively). No significant associations were identified in normal-weight subjects. CONCLUSIONS Significant associations between visceral fat amount and alterations in serum fatty acid composition were identified, but only in overweight individuals.

Low concentrations of serum n-3 polyunsaturated fatty acids in non-alcoholic fatty liver disease patients with liver injury

Tomonori Kishino*, Hiroaki Ohnishi, Kouki Ohtsuka, Satsuki Matsushima, Tsuyoshi Urata, Keiko Watanebe, Yukihisa Honda, Yoshitake Mine, Motoko Matsumoto, Kaori Nishikawa, Hideaki Mori, Shin’ichi Takahashi, Hitoshi Ishida and Takashi Watanabe 1 Department of Laboratory Medicine, Kyorin University School of Medicine, Tokyo, Japan 2 Department of Clinical Laboratory, Kyorin University Hospital, Tokyo, Japan 3 The Third Department of Internal Medicine, Kyorin University School of Medicine, Tokyo, Japan

Changes in liver fatty acid unsaturation after partial hepatectomy in the rat

The purpose of this study was to assess changes in the degree of fatty acid unsaturation in rat liver after partial hepatectomy. This is the first study in which liver fatty acid unsaturation has been analyzed over a long period of regeneration until day 28 after operation. The relationship between changes in unsaturation and fatty acid composition in the regenerating liver were also investigated in this study. Proton nuclear magnetic resonance spectroscopy revealed significantly elevated levels of unsaturation with a maximum on day 5 after partial hepatectomy, compared with untreated controls (11.72±0.55 vs. 11.05±0.26%, P<0.05). No significant changes in unsaturation were found in day 1 regenerating liver, which is rich in absolute amounts of fatty acids. Based on gas-liquid chromatography, the relative amounts of oleic acid (18∶1n−9) and linoleic acid (LA; 18∶2n−6) were increased, while polyunsaturated fatty acids such as arachidonic acid (20∶4n−6) and docosahexaenoic acid (DHA; 22∶6n−3) were decreased on day 1. On the other hand, on day 5 of regeneration, while most fatty acids were returning to their preoperative control levels, only DHA was higher than the control value (7.69±0.58 vs. 5.57±0.37%, P<0.001). The high levels of unsaturation on day 5 were found to be partly due to the increase in DHA. The findings suggest that some significant signals are transmitted during the regeneration process owing to alterations in the membrane structure by the high levels of fatty acid unsaturation and the increase in DHA levels on day 5 after partial hepatectomy.

Effect of Carryover of Clot Activators on Coagulation Tests During Phlebotomy

We investigated the effect of clot activators carried over from the serum tube on major coagulation tests during phlebotomy. First, blood specimens from 30 normal subjects were mixed with small amounts of fluid containing clot activators, and their effects on various coagulation tests were determined. Only the value of fibrin monomer complex displayed a remarkable change when thrombin-containing fluid was added to the blood specimens. Subsequently, 100 paired blood specimens (taken from 75 healthy volunteers and 25 patients taking warfarin) were collected in coagulation tubes before and after the serum tube using standard phlebotomy procedures. Various coagulation tests were performed to determine the effect of contamination of thrombin-containing blood on coagulation parameters. Differences between the 2 tubes were minimal but significant for some of the coagulation tests. Therefore, we conclude that the effect of clot activators in the serum tube on coagulation tests is minimal when standard phlebotomy procedures are used.

Downregulated ABCG2 Enhances Sensitivity to Topoisomerase I Inhibitor in Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitor-Resistant Non-small Cell Lung Cancer

