Availability of a Magnetic Method for Hepatocyte Transplantation.

INTRODUCTION: Hepatocyte transplantation is a promising alternate for the treatment of hepatic diseases. Hypothermic preservation of isolated human hepatocytes is potentially a simple and convenient strategy to provide on-demand hepatocytes in the quantity sufficient and the quality required for biotherapy. Isolated fresh hepatocytes include damaged cells that are also early apoptotic cells, which is not ideal for hepatocyte transplantation. However, this does not reflect cell viability, although it is considered that it adversely affects cell survival after transplantation. We aimed to harvest these hepatocytes and filter the apoptotic cells using a magnetic method to provide a transplantation source. MATERIALS AND METHODS: Rat hepatocytes were isolated from caudate lobes using manual enzymatic perfusion. The hepatocyte yield was 5.3 ± 0.66 × 109 cells/g of liver tissue, with a viability of 82.3 ± 3.5%. Two samples of hepatocytes were freshly isolated, one using the magnetic method, and the other without. The magnetic method was performed using DynaMag-15 Magnet, and Annexin V Antibody was used on the early apoptotic cells. We evaluated the viability and plate efficiency of the cells after 24 hours at 37°C. Hepatocytes were isolated using cell separation method, and 30 × 106 cells were mixed with 1.0 mL of Dulbecco's Modified Eagle's Medium (DMEM) and directly injected into the spleen of Lewis rats (150-250 g) using 24-gauge needles. Blood samples were collected on days 0, 3, 7, and 14, and the blood albumin level was measured using enzyme-linked immunosorbent assay (ELISA):G1, control (medium injection); G2, fresh hepatocyte transplant using the magnetic method; and G3, fresh hepatocyte transplant without the magnetic method. RESULTS: The viability was 84.9 ± 2% for fresh hepatocytes and 80.7 ± 1.2% for hepatocytes isolated using the magnetic method. The magnetic method does not damage the cells (73.5 ± 2% vs 35.2 ± 2% after 24 hours), preserving hepatocyte. The albumin level accepted significantly increased in the magnet-treated group compared with the nonmagnet group. Simultaneously, the spleen in which these hepatocytes were transplanted could be used to observe the hepatocytes; the cells were transplanted 14 days later, and the magnet-treated group had significantly higher levels of hepatocytes than the nonmagnet group. CONCLUSION: We developed an effective technique for hepatocyte isolation for short-term preservation. As a result, we believe that transplantation not only improves the cell transplantation effect but also allows the cells to be stored efficiently using the magnetic method. These results demonstrate the usefulness of hepatocyte hypothermic preservation for cell transplantation.

A quantitative evaluation of fluid leakage around a polyvinyl chloride tapered endotracheal tube cuff using an in-vitro model.

INTRODUCTION: This study was designed to quantitatively evaluate the sealing effect of a polyvinyl chloride tapered endotracheal tube cuff and to compare the tapered cuff with cylindrical endotracheal tube cuffs using an in vitro viscous fluid model. METHODS: Five types of 8.0 mm inner diameter endotracheal tubes (TaperGuard, PortexSacett, PortexSoftseal, Sheridan HVT, Sheridan CF) were fixed in vertically placed 20mm internal diameter acrylic tubes. The cuffs were inflated to 25 mmHg pressure and water was added to the top of the cuff. The amount of water leaking around the cuff at five minutes was measured. Afterwards a viscous fluid was poured above the cuff and the amount of fluid passing around the cuff at five minutes and four hours was measured. Each determination was repeated five times and the data analyzed. RESULTS: The median (range) amount (ml) of dyed water passing around the cuff at 5 minutes was TaperGuard tube 0 (0-0.2), PortexSacett 81.1 (44.6-107.9), PortexSoftseal 95.4 (91.4-113.7), Sheridan HVT 46.5 (32.7-74.6) and Sheridan CF 52.6 (31.9-62.2), suggesting that TaperGuard significantly (P<0.05) reduced fluid leakage. The viscous fluid leakage at 5 minutes was not significantly different among the groups while the amount of leaked viscous fluid at 4 hours was 0.2 (0-1.1) for the TaperGuard, 57.6 (11.9-117.4) for the PortexSacett, 23.2 (13.8-33.0) for the PortexSoftseal, 5.5 (2.7-15.4) for the Sheridan HVT and 5.6 (1.8-7.9) for the Sheridan CF. CONCLUSION: An endotracheal tube with a tapered polyvinyl chloride cuff had significantly less fluid leakage compared to four tubes with cylindrical cuffs.

