Novel invagination procedure for pancreaticojejunostomy using double purse string sutures: A technical note.

BACKGROUND: Postoperative pancreatic fistula (POPF) is one of the most serious complications after pancreaticoduodenectomy (PD), and the choice of pancreaticojejunostomy (PJ) is considered a key factor affecting the occurrence of POPF. Numerous anastomotic methods and their modifications have been proposed, and there is no method that can completely avoid the occurrence of POPF. Based on our team's experience in pancreatic surgery and a review of relevant literature, we describe a novel invagination procedure for PJ using double purse string sutures, which has resulted in favourable outcomes. AIM: To describe the precise procedural steps, technical details and clinical efficacy of the novel invagination procedure for PJ. METHODS: This study adopted a single-arm retrospective cohort study methodology, involving a total of 65 consecutive patients who underwent PD with the novel invagination procedure for PJ, including the placement of a pancreatic stent, closure of the residual pancreatic end, and two layers of purse-string suturing. Baseline data included age, sex, body mass index (BMI), pancreatic texture, pancreatic duct diameter, operation time, and blood loss. Clinical outcomes included the operation time, blood loss, and incidence of POPF, postoperative haemorrhage, delayed gastric emptying, postoperative pulmonary infection, postoperative abdominal infection, and postoperative pulmonary infection. RESULTS: The mean age of the patients was 59.12 (± 8.08) years. Forty males and 25 females were included, and the mean BMI was 21.61 kg/m2 (± 2.74). A total of 41.53% of patients had a pancreatic duct diameter of 3 mm or less. The mean operation time was 263.83 min (± 59.46), and the mean blood loss volume was 318.4 mL (± 163.50). Following the surgical intervention, only three patients showed grade B POPF (4.62%), while no patients showed grade C POPF. Five patients (5/65, 7.69%) were diagnosed with postoperative haemorrhage. Six patients (6/65, 9.23%) experienced delayed gastric emptying. Four patients (4/65, 6.15%) developed postoperative pulmonary infection, while an equivalent number (4/65, 6.15%) exhibited postoperative abdominal infection. Additionally, two patients (2/65, 3.08%) experienced postoperative pulmonary infection. CONCLUSION: The novel invagination technique for PJ is straightforward, yields significant outcomes, and has proven to be safe and feasible for clinical application.

Liver dual arterial blood supply maintains liver regeneration: Analysis of signaling pathways in rats.

Liver dual arterial blood supply (LDABS) could increase blood supply to the liver and maintain normal liver regeneration in patients with compromised portal vein. The current study attempted to examine the underlying molecular mechanisms. Male Sprague­Dawley rats randomly received partial hepatectomy (PH) alone or PH followed by LDABS. Liver regeneration was assessed by histological examination, liver function and liver regeneration rate (LRR). Whole­genome oligo microarray analysis was used to compare gene expression profile between rats receiving PH and rats receiving PH plus LDABS. Key genes identification was validated using a MAPK signaling polymerase chain reaction (PCR) array. The extent of liver regeneration in rats receiving PH plus LDABS was comparable to that in rats receiving PH alone. The differentially expressed genes were enriched in 12 signaling pathways in two groups. MAPK signaling pathway, NF­kappa B signaling pathway, and Toll­like receptor signaling pathway were involved in LDABS­mediated liver regeneration, with Retinoblastoma 1 (Rb1), Cyclin D1, Cyclin­dependent kinase 4, Mitogen­activated protein kinase 10 (Mapk10) and CAMP responsive element binding protein 1 genes in the initiation phase, Kirsten rat sarcoma viral oncogene homolog (Kras), tumor protein 53, MYC proto­oncogene, BHLH transcription factor, Cyclin E1 and Heat shock protein family B (small) member 1 genes in the proliferation phase, Kras, Rb1, Jun proto­oncogene, AP­1 transcription factor subunit, Cyclin D2 and Mapk10 genes in the termination phase were identified as key genes in LDABS­mediated liver regeneration using MAPK signaling PCR array analysis.

RRS1 silencing suppresses colorectal cancer cell proliferation and tumorigenesis by inhibiting G2/M progression and angiogenesis.

