Laparoscopic liver resection with open partial resection of the thoracoabdominal wall for cancer along the transhepatic biliary drainage tract: A case report.

INTRODUCTION AND IMPORTANCE: Recently, laparoscopic liver resection has been established as a standard procedure for a certain group of hepatic resections; its indications and procedures have been expanding. CASE PRESENTATION: A 67-year-old woman was referred with a chief complaint of left subcostal pain. Eight years and nine months before, she underwent pancreatoduodenectomy for cancer of the ampulla of Vater after endoscopic retrograde biliary drainage for obstructive jaundice. A trans-anastomotic internal-external retrograde transhepatic biliary drainage (RTBD) catheter was placed intraoperatively. Based on radiological findings, intrahepatic cholangiocarcinoma, or local recurrence of cancer of the ampulla of Vater, invading the anterior thoracoabdominal wall, diaphragm, and pericardium, without distant metastasis, were considered. Laparoscopic lateral segmentectomy of the liver combined with open en bloc resection of the invaded portion of the anterior thoracoabdominal wall with costal cartilage, diaphragm, and pericardium was performed. The postoperative course was uneventful, and the patient was discharged 17 days post-surgery. The patient died of cancer recurrence 46 months later. CLINICAL DISCUSSION: The cancer invaded along the RTBD tract. Although the mechanisms of carcinogenesis remain unclear, transhepatic biliary drainage is a risk factor for cancer invasion along the tract. Laparoscopic liver resection before open thoracoabdominal wall resection ensured clear delineation of the cancer margins invading the thoracoabdominal wall and minimized the defect of the thoracoabdominal wall. CONCLUSION: Laparoscopic liver resection for hepatic cancer invading adjacent structures or organs would be feasible and useful in selected patients, even if resection of the invaded area is performed by open surgery.

Bacterial pyruvate production from alginate, a promising carbon source from marine brown macroalgae.

Marine brown macroalgae is a promising source of material for biorefining, and alginate is one of the major components of brown algae. Despite the huge potential availability of alginate, no system has been reported for the production of valuable compounds other than ethanol from alginate, hindering its further utilization. Here we report that a bacterium, Sphingomonas sp. strain A1, produces pyruvate from alginate and secretes it into the medium. High aeration and deletion of the gene for d-lactate dehydrogenase are critical for the production of high concentrations of pyruvate. Pyruvate concentration and productivity were at their maxima (4.56 g/l and 95.0 mg/l/h, respectively) in the presence of 5% (w/v) initial alginate, whereas pyruvate produced per alginate consumed and % of theoretical yield (0.19 g/g and 18.6%, respectively) were at their maxima at 4% (w/v) initial alginate. Concentration of pyruvate decreased after it reached its maximum after cultivations for 2 or 3 days at 145 strokes per minute. Our study is the first report to demonstrate the production of other valuable compounds than ethanol from alginate, a promising marine macroalgae carbon source.

Fine tuning of the redox function of Pseudomonas aeruginosa cytochrome c551 through structural properties of a polypeptide loop bearing an axial Met residue.

Pseudomonas aeruginosa cytochrome c(551) (PA) possesses a long polypeptide loop near its heme, and a unique hydrogen bond network among Ser52, axial Met61, and the heme 13-propionate side chain, i.e., Ser52 amide NH is hydrogen bonded to axial Met61 carbonyl CO, Met61 amide NH to Ser52 carbonyl CO, and Ser52 side chain OH to the heme 13-propionate side chain, contributes to stabilization of the structure of the loop [Y. Matsuura, T. Takano, R.E. Dickerson, J. Mol. Biol. 156 (1982) 389-409]. In this study, the structure and redox function of S52N and S52G mutants were characterized in order to elucidate the role of Ser52 in functional regulation of the protein. We found that the redox function of PA was hardly affected by an S52N mutation, but was slightly by an S52G one. The functional similarity between the wild-type protein and the S52N mutant demonstrated that Asn52 in the mutant plays a similar pivotal role in the formation of the unique hydrogen bond network that stabilizes the structure of the loop as Ser52 in the wild-type protein does. On the other hand, the functional alteration induced by the S52G mutation can be attributed to a structural change of the loop due to the lack of the hydrogen bond between the Gly52 and heme 13-propionate side chain in the mutant. Thus, this study demonstrated that the function of the protein can be tuned through the structural properties of the polypeptide loop near its heme.