Robot-assisted laparoscopic pancreaticoduodenectomy.

BACKGROUND: Robotic surgery is the most advanced development in minimally invasive surgery. However, the number of reports on robot-assisted endoscopic gastrointestinal surgery is still very small. In this article, we describe total laparoscopic pancreaticoduodenectomy (PD) undertaken using the da Vinci Surgical System® (Intutive Surgical). METHODS: Three patients underwent robotic PD between November 2009 and February 2010. Following resection of the pancreatic head, duodenum, and the distal stomach, intracorporeal anastomosis was accomplished by Child's method of reconstruction, which includes a two-layered end-to-side pancreaticojejunostomy, an end-to-side choledochojejunostomy, and a side-to-side gastrojejunostomy. RESULTS: The time required for surgery was 703 ± 141 min, and blood loss was 118 ± 72 mL. The average hospital stay period was 26 ± 12 days. As a postoperative complication, pancreatic juice leak occurred in one case, but it was managed with conservative treatment. Of the three patients, one had cancer of the papilla of Vater, one had cancer of the pancreatic head, and one had a solid pseudopapillary neoplasm. In all cases, the surgical margin was negative for tumor. CONCLUSIONS: Robot-assisted PD required a long time, but organ removal with less bleeding was able to be safely performed owing to the high degree of freedom associated with the forceps manipulation and the magnified view. Similarly, pancreatojejunostomy could certainly be conducted. No major postoperative complications were found. Accumulation of da Vinci PD experience in the future will lead to safer and faster PD.

STAT3 inhibitor WP1066 as a novel therapeutic agent for renal cell carcinoma.

BACKGROUND: Signal transducer and activator of transcription 3 (STAT3) regulates the expression of genes that mediate cell survival, proliferation, and angiogenesis and is aberrantly activated in various types of malignancies, including renal cell carcinoma (RCC). We examined whether it could be a novel therapeutic target for RCC by using the STAT3 inhibitor WP1066. METHODS: The antitumour activities and related mechanisms of WP1066 were investigated in vitro on renal cancer cell lines and in vivo on murine xenografts. RESULTS: In Caki-1 and 786-O renal cancer cells, 5 muM WP1066 prevented the phosphorylation of STAT3, and 2.5 muM WP1066 significantly (P<0.01) inhibited cell survival and proliferation. WP1066 suppressed the expression of Bcl-2, induced apoptosis, and inhibited the basal and hypoxia-induced expression of HIF1alpha and HIF2alpha, as well as vascular endothelial growth factor secretion into cell culture medium. Human umbilical vascular endothelial cells cocultured with media from WP1066-treated cells showed significantly reduced tubulogenesis (P<0.05). Systemic oral administration of WP1066 to mice for 19 days significantly inhibited the growth of Caki-1 xenograft tumours (P<0.05), and pathological analysis of xenografts of WP1066-treated mice showed decreased immunostaining of phosphorylated STAT3 and reduced length of CD34-positive vessels (P<0.05). CONCLUSION: Our results suggest that using WP1066 to inhibit the STAT3 signalling pathway could be a novel therapeutic strategy against RCC.

Neutral endopeptidase inhibits prostate cancer tumorigenesis by reducing FGF-2-mediated angiogenesis.

Neutral endopeptidase (NEP) is a cell surface peptidase that catalytically inactivates a variety of physiologically active peptides including basic fibroblast growth factor (FGF-2). We investigated the effect of using lentivirus to overexpress NEP in NEP-deficient DU145 prostate cancer cells. Third-generation lentiviral vectors encoding wild-type NEP (L-NEP), catalytically inactive mutant NEP (L-NEPmu), and green fluorescent protein (L-GFP) were stably introduced into DU145 cells. FGF-2 levels in cell culture supernatants decreased by 80% in L-NEP-infected DU145 cells compared to cells infected with L-NEPmu or L-GFP (P<0.05) while levels of other angiogenic factors were not altered. In vitro tubulogenesis of human vascular endothelial cells induced by conditioned media from DU145 cells infected with L-NEP was significantly reduced compared with that from DU145 cells infected with L-GFP (P<0.05). Tumor xenografts from L-NEP-infected DU145 cells were significantly smaller compared to control cell xenografts and vascularity within these tumors was decreased (P<0.05). Our data suggest that stable expression of NEP in DU145 cells inhibits prostate cancer tumorigenicity by inhibiting angiogenesis, with a probable mechanism being proteolytic inactivation of FGF-2.

Lentiviral vector neutral endopeptidase gene transfer suppresses prostate cancer tumor growth.

Neprilysin (neutral endopeptidase, NEP) is a cell surface peptidase whose expression is lost in approximately 50% of prostate cancers (PC). NEP normally functions to inactivate peptides such as bombesin and endothelin-1, and potentiates the effects of the PTEN tumor suppressor via a direct protein-protein interaction. NEP loss contributes to PC progression. We investigated the therapeutic efficacy of using a lentiviral vector system to restore NEP expression in PC cells. Third-generation lentiviral vectors encoding wild-type NEP (L-NEP) or green fluorescent protein (L-GFP) were introduced into NEP-deficient 22RV1 PC cells. Cells infected with L-NEP or L-GFP at a multiplicity of infection of 10 demonstrated NEP enzyme activity of 1171.2+/-4.9 and 17.2+/-5.3 pmol/microg/min (P<0.0001), respectively. Cell viability, proliferation and invasion were each significantly inhibited in 22RV1 cells expressing NEP compared with control cells infected with L-GFP (P<0.01). Analysis of known downstream effects of NEP showed NEP-expressing cells exhibiting decreased Akt and focal adhesion kinase phosphorylation and increased PTEN protein expression. Finally, injection of L-NEP into established 22RV1 xenograft tumors significantly inhibited tumor growth (P<0.01). These experiments demonstrate that lentiviral NEP gene transfer is a novel targeted strategy for the treatment of NEP-deficient PC.

Akt activation in renal cell carcinoma: contribution of a decreased PTEN expression and the induction of apoptosis by an Akt inhibitor.

BACKGROUND: Akt has been implicated in the oncogenesis of human malignant tumors, because Akt regulates many key effector molecules involved in cell survival. PTEN (phosphatase and tensin homolog deleted on chromosome 10) negatively regulates Akt activation. MATERIALS AND METHODS: The expression of phosphorylated Akt (p-Akt), total Akt and PTEN was analyzed by Western blotting in 45 renal cell carcinoma (RCC) patients. The Bad and phosphorylated Bad (p-Bad) statuses were analyzed in 20 RCC patients. A phosphatidylinositol ether analog was used as an Akt inhibitor to treat four RCC cell lines, namely Caki-1, KU19-20, SW839 and Caki-2. RESULTS: The PTEN expression in RCC was observed to decrease and p-Akt expression to increase significantly in comparison with that in the corresponding normal kidney tissue. The PTEN expression inversely correlated with the p-Akt expression. These alterations were specific for clear cell type RCC, but not for papillary or chromophobe type RCC. Alterations in Bad phosphorylation were also specifically observed in clear cell type. The Akt inhibitor induced apoptosis in KU19-20 and Caki-2 cells with a high Akt activity. CONCLUSIONS: A decreased expression of PTEN may be an underlying mechanism for Akt activation. An Akt inhibitor may be a therapeutic option for a subset of RCC with an elevated Akt activity.

A new technique of laparoscopic surgery for rectal disease.

Laparoscopic rectal surgery is a technique that has to be done in a narrow space: the pelvis. If an immoderate operation is performed with a difficult view, for example in female where the field is disturbed by uterus, it is possible to produce organ trauma or an unexpected bleeding. Taking these problems in mind, we performed laparoscopic surgery in 44 cases of rectal disease with several techniques which we have invented. In these cases either the uterus or the rectum was retracted in the narrow space, and if an anterior resection was to be done, the tape was tied tightly around the rectum below the tumor to avoid touching the tumor and leaving adequate vascular irrigation to the remnant rectum. With the use of our techniques, we did not have female intestinal injury or unexpected bleeding. In addition laparoscopic anterior resection of rectum did not cause any intestinal injury, or unexpected bleeding or anastomotic leakage; also we did not have any local tumor recurrence. It is our belief that these techniques can decrease complications that traumatize the grasping intestine with intestinal forceps and prevent implantation in the anastomosis. This technical report validate that our technique modifications for rectal laparoscopic surgery are useful when a surgeon has to work in a narrow space.

[A case of Collet-Sicard syndrome caused by skull base metastasis of prostate carcinoma].

A case of Collet-Sicard Syndrome caused by skull base metastasis of prostate carcinoma is reported. A fifty-five years old man presenting multiple lymph node and bone metastases of prostate carcinoma was treated with LH-RH agonist and Flutamide, which induced transient decrease in serum PSA levels and size of lymph node metastases. After 8 months of the treatment, the patient started complaining headache, dysphagia and dysarthria. Brain CT and MRI demonstrated a soft tissue mass replacing left pyramidal bone and occipital bone around left jugular foramen. The tumor was diagnosed as skull base metastasis of the prostate carcinoma and was treated with 50Gy of radiation. The symptom improved after the radiation but died of the disease in 4 months. The autopsy revealed the skull base metastasis of the prostate carcinoma and the tumor was proved to be poorly differentiated adenocarcinoma, which was positively stained by anti-PSA antibody. The case showed cranial nerve palsy of IX to XII, which is usually called Collet-Sicard syndrome. This is the third case report of Collet-Sicard syndrome caused by the skull base metastasis of prostate carcinoma, and it is the first case in Japan.

Fat absorption after pylorus-preserving pancreatoduodenectomy reconstructed with Billroth II pancreaticojejunostomy or Billroth I pancreaticogastrostomy.

BACKGROUND/AIMS: The aim of this study was to determine whether Billroth I pancreaticogastrostomy (PG-I) or Billroth II pancreaticojejunostomy (PJ-II) after pylorus-preserving pancreatoduodenectomy is associated with better postoperative fat absorption, based on residual pancreatic exocrine function. Several reconstructive operations have been employed after pylorus-preserving pancreatoduodenectomy to maximize postoperative nutrition. However, no single-institution study has been published comparing the reconstructive procedures with respect to digestion and absorption of fat. METHODOLOGY: Fat absorption was studied using the 13C-trioctanoin breath test in patients who were grouped according to the degree of fibrosis of the pancreatic remnant, which was determined by histologic examination of the resection specimen. The fibrosis was graded: grade 0, < 10% fibrosis; grade 1, 10-30% fibrosis; and grade 2, > 30% fibrosis. There were 22 patients in the PG-I group and 22 patients in the PJ-II group. RESULTS: There were no significant differences between the PG-I and PJ-II groups in the cumulative excretion of labeled carbon dioxide in the patients with grade 0 pancreatic fibrosis. The cumulative excretion in the PG-I group was better than in the PJ-II group in the patients with grade 1 and grade 2 pancreatic fibrosis. CONCLUSIONS: Fat absorption after PG-I is superior to that after PJ-II in patients with disordered exocrine function of the pancreatic remnant. Billroth I pancreaticogastrostomy allows more effective utilization of the exocrine enzymes of the pancreatic remnant due to elimination of the blind loop characteristic of the Billroth II pancreaticojejunostomy.

Portal vein resection without reconstruction during Appleby operation in a patient with pancreatic body carcinoma with cavernous transformation.

The prognosis of pancreatic body carcinoma has been poor due to cancerous invasion of major vessels. Resection of the involved vessels may improve resectability and prognosis. We report a patient who had a pancreatic body carcinoma with cavernous transformation of the portal vein, in whom the portal vein was resected without reconstruction during an Appleby operation. A 67 year-old man was admitted for evaluation of back pain. Enhanced computed tomography showed no main trunk of the portal vein but a developed collateral circulation. Celiac angiography revealed encasement of the common hepatic, splenic and celiac artery. Venous angiography revealed obstruction of the portal and splenic veins with cavernous transformation surrounding these veins. Pre-operative diagnosis was carcinoma in the pancreatic body, which invaded the portal vein, the celiac and common hepatic arteries. The Appleby operation combined with resection of the portal vein without reconstruction could be performed, by preserving collateral vessels and monitoring hepatic venous oxygen saturation (ShvO2) to prevent hepatic ischemia caused by occlusion of the portal vein. The post-operative course was uneventful.

[Dynamics of prostate-specific antigen as prognostic factors in endocrine treatment for prostate cancer].

Prostate-specific antigen values provide important and unique information about prostate cancer to clinicians. However, there is conflicting information about the prognostic significance of the dynamics of PSA decline and elevation after treatment. To provide further insight into the dynamics of PSA as prognostic factors, we herein studied various PSA values as possible prognostic factors in 93 patients with prostate cancer treated with endocrine therapy. Thirteen (14.0%) had stage B tumors, 20 (21.5%) had stage C tumors, and 60 (64.5%) had stage D tumors. The overall 5-year survival rate was 41.2%. Relapse was observed in 32 (34.4%) patients. The influence of pre- and post-treatment PSA (both absolute and percentage values) on survival was analyzed. Normalization of PSA value was associated with prolonged survival regardless of interval to reach the lowest PSA level. The absolute value of PSA at 3 or 6 months following treatment was a significant discriminator, while the pre-treatment PSA level and percentage values of post-treatment PSA were not. These data show that the PSA nadir and PSA value at 3 or 6 months following treatment provide important prognostic information.

[Giant vesico-vaginal stone: a case report].

A 74-year-old female with the chief complaint of lower abdominal and anal pain had been suffering from total incontinence due to cerebral palsy since her childhood. A giant stone was palpable on vaginal examination. A radiograph showed a giant calcification in the pelvis. Magnetic resonance imaging (MRI) revealed a giant vesico-vaginal stone, which occupied most of the bladder and vagina. Cystolithotomy was performed. The removed stone weighed 435 g, and measured 9.0 x 6.5 x 5.5 cm, and was composed of magnesium ammonium phosphate. To our knowledge only eight cases of female giant vesical stone have been reported. We herein report a rare case of vesico-vaginal stone unrelated to gynecological procedures.

[A case of septic shock following incarceration of the penis].

An 84-year-old male presented to the emergency room with the chief complaint of painful, swollen penis following the use of a constriction ring to maintain penile erection. A high fever, chills and hypotension were recognized. Septic shock was presumed, and administration of antibiotics was started. Microbiologic cultures revealed Escherichia coli in blood. We herein report a rare but serious complication accompanying incarceration of the penis.

[Peripelvic extravasation due to peritoneal dissemination of pancreatic carcinoma: a case report].

We herein report a rare case of peripelvic extravasation due to peritoneal dissemination of pancreatic carcinoma. A 75-year-old female with left flank pain was admitted. Computed tomographic scan demonstrated a huge urinoma medial and posterior to the left kidney, and an irregular mass around the left lower ureter. Retrograde pyelography showed complete ureteral obstruction. The probable diagnosis was spontaneous peripelvic extravasation due to left ureteral tumor or ovarian tumor. Laparotomy revealed a tumor involving the body and tail of the pancreas and some disseminated tumors in the retroperitoneum. Pathological diagnosis was metastatic carcinoma of the pancreas. Malignant tumors in the digestive organs should be taken into consideration in the differential diagnosis of peripelvic extravasation.

Intraductal papillary adenocarcinoma with mucin hypersecretion and coexistent invasive ductal carcinoma of the pancreas with apparent topographic separation.

We report a 66-year-old man who had a cystic intraductal papillary adenocarcinoma containing a papillary adenoma, in the head of the pancreas and a coexistent invasive, well differentiated solid tubular adenocarcinoma in the tail of the pancreas. He was hospitalized with acute epigastralgia. Computed tomography demonstrated a multilocular cystic mass in the head of the pancreas and a solid tumor in the tail. Endoscopic retrograde pancreatography showed mucin secretion from an enlarged papilla of Vater, marked dilatation of the main pancreatic duct in the head and body, cystic dilatation of the uncinate branch, and irregular narrowing of the main pancreatic duct in the tail. Total pancreatectomy was performed. Between the cystic tumor and the solid tumor there was a distance of 4.8 cm of normal pancreatic parenchyma and duct, recognized both grossly and microscopically. The patient died 35 months after the operation. At autopsy, peritonitis carcinomatosa was found in the abdominal cavity. Microscopically, disseminated nodules were also well differentiated tubular adenocarcinoma. The apparent anatomic separation of these two tumors within the pancreas is extremely unusual.

Estimation of fat absorption with the 13C-trioctanoin breath test after pancreatoduodenectomy or pancreatic head resection.

The aim of this study was to determine if fat absorption is better after pylorus-preserving pancreatoduodenectomy (PPPD) or duodenum-preserving pancreatic head resection (DPPHR) compared with that following pancreatoduodenectomy with gastrectomy (SPD), provided the patients have similar pancreatic exocrine function. Fat absorption was studied using the 13C-trioctanoin breath test in patients who were grouped according to the degree of fibrosis of the pancreatic remnant. The latter was judged by histologically measuring the fibrosis in a transectional wedge of resected pancreas. We evaluated 11 SPD cases, 25 PPPD cases, and 9 DPPHR cases. The 13C excretion rates and cumulative excretion values following DPPHR or PPPD were significantly better than those following SPD. The 13C excretion rates and cumulative values for the patients with > 30% fibrosis of the pancreas were lower than those in patients with < 30% pancreatic fibrosis, regardless of the surgical procedure. The cumulative value in the SPD group, however, was lower than that in the PPPD or DPPHR patients with < 30% pancreatic fibrosis. The results suggested that fat absorption following PPPD or DPPHR is superior to that after SPD in patients with the same fibrotic area of the pancreatic remnant and depends on the degree of fibrosis in the pancreatic remnant.