Colorectal cancer (CRC) is one of the most common malignancies worldwide. Ribosome biogenesis regulatory protein homolog (RRS1) is an essential factor involved in ribosome biogenesis, while its role in CRC remains largely unclear. Here, we found that RRS1 expression was significantly higher in CRC tissues compared with adjacent normal tissues. RRS1 High expression also predicted poor overall survival of CRC patients. Knockdown of RRS1 induced the G2/M cell cycle arrest, apoptosis and suppressed the proliferation of RKO and HCT-116 CRC cells. Furthermore, angiogenesis was also reduced in CRC cells after RRS1 knockdown. In addition, suppression of RRS1 blunted the tumor formation of CRC cells in nude mice. At the molecular level, silencing of RRS1 decreased the expression of M-phase inducer phosphatase 3 (CDC25C), Cyclin-dependent kinase 1 (CDK1), antigen KI-67 (KI67) and increased the protein level of cyclin-dependent kinase inhibitor 1 (CDKN1A) and tumor suppressor p53 (p53). Taken together, our findings provide evidence that RRS1 may promote the development of colon cancer. Therefore, targeting RRS1 may be a promising therapeutic strategy for CRC patients.

The Wnt/ß-catenin signaling pathway mechanism for pancreatic cancer chemoresistance in a three-dimensional cancer microenvironment.

ß-catenin is a key protein that is encoded by the CTNNB1 gene in the Wnt signaling pathway. This study investigated the associations between ß-catenin expression and implications for the efficacy of gemcitabine on pancreatic cancer cells in a three-dimensional (3-D) cancer microenvironment. For low ß-catenin expression pancreatic carcinoma cells, the inhibition rates (IRs) for low, middle, and high doses of gemcitabine were 0.615 ± 0.079, 0.691 ± 0.093, and 0.765 ± 0.061, respectively. For the high ß-catenin expression pancreatic carcinoma cells, the IRs for the same doses were 0.325 ± 0.072, 0.453 ± 0.075, and 0.537 ± 0.056, respectively. Additionally, the evaluation of ß-catenin immunoreactivity in 31 pancreatic cancer patients revealed that the low ß-catenin protein expression group had significantly longer overall survival (OS) and disease free survival (DFS) than the high ß-catenin protein group (P < 0.05). Overall, ß-catenin protein expression levels were significantly correlated to gemcitabine sensitivity in seven pancreatic carcinoma cell lines in the 3-D cancer microenvironment. These data suggest that large-scale clinical studies are warranted to assess the role of the Wnt/ß-catenin signaling pathway on ß-catenin protein expression and chemosensitivity to gemcitabine in pancreatic cancer.

Studied microRNA gene expression in human hepatocellular carcinoma by microRNA microarray techniques.

AIM: To achieve a better understanding of the molecular mechanisms of microRNA expression changes involved in hepatocellular carcinoma. METHODS: In this research process, patients were not treated with antivirals, immunosuppressants or immunomodulators for at least 6 mo before collecting serum. The study population was composed of 35 outpatient hepatitis B virus (HBV) cases and 12 healthy control cases from the Affiliated Hospital of Inner Mongolia Medical University (Inner Mongolia, China) from July 2013 to April 2014. The 35 HBV cases were divided into two groups: a hepatocirrhosis group with 20 cases and a liver cancer group with 15 cases. All 35 cases carried HBsAg. The diagnostic criteria followed the European Association for the Study of the Liver 2012 (EASL2012) standards. MicroRNA (miRNA) was extracted from a control group of patients, a group with hepatocirrhosis and a group with liver cancer and its quality was analyzed using the human V2 microRNA expression beadchip. Cluster analysis and a radar chart were then applied to the miRNA changes. RESULTS: The miRNA-qualified rate of human serum samples was 93%. The concentration of a single sample was > 200 ng/µL and the volume was > 5 µL. All miRNA serum samples were uncontaminated by the genome. The Mann-Whitney test showed significant differences in miRNA between each group, with a detection P-value of < 0.05. Illumina software was set up with Diff Score set to ± 13, meaning that P = 0.001.There were significant changes in miRNA expression between the three groups. miRNA-183 was the most up-regulated, followed by miRNA-373. miRNA-129 and miRNA-188 were both strongly down-regulated and miRNA-378 was down-regulated a small amount. The liver cancer group had greater changes, which indicated that changes in miRNA expression levels were caused by hepatocirrhosis. The liver cancer disease course then further increased these changes. In the pentagon created by these five miRNAs, three groups showed significant deviation. The liver cancer group had a bigger deviation trend. The chart indicated that miRNA expression changes occurred in the hepatocirrhosis group, which increased in the liver cancer disease course and were irreversible. CONCLUSION: There was a significant relationship between the irreversible up-regulation of miRNA-183/373 and down-regulation of miRNA-129/188/378 and incidences of hepatocirrhosis and liver cancer.