Enhancement of the thermostability of Hydrogenobacter thermophilus cytochrome c(552) through introduction of an extra methylene group into its hydrophobic protein interior.

Careful scrutiny of the protein interior of Hydrogenobacter thermophilus cytochrome c(552) (HT) on the basis of its X-ray structure [Travaglini-Allocatelli, C., Gianni, S., Dubey, V. K., Borgia, A., Di Matteo, A., Bonivento, D., Cutruzzola, F., Bren, K. L., and Brunori, M. (2005) J. Biol. Chem. 280, 25729-25734] indicated that a void space, which is large enough to accommodate a methyl group, exists in the hydrophobic protein interior near the heme. We tried to reduce the void space through the replacement of a Val by Ile or Leu (Val/Ile or Val/Leu mutation), and then the structural and functional consequences of these two mutations were characterized in order to elucidate the relationship between the nature of the packing of hydrophobic residues and the functional properties of the protein. The study demonstrated striking differences in the structural and functional consequences between the two mutations. The Val/Ile mutation was found to cause further enhancement of the thermostability of the oxidized HT, as reflected in the increase of the denaturation temperature (T(m)) value by ∼ 3 deg, whereas the thermostability of the reduced form was essentially unaffected. As a result, the redox potential (E(m)) of the Val/Ile mutant exhibited a negative shift of ∼ 50 mV relative to that of the wild-type protein in an enthalpic manner, this being consistent with our previous finding that a protein with higher stability in its oxidized form exhibits a lower E(m) value [Terui, N., Tachiiri, N., Matsuo, H., Hasegawa, J., Uchiyama, S., Kobayashi, Y., Igarashi, Y., Sambongi, Y., and Yamamoto, Y. (2003) J. Am. Chem. Soc. 125, 13650-13651]. In contrast, the Val/Leu mutation led to a decrease in thermostability of both the redox forms of the protein, as reflected in the decreases of the T(m) values of the oxidized and reduced proteins by ∼ 3 and ∼ 5 deg, respectively, and the E(m) value of the Val/Leu mutant happened to be similar to that of the Val/Ile one. The E(m) value of the Val/Leu mutant could be reasonably interpreted in terms of the different effects of the mutation on the stabilities of the two different redox forms of the protein. Thus, the present study demonstrated that the stability of the protein is affected quite sensitively by the contextual stereochemical packing of hydrophobic residues in the protein interior and that the structural properties of the hydrophobic core in the protein interior are crucial for control of the redox function of the protein. These findings provide novel insights as to functional control of a protein, which could be utilized for tuning of the T(m) and E(m) values of the protein by means of protein engineering.

Comparison of the outcomes between anatomical resection and limited resection for single hepatocellular carcinomas no larger than 5 cm in diameter: a single-center study.

BACKGROUND/PURPOSE: Liver resection is a widely preferred treatment modality for hepatocellular carcinomas (HCCs). This study aimed to compare the survival impact of anatomical resection with that of limited resection, in patients with single HCCs no larger than 5 cm in diameter. METHODS: A cohort study was carried out on 209 consecutive patients who underwent hepatic resection for a single HCC no larger than 5 cm in diameter between January 1994 and March 2007 at Osaka City General Hospital. RESULTS: The cumulative 5-year overall survival and disease-free survival rates in the anatomical resection group (n = 111) were 71 and 40%, respectively, both of which were significantly better than the 48 and 25% seen in the limited resection group (n = 98) (P = 0.0043 and P = 0.0232, respectively). Better effects of the anatomical resection on both overall and disease-free survival were seen in patients having HCC larger than 2 cm in diameter and in patients with moderately or poorly differentiated HCC. But no significant difference in either overall or disease-free survival was seen between the groups in patients with a HCC 2 cm or less in diameter or in the patients with well-differentiated HCC. Using Cox's regression model, anatomical resection was confirmed to be an independent favorable factor for both overall and disease-free survival. CONCLUSIONS: Anatomical resection is therefore recommended for histologically advanced single HCCs ranging from 2 to 5 cm in diameter.

