Shear-Induced Structuring for Multiple Parallel Gel Filaments Obtained from Casein-Alginate Hybrids.

The addition of more than 1 wt % sodium alginate to a 7.5 wt % sodium caseinate solution induced aqueous two-phase separation (ATPS). Alginate and caseinate were effectively condensed in the upper and lower phases, respectively, thereby forming an alginate-casein assembly. The rigid structure of the alginate, which was formed by repeated regular hydrogen bonds between guluronic acid units, became supple when casein micelles penetrated and were adsorbed into the coiled alginate chains to weaken the diaxial linkages of the alginate hydrogen bonds. Protein-polysaccharide hybrid gel fibrils with a bundled structure were formed due to the deformation of the alginate-casein assembly in an aqueous solution by shear in a co-flow double tube, followed by cross-linking with Ca2+ supplied as the sheath fluid. The combination of ATPS and shear-induced elongation of the alginate-casein assemblies enabled the fabrication of hundreds of parallel gel filaments (φ = 1-10 µm) along the flow direction. These multiple parallel gel filaments can be applied to biomimetic chemistry for fibrous living tissues, as a biodegradable scaffold for cell engineering, and as a release carrier of physiologically active substances or drugs. Our proposed technique enables the formation of biomimetic protein-polysaccharide hybrid gel filaments with a bundled structure using Ca2+ chelation under laminar flow in a capillary, without the need for enzymatic cross-linking and multihole nozzles.

Switch of SpnR function from activating to inhibiting quorum sensing by its exogenous addition.

The opportunistic human pathogen Serratia