Introduction: Understanding the mechanisms of drug resistance to epidermal growth factor receptor tyrosine kinase inhibitor (EGFR TKI) is essential to develop novel chemotherapies for non-small cell lung cancer (NSCLC). Therefore, we analyzed the expression and function of ATP-binding cassette (ABC) transporters in EGFR TKI-resistant NSCLC. Methods: In three newly established AG1478-resistant NSCLC cell lines, we evaluated the expression profile of ABC transporters and genotyping of ABCG2 by real-time polymerase chain reaction and elucidated their function by Hoechst dye efflux analyses. The growth-inhibitory effect of the topoisomerase I inhibitor Hoechst 33342, which is extruded by ABCG2, was also investigated in these cells using 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay. Results: In AG1478-resistant cells, significantly less ABCG2 was expressed, and the ratios of the cells with a strong ability to extrude Hoechst dye were remarkably smaller than in the parent cells. Because of the ABCG2 downregulation and loss of function due to C421A/C421A homozygosity, PC-14AG50R was thus considered to be more sensitive to Hoechst 33342 than the parental cells. All AG1478-resistant cells were more sensitive to the combination of Hoechst 33342 and AG1478 than to single agent. Conclusions: Resistance to EGFR TKI in NSCLC is associated with the downregulation of ABCG2 expression. A topoisomerase I inhibitor alone or in combination with EGFR TKI might offer a promising strategy for treating NSCLC that is resistant to EGFR TKI.

V843I, a Lung Cancer Predisposing EGFR Mutation, Is Responsible for Resistance to EGFR Tyrosine Kinase Inhibitors

Introduction: We previously demonstrated that a family predisposed to lung cancer harbored a V843I substitution in the epidermal growth factor receptor (EGFR) protein. We report here the further characterization of this mutant EGFR protein in the context of tumorigenicity and resistance to tyrosine kinase inhibitors (TKIs) of EGFR activity. Methods: Phosphorylation of EGFR and downstream signaling proteins of lung adenocarcinoma cell lines with EGFR mutations was assayed by flow cytometry. Susceptibility to TKIs of these cell lines, with or without suppression of mutant EGFR expression by small inhibitory RNA (siRNA), was investigated using a cellular viability assay. Furthermore, protein modeling was used to predict TKI binding to EGFR protein carrying the V843I mutation. Results: Phosphorylation of EGFR and downstream signaling proteins was elevated upon transfection with an EGFR gene with the V843I. Although the cell line with V843I + L858R demonstrated resistance to EGFR-TKIs, the cells became susceptible to TKIs upon incubation with siRNA specific for the V843I allele. The structural analysis suggested that TKI binding to EGFR would be sterically hindered by Arg841 in the double-mutant (V843I + L858R) EGFR. Conclusions: The V843I mutation contributes to tumorigenesis by promoting phosphorylation of EGFR and its downstream signaling proteins. This mutation also appears to provide resistance to EGFR-TKIs through structural modification of EGFR. These features are comparable with those in EGFR T790M mutation, suggesting that cases with germ-line V843I or T790M mutations could be categorized as a class of familial lung cancer syndrome with resistance to EGFR-TKIs.

Hepatocellular carcinoma containing sarcomatous lesions in a normal liver, accompanied by secondary Budd-Chiari syndrome

To the Editor: We experienced a huge tumor in an otherwise-intact liver of a 53-year-old woman without any known predisposing causes. The tumor was a moderately differentiated hepatocellular carcinoma (HCC), intermingled with sarcomatous lesions. HCC in this case was complicated by secondary Budd-Chiari syndrome (BCS). It is quite unusual that such events occur simultaneously in a patient, although the precise pathoetiological link among them remains unclear. The patient was admitted to Kyorin University Hospital complaining of abdominal distension. She had received a blood transfusion during labor at the age of 25, but had had no episode of liver damage. There was no history of alcohol abuse, exposure to chemical carcinogens, or any chronic liver diseases. On physical examination, a hard liver was palpable. Longitudinal dilated veins were visible on the abdominal wall. Laboratory examinations showed an elevated lactate dehydrogenase (LDH) level of 884 IU/L and a decreased hemoglobin value of 8.5 g/dL. Liver function tests remained within the normal limit. Hepatitis virus infection, including hepatitis B, C, or other possible viruses, was negative. HCC tumor markers such as -fetoprotein were also negative. Computed tomography revealed a huge hepatic mass in the right lobe with a mixed appearance of solid and cystic components. The solid regions were stained with contrast medium until the delayed phase of the image. Duplex Doppler ultrasonography demonstrated reversed flat flows in the right and middle hepatic veins, which reflected dysfunction of the venous outlet into the inferior vena cava (IVC) possibly due to the compression of the mass, while a normal flow was confirmed in the left hepatic vein toward the IVC. Intrahepatic collateral vessels were also recognized communicating between the middle and left hepatic veins. These findings on ultrasonography were compatible with those of BCS. However, no membranous obstruction was found in the IVC or hepatic veins. Tumor cytology revealed malignant cells with round to spindle nuclei. All these clinical data led us to suspect the mass could be a hepatic sarcoma, and a right lobectomy was performed. The tumor occupied 23 × 22 × 12 cm of the resected hepatic lobe of 36 × 23 × 15 cm (3875 g) and contained large hemorrhagic necrosis foci. Pathologic examination revealed that the tumor was primarily a moderately differentiated HCC. However, lesions of sarcomatous appearance were also observed sporadically. These lesions were composed mainly of marked pleomorphic cells, including bizarre multi-nucleated giant cells, and partly of spindle-shaped cells lacking mutual adherence, resembling so-called pleomorphic sarcoma and spindle cell sarcoma, respectively. Noncancerous liver tissue showed no signs of chronic liver diseases except that mild dilatation of the central veins were noted in Zone 3. When multiple recurrences were found in the liver 19 months later, serum liver function tests and tumor markers were again within the normal limit, including LDH and hemoglobin. All these clinicopathologic findings indicate that the HCC in the present case was of unknown etiology. Although the development of HCC is often associated with chronic liver diseases, as represented by liver cirrhosis, it is unusual that HCC develops when liver tissues are intact. However, we cannot exclude the possibility that an unknown virus infection at the time of blood transfusion at the age of 25 was responsible for the carcinogenesis. A sarcomatous appearance has been observed in 9.4% of HCC cases in an autopsy study and in 1.8% of surgically resected cases. It is noteworthy that most of these cases have been shown to be related to at least one of a number of factors including chronic liver diseases, hepatitis virus infection, or sarcomatous transformation of HCC due to anticancer therapy, none of which, however, were confirmed in the present case. In comparison with ordinary HCC, this type of tumor tends to show certain characteristics such as low or negative levels of serum tumor markers, huge mass including hemorrhagic necrosis, and poor prognosis with intrahepatic metastasis, as were found in the present case. Another remarkable feature of this case is that HCC was complicated by the presence of secondary BCS. Although primary BCS could be a carcinogenic cause of HCC, in this case, BCS was considered to be the consequence of HCC. Collateral vessels disappeared in the remaining left lobe after surgery, which also supports this view. Although HCC is a possible though rare cause of BCS, occurring in only 0.6 to 3.2% of all BCS cases, atypical features in HCC in the present case, or HCC containing sarcomatous lesions, may be related to the pathogenesis of BCS.

Unusual sonographic appearance of synovial sarcoma of the anterior abdominal wall

We report a case of synovial sarcoma in a 58‐year‐old woman arising from the anterior abdominal wall. Sonography demonstrated a large mass with an unusual complex “honeycomb” echotexture. Doppler imaging displayed increased vascularity in the solid parts of the tumor.

Meal Ingestion and Hemodynamic Interactions Regarding Renal Blood Flow on Duplex Sonography: Potential Diagnostic Implications

Splanchnic blood flow changes dramatically after meal ingestion. The present study evaluated physiologic interactions between meal ingestion and hemodynamics with respect to renal blood flow on duplex sonography, assessing the possible influence on Doppler parameters used as diagnostic criteria for renal artery stenosis. Subjects comprised 26 healthy young men (mean age: 22 ± 2 y). Sonographic measurements were made shortly after breakfast and every 1 h thereafter and were compared with values measured before the meal. Peak systolic velocity in the renal artery was elevated post-prandially, peaking at 1 h (90 ± 12 cm/s), compared with pre-prandially (73 ± 10 cm/s, p < 0.01). Similarly, acceleration time at the intra-renal segmental artery shortened to a minimum at 1 h (45 ± 5 ms) compared with baseline (51 ± 6 ms, p < 0.01). The present study indicates that renal blood flow is altered for a few hours after meal ingestion. Attention should be paid to the interpretation of data measured after meals on duplex sonography for diagnosis of renal artery stenosis.