Role of a highly conserved electrostatic interaction on the surface of cytochrome C in control of the redox function.

In Hydrogenobacter thermophilus cytochrome c(552), an electrostatic interaction between Lys8 and Glu68 in the N- and C-terminal helices, respectively, stabilizes its protein structure [Travaglini-Allocatelli, C., Gianni, S., Dubey, V. K., Borgia, A., Di Matteo, A., Bonivento, D., Cutruzzola, F., Bren, K. L., and Brunori, M. (2005) J. Biol. Chem. 280, 25729-25734], this electrostatic interaction being a highly conserved structural feature of the cytochrome c family. In the present study, the functional consequences of removal of the interaction through replacement of Lys8 by Ala have been investigated in order to elucidate the molecular mechanisms responsible for functional control of the protein. The mutation resulted in a decrease in protein stability, as reflected in lowering of the denaturation temperature by approximately 2-9 degrees C, and a negative shift by approximately 8 mV of the redox potential (E(m)) of the protein. The decrease in the protein stability was attributed to the enthalpic loss due to the removal of the intramolecular interaction. The negative shift of the E(m) value was shown to be due to the effect of the mutation on the entropic contribution to the E(m) value. The small, but subtle, effects of removal of the conserved electrostatic interaction, occurring at approximately 1.4 nm away from heme iron, on the thermodynamic properties of the protein demonstrated not only that the interaction is important for maintaining the functional properties of the protein but also that amino acid residues relatively remote from the heme active site play sizable roles in functional control of the protein.

The First Case of Feline Infectious Peritonitis-like Pyogranuloma in a Ferret Infected by Coronavirus in Japan.

A male ferret, which was purchased from abroad at 9 months of age, had shown significant weight loss starting at 13 months of age. The ferret subsequently showed decreasing motor activity and recumbency and was euthanized at 14 months of age. At necropsy, a white, quail egg-sized mass was found in the mesentery. Histopathologically, multifocal granulomas consisting of necrotic foci, macrophages, fibroblasts and plentiful fibrous connective tissues were observed in the mesenteric mass. Surrounding the granulomas, inflammatory cell infiltration consisting of neutrophils, lymphocytes and plasmacytes was observed diffusely and significantly. Immunohistochemistry revealed small numbers of macrophages around necrotic foci that were positively stained for anti-mouse feline coronavirus. Electron microscopically, the cytoplasm of the macrophages contained viral particles, which were identified as coronavirus. The histopathological features in this ferret were similar to those in cats with feline infectious peritonitis (FIP). This was the first case in ferrets in Japan.

Effect of the redox-dependent ionization state of the heme propionic acid side chain on the entropic contribution to the redox potential of Pseudomonas aeruginosa cytochrome c551.

The thermodynamic properties of the redox potentials (E(m)) of Pseudomonas aeruginosa cytochrome c(551) (PA) and its mutants possessing a variety of pK(a) values for the heme 17-propionic acid side chain, which ranged from approximately 5 to approximately 8, have been investigated to elucidate the role of ionization of the heme side chain in the E(m) control. Since the pK(a) values of the heme 17-propionic acid side chains of the oxidized and reduced forms of PA are 5.9 +/- 0.2 and 7.0 +/- 0.2, respectively [Takayama, S. J., Mikami, S., Terui, N., Mita, H., Hasegawa, J., Sambongi, Y., and Yamamoto, Y. (2005) Biochemistry 44, 5488-5494], the ionization state of the heme 17-propionic acid side chain at physiological pH depends on the oxidation state of the protein. This redox-dependent ionization state of the heme 17-propionic acid side chain was found to have a large effect on the entropic contribution (DeltaS) to the E(m) value. The magnitude of the E(m) control through the DeltaS value due to the redox-dependent ionization state of the heme 17-propionic acid side chain was shown to be about 170 mV and hence is considerably larger than that through the enthalpic contribution (DeltaH) to the E(m) value due to stabilization of the cationic ferriheme in the oxidized protein through partial neutralization of its positive charge by the heme 17-propionate side chain, i.e., about 60 mV [Takayama, S. J., Mikami, S., Terui, N., Mita, H., Hasegawa, J., Sambongi, Y., and Yamamoto, Y. (2005) Biochemistry 44, 5488-5494]. The present study revealed that the heme 17-propionic acid side chain of the protein plays a pivotal role in the E(m) control of the protein.

Feasibility of ultrasonography in determining the diagnosis and treatment of perforated duodenal ulcer.

PURPOSE: We diagnosed and treated perforated duodenal ulcers (PDUs) based on the results of ultrasonography (US). We obtained useful ultrasonographic findings regarding the diagnosis and treatment of PDU. METHODS: We experienced 24 PDU cases over 6 years and 4 months (March 2002 to June 2008), and evaluated five useful ultrasonographic findings regarding PDU: (1) Two-way transmigration of liquid and air bubbles through the perforated duodenal wall, (2) hyperechoic band penetration out through the duodenal wall, (3) free air exiting the PDU, (4) fluid exiting the PDU, and (5) liver covering the PDU. RESULTS: Of the 24 cases, two-way transmigration was observed in 4 cases, hyperechoic band penetration was observed in 18 cases, and exiting free air and fluid was observed in all cases. In all patients with two-way transmigration of liquid and air bubbles, we could immediately diagnose PDU. For the PDU cases falling under categories 2, 3, and 4 above, upon analysis, a large majority provided useful ultrasonographic findings for diagnosing PDU. Of the 5 cases in which the liver did not cover the PDU at all, two-way transmigration of liquid and air bubbles was observed in 4 cases, and surgery was performed in all 5 cases. Of the 19 cases where the PDU was completely covered by the surrounding organs (the liver in 16 cases and fatty organs such as the round ligament or the omentum in 3 cases), nonsurgical treatment was selected in 18 cases. As a result, patients with PDUs covered by surrounding organs, mainly by the liver, received nonsurgical treatment, whereas those with noncovered PDUs underwent surgery. CONCLUSION: The findings of ultrasonography are useful for diagnosing and determining the treatment of PDU.

Local conformational transition of Hydrogenobacter thermophilus cytochrome c552 relevant to its redox potential.

In order to elucidate the molecular mechanisms responsible for the apparent nonlinear behavior of the temperature dependence of the redox potential of Hydrogenobacter thermophilus cytochrome c552 [Takahashi, Y., Sasaki, H., Takayama, S. J., Mikami, S., Kawano, S., Mita, H., Sambongi, Y., and Yamamoto, Y. (2006) Biochemistry 45, 11005-11011], its heme active site structure has been characterized using variable-temperature and -pressure NMR techniques. The study revealed a temperature-dependent conformational transition between protein structures, which slightly differ in the conformation of the loop bearing the Fe-bound axial Met residue. The heme environment in the protein structure which arises at lower temperature was found to be more polar, as a result of the altered orientation of the loop with respect to the heme due to its conformational change, than that arising at higher temperature. The present study demonstrated the importance of the structural and dynamic properties of the polypeptide chain in close proximity to the heme for redox regulation of the protein.

Further enhancement of the thermostability of Hydrogenobacter thermophilus cytochrome c552.

Thermophile Hydrogenobacter thermophilus cytochrome c(552) (HT) is a stable protein with denaturation temperatures (T(m)) of 109.8 and 129.7 degrees C for the oxidized and reduced forms, respectively [Uchiyama, S., Ohshima, A., Nakamura, S., Hasegawa, J., Terui, N., Takayama, S. J., Yamamoto, Y., Sambongi, Y., and Kobayashi, Y. (2004) J. Am. Chem. Soc. 126, 14684-14685]. The removal of a single hydroxyl group from the hydrophobic core of HT, through the replacement of a Tyr by Phe, resulted in further elevation of the T(m) value of the oxidized form by approximately 6 degrees C, the T(m) value of the reduced one remaining essentially unaltered. As a result, the redox potential of the mutant with higher stability in the oxidized form exhibited a negative shift of approximately 20 mV relative to that of wild-type HT in an enthalpic manner. These findings indicated that the redox function of a protein can be enthalpically regulated through the stability of the oxidized form by altering the contextual stereochemical packing of hydrophobic residues in the protein interior using protein engineering.