Osteopontin knockdown suppresses the growth and angiogenesis of colon cancer cells.

AIM: To investigate the effects of osteopontin (OPN) gene expression knockdown on colon cancer Lovo cells in vitro. METHODS: Four candidate small interfering RNA (siRNA) constructs targeting the OPN gene and a scrambled control sequence (NC-siRNA) were synthesized and inserted into a pGPU6/GFP/Neo expression vector. After confirmation by restriction enzyme digestion and DNA sequencing, the recombinant plasmids were subsequently transfected into a human colon cancer cell line (Lovo) using a liposome transfection method. Stably transfected cells were maintained with G418 selection and referred to as Lovo-OPN-1, -2, -3, -4, and Lovo-NC cells. Knockdown efficiency of each of the four siRNA constructs was determined by real-time reverse transcription polymerase chain reaction assays and western blotting, and the construct with the most effective silencing was used for subsequent experiments. Cell proliferation, adhesion, and Matrigel invasion assays were performed to analyze the effects of OPN knockdown in stably transfected Lovo cells. The levels of four angiogenic factors, namely vascular endothelial growth factor (VEGF), matrix metalloproteinase (MMP)-2, MMP-9 and urokinase plasminogen activator were detected by enzyme-linked immunosorbent assays (ELISA). RESULTS: Recombinant vectors containing OPN-specific and scrambled siRNA sequences were successfully constructed and stably transfected into Lovo cells. Compared with the control Lovo and Lovo-NC cells, the levels of OPN mRNA and protein expression in Lovo-OPN-1, -2, -3, and -4 were significantly reduced (all P < 0.05), with the most efficient reduction observed in Lovo-OPN-4 cells (P < 0.05). Relative to untransfected Lovo cells, OPN mRNA expression levels in Lovo-NC and Lovo-OPN-4 cells were 1.008 ± 0.067 and 0.160 ± 0.023, respectively. The relative OPN protein expression levels in Lovo, Lovo-NC, and Lovo-OPN-4 cells were 3.024 ± 0.211, 2.974 ± 0.630, and 0.121 ± 0.008, respectively. Moreover, transfection with the scrambled sequence had no effect on the expression of OPN. After 24, 48, 72, and 96 h of cultivation, absorption values at 450 nm to assess proliferation of Lovo-OPN-4 cells were 0.210 ± 0.017, 0.247 ± 0.024, 0.314 ± 0.037, and 0.359 ± 0.043, respectively, which were significantly lower than those of Lovo (0.244 ± 0.031, 0.313 ± 0.024, 0.513 ± 0.048 and 0.783 ± 0.051) and Lovo-NC cells (0.241 ± 0.029, 0.309 ± 0.022, 0.563 ± 0.023, and 0.735 ± 0.067) (all P < 0.05). The absorption values at 595 nm, which were measured in a cell adhesion assay, showed that adhesion of Lovo-OPN-4 cells (0.215 ± 0.036) was significantly decreased compared to Lovo (0.490 ± 0.037) and Lovo-NC cells (0.462 ± 0.043) (P < 0.05). The number of invasive Lovo-OPN-4 cells (16.1 ± 1.9) was also significantly decreased compared to Lovo (49.9 ± 5.4) and Lovo-NC cells (48.8 ± 4.5) (P < 0.05). ELISA assays showed significant reductions in Lovo-OPN-4 cells compared to Lovo and Lovo-NC cells with regard to the expression of VEGF (1687.85 ± 167.84 ng/L vs 2348.54 ± 143.80 ng/L and 2284.39 ± 138.62 ng/L, respectively), MMP-2 (2966.07 ± 177.36 µg/L vs 4084.74 ± 349.54 µg/L and 4011.41 ± 424.48 µg/L, respectively), MMP-9 (3782.89 ± 300.64 µg/L vs 5062.90 ± 303.02 µg/L and 4986.38 ± 300.75 µg/L, respectively) and uPA (1152.69 ± 120.79 µg/L vs 1380.90 ± 147.25 µg/L and 1449.80 ± 189.92 µg/L, respectively) (all P < 0.05). CONCLUSION: Knockdown of OPN gene expression suppresses colon cancer cell growth, adherence, invasion, and expression of angiogenic factors.

Probing the hepatic progenitor cell in human hepatocellular carcinoma.

Objective. The intrahepatic stem cells, also known as hepatic progenitor cells (HPCs), are able to differentiate into hepatocytes and bile duct epithelia. By exposure of different injuries and different hepatocarcinogenic regimens, the mature hepatocytes can no longer effectively regenerate; stem cells are involved in the pathogenesis of hepatocellular carcinoma. Methods. Immunohistochemistry was performed on 107 paraffin-embedded hepatocellular carcinoma specimens with the marker of hepatocyte and hepatocellular carcinoma (HepPar1), biliary differentiation (CK7,CK19), haemopoietic stem cell (HSC) (c-kit/CD117, CD34, and Thy-1/CD90), HPC specific markers (OV-6), and Ki-67, p53 protein. Results. HPCs can be identified in the tumor nodules, around the edge of tumor nodules, and in the portal tracts of the paracirrhosis nodules being positive in HepPar1, CK7, CK19, and OV-6, but they failed to immunostain with CD117, CD34, and CD90. The HPCs positive in Ki-67 are observed in the tumor and paracirrhosis tissues. In 107 specimens, 40.2% (43/107) HCC tissues expressed p53 protein, lower than that of the HPCs around the tumor nodules (46.7%, 50/107) and much higher than that of the HPCs around the paracirrhosis nodules (8.41%, 9/107). Conclusion. Human hepatocellular carcinogenesis may be based on transformation of HPCs, not HSCs, through the formation of the transitional cells (hepatocyte-like cells and bile ductal cells).

Development of a new auxiliary heterotopic partial liver transplantation technique using a liver cirrhosis model in minipigs: Preliminary report of eight transplants.

This study aimed to develop a new auxiliary heterotopic partial liver transplantation (AHPLT) technique in minipigs using a model of liver cirrhosis. Based on our previous study, 14 minipigs were induced to cirrhosis by administration of carbon tetrachloride (CCl(4)) through intraperitoneal injection. All of the cirrhotic animals were utilized as recipients. The donor's liver was placed on the recipient's splenic bed, and the anastomosis was performed as follows: end-to-end anastomosis between the donor's portal vein and the recipient's splenic vein, end-to-side anastomosis between the donor's suprahepatic vena cava and the recipient's suprahepatic vena cava, and end-to-end anastomosis between the donor's hepatic artery and the recipient's splenic artery. The common bile duct of the donor was intubated and bile was collected with an extracorporeal bag. Vital signs, portal vein pressure (PVP), hepatic venous pressure (HVP) and portal vein pressure gradient (PVPG) were monitored throughout the transplantation. All 8 minipigs that developed liver cirrhosis were utilized to establish the new AHPLT; 7 cases survived. Following the surgical intervention, the PVP and PVPG of the recipients were lower than those prior to the operation (P<0.05), whereas the PVP and PVPG of the donors increased significantly compared to those of the normal animals (P<0.05). A new operative technique for AHPLT has been successfully described herein using a model of liver cirrhosis.

Right hepatic arterial pseudoaneurysm with hemobilia following minilaparotomy cholecystectomy: a rare complication.

Hepatic arterial pseudoaneurysm with hemobilia occurs less frequently as a complication of minilaparotomy cholecystectomy than laparoscopic cholecystectomy; however, given its severe nature, it needs to be managed promptly. This report presents a case of right hepatic artery pseudoaneurysm with hemobilia in a 36-year-old woman who underwent minilaparotomy cholecystectomy 5 weeks earlier. Angiography with embolization was carried out as definitive treatment.