Immunohistologic attempt to find carcinogenesis from hepatic progenitor cell in hepatocellular carcinoma.

AIM: To clarify whether hepatocellular carcinoma (HCC) originates from hepatic progenitor cells and whether there is any correlation with the clinicopathologic factors of HCC, we reviewed 217 resected HCC specimens. METHODS: Immunohistochemical examination of cytokeratin (CK) 7, CK19, CD34, and CD117 (c-KIT) was performed. Overexpression of CK7 and CK19 indicates differentiation from cholangiocellular and hepatic progenitor cells, while overexpression of CD34 and CD117 indicates hepatic stem cells. Fresh specimens were obtained from 20 HCC patients for mutation of the c-KIT gene. RESULTS: CK7, CK19, and CD117 were positive in 41, 9.7, and 0.9% of the HCC specimens, respectively, and CD34 was never positive. None of the fresh HCC specimens demonstrated a c-KIT mutation. CK19 positivity was significantly correlated with a positive hepatitis B core antibody, and with poor survival outcome, and tended to correlate with poor histologic differentiation. CONCLUSION: These results suggest that: (i) about 10% of HCCs with typical histologic features originate from an intermediate hepatic progenitor cell, such as the canal of Hering and oval cells in the rat, or acquire the characteristics of cholangiocellular epithelium by metaplasia; (ii) HCC with typical histologic features rarely originates from hepatic stem cells, and (iii) patients with CK19-positive HCC have a poor prognosis.

Control of the redox potential of Pseudomonas aeruginosa cytochrome c551 through the Fe-Met coordination bond strength and pKa of a buried heme propionic acid side chain.

Pseudomonas aeruginosa cytochrome c(551) and a series of its mutants exhibiting various thermostabilities have been studied by paramagnetic (1)H NMR and cyclic voltammetry in an effort to elucidate the molecular mechanisms responsible for control of the redox potentials (E degrees ') of the proteins. The study revealed that the E degrees ' value of the protein is regulated by two molecular mechanisms operating independently of each other. One is based on the Fe-Met coordination bond strength in the protein, which is determined by the amino acid side chain packing in the protein, and the other on the pK(a) of the heme 17-propionic acid side chain, which is affected by the electrostatic environment. The former mechanism alters the magnitude of the E degrees ' value throughout the entire pH range, and the latter regulates the pK values reflected by the pH profile of the E degrees ' value. These findings provide novel insights into functional regulation of the protein, which could be utilized for tuning the E degrees ' value of the protein by means of protein engineering.

Evaluation of the effect of portal vein embolization on liver function by (99m)tc-galactosyl human serum albumin scintigraphy.

BACKGROUND: Preoperative percutaneous transhepatic portal vein embolization (PTPE) increases the safety of liver resection and improves the outcome after surgery for hepatocellular carcinoma. Scintigraphy with (99m)Tc-galactosyl human serum albumin (GSA) causes specific binding to viable hepatocytes and serves as an index of liver function. MATERIALS AND METHODS: (99m)Tc-GSA scintigraphy was performed before and 2 weeks after PTPE of the right portal vein in 16 patients. The total receptor index, reflecting overall liver function, right receptor index (right lobe), and left receptor index (left lobe) were calculated. RESULTS: After PTPE, the proportion of the volume of the nonembolized lobe (left lobe) increased (P = 0.0002). The total receptor index slightly decreased after PTPE (P = 0.090), the right receptor index decreased (P < 0.0001), and the left receptor index increased (P < 0.0001). The average increase rate in the left receptor index was 30% of the pre-PTPE value. In 2 patients with portal hypertension (> or =30 cm H(2)O) after PTPE, the left receptor index did not change. In 4 patients whose left receptor index after PTPE (including the 2 patients with portal hypertension) was <0.35, right lobectomy was not performed. CONCLUSIONS: (99m)Tc-GSA scintigraphy demonstrated that PTPE induces a shift in hepatic function from the embolized part to the nonembolized part of the liver. PTPE of the right portal vein increases the hepatic functional reserve of the left lobe as well as its volume. The changes in (99m)Tc-GSA uptake following PTPE may predict the response to liver resection.