A relative quantitative method to detect OCT4A gene expression by exon-junction primer and locked nucleic acid-modified probe.

OBJECTIVE: OCT4A has been known to play a critical role in the maintenance of pluripotency of embryonic stem cells. Recent research has shown that OCT4A is also expressed in partial tumor cell lines and tissues. This study is aimed to develop a real-time reverse transcriptase polymerase chain reaction (RT-PCR) assay for relative quantitative detection of OCT4A mRNA and discrimination from OCT4B, pseudogene, and genomic contaminations. METHODS: A locked nucleic acid (LNA)-modified probe was designed to discern the single base difference 352A/C to identify OCT4A mRNA. An exon-junction primer was designed to avoid false positive caused by genomic contaminations. In addition, a house keeping gene glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was measured in parallel to normalize the differences between samples and operations. RESULTS: Experiments showed that the newly established RT-PCR assay amplified the OCT4A mRNA selectively; OCT4A analogues gave negative signals. Cell lines nTERA-2 and HepG2 showed positive results in OCT4A expression, while for HeLa and 293 cell lines, as well as primary peripheral blood mononuclear cells (PBMCs), OCT4A expression was negative. Additionally, the relative quantity of OCT4A mRNA was calculated by cycle threshold (C(t)) method and house keeping gene normalization. CONCLUSIONS: This technique proved to be effective for relative quantitation of OCT4A mRNA with high specificity.

Hepatocellular carcinoma-specific immunotherapy with synthesized α1,3- galactosyl epitope-pulsed dendritic cells and cytokine-induced killer cells.

AIM: To evaluate the safety and clinical efficacy of a new immunotherapy using both α-Gal epitope-pulsed dendritic cells (DCs) and cytokine-induced killer cells. METHODS: Freshly collected hepatocellular carcinoma (HCC) tumor tissues were incubated with a mixture of neuraminidase and recombinant α1,3-galactosyltransferase (α1,3GT) to synthesize α-Gal epitopes on carbohydrate chains of the glycoproteins of tumor membranes. The subsequent incubation of the processed membranes in the presence of human natural anti-Gal IgG resulted in the effective phagocytosis to the tumor membrane by DCs. Eighteen patients aged 38-78 years with stage III primary HCC were randomLy chosen for the study; 9 patients served as controls, and 9 patients were enrolled in the study group. RESULTS: The evaluation demonstrated that the procedure was safe; no serious side effects or autoimmune diseases were observed. The therapy significantly prolonged the survival of treated patients as compared with the controls (17.1 ± 2.01 mo vs 10.1 ± 4.5 mo, P = 0.00121). After treatment, all patients in the study group had positive delayed hypersensitivity and robust systemic cytotoxicity in response to tumor lysate as measured by interferon-γ-expression in peripheral blood mononuclear cells using enzyme-linked immunosorbent spot assay. They also displayed increased numbers of CD8-, CD45RO- and CD56-positive cells in the peripheral blood and decreased α-fetoprotein level in the serum. CONCLUSION: This new tumor-specific immunotherapy is safe, effective and has a great potential for the treatment of tumors.

Protective effect of pyrroloquinoline quinone against Abeta-induced neurotoxicity in human neuroblastoma SH-SY5Y cells.

The neurotoxicity of aggregated beta-amyloid (Abeta) has been implicated as a critical cause in the pathogenesis of Alzheimer's disease (AD). It can cause neurotoxicity in AD by evoking a cascade of oxidative damage-dependent apoptosis to neurons. In the present study, we for the first time investigated the protective effect of pyrroloquinoline quinone (PQQ), an anionic, water soluble compound that acts as a redox cofactor of bacterial dehydrogenases, on Abeta-induced SH-SY5Y cytotoxicity. Abeta(25-35) significantly reduced cell viability, increased the number of apoptotic-like cells, and increased ROS production. All of these phenotypes induced by Abeta(25-35) were markedly reversed by PQQ. PQQ pretreatment recovered cells from Abeta(25-35)-induced cell death, prevented Abeta(25-35)-induced apoptosis, and decreased ROS production. PQQ strikingly decreased Bax/Bcl-2 ratio, and suppressed the cleavage of caspase-3. These results indicated that PQQ could protect SH-SY5Y cells against beta-amyloid induced neurotoxicity.

[Expression of CK20 mRNA in the peripheral blood around postoperative chemotherapy in patients with gastric cancer and its clinical significance].

OBJECTIVE: To investigate whether CK20 mRNA expression level could be considered as an effective molecular indicator for evaluation of chemotherapy sensitivity. METHODS: All samples of peripheral blood were taken from 31 gastric cancer patients undergone radical operation a week before postoperative chemotherapy, at the first day of chemotherapy point, and after the first cycle of chemotherapy respectively, and subjected to FQ RT-PCR assay for CK20 mRNA. The chemotherapy scheme was FOLFOX 4. The control group was 15 healthy volunteers. RESULTS: Aomng the 31 gastric cancer patients, the value of CK20 mRNA before postoperative chemotherapy was increased (2.96+/-2.27 vs 2.22+/-2.12, t=2.10, P<0.05) in 25 positive cases, and then declined after chemotherapy(2.05+/-1.86 vs 2.96+/-2.27, t=2.50, P<0.05) in 24 positive cases. The expression level of CK20 mRNA in patients before chemotherapy was increased in 16 cases(51.6%), declined in 9 cases(29.0%) and stabilized as negative in 6 cases(19.4%). After chemotherapy the level of CK20 mRNA was increased in 7 cases(22.6%), declined in 17 cases (54.8%) and stabilized as negative in 7 cases(22.6%), there was significant difference between the two groups(chi(2)=6.06, P<0.05). CONCLUSIONS: The expression level of CK20 mRNA in the peripheral blood detected by FQ RT-PCR in patients with gastric cancer declines after postoperative adjuvant chemotherapy. Different individuals have different sensitivity to chemotherapeutics. Dynamic monitoring CK20 mRNA should be considered as an effective index to evaluate the efficacy of postoperative adjuvant chemotherapy.

[Wound healing after pancreaticojejunostomy in piglets: a comparison between two anastomotic methods].

OBJECTIVE: To evaluate wound healing after types of pancreaticojejunostomy. METHODS: After resection of the pancreatic head, 38 domestic piglets were divided into two groups according to the types of anastomoses: group I: binding pancreaticojejunostomy, a new technique designed and advocated by professor Peng Shuyou; group II: end-to-end pancreaticojejunal invagination. Anastomotic strength in vivo and histopathological findings were assessed on operative day and postoperative day 5 and 10. RESULTS: Bursting pressure was 139.7 +/- 8.0, 178.7 +/- 9.7 and 268.8 +/- 12.8 mm Hg in group I on day 0, 5 and 10, whereas 67.3 +/- 7.9, 96.2 +/- 10.4 and 130.6 +/- 9.3 mm Hg in group II. The gain on day 0 to 5 and 5 to 10 was 27.9% and 50.5% in group I and 42.9% and 35.7% in group II, respectively. A significant difference was observed between group I and group II, and between 5 and 10 day after anastomoses (P < 0.01). Breaking strength was 4.5 +/- 0.4, 6.6 +/- 0.4 and 10.0 +/- 0.6 N in group I on day 0, 5 and 10 and 4.6 +/- 0.6, 5.8 +/- 0.5 and 7.1 +/- 0.6 N in group II. Although a similar value was shown in both types of anastomoses on day 0, a rapider gain was demonstrated on day 0 to 5 and 5 to 10 in group I (44.8% and 52.9%) than in group II (25.4% and 22.0%). A significant difference was found on day 5 and 10 between the two types of anastomoses (P < 0.05 and P < 0.01). Anastomotic site was well repaired by connective tissue and the cut surface of pancreatic stump was covered by mucosal epithelium in group I on day 10, but the cut surface was incompletely repaired by granulation tissue and no, regeneration of the epithelium was found in group II. CONCLUSION: Anastomotic strength of binding pancreaticojejunostomy was stronger than end-to-end pancreaticojejunal invagination and the healing was better and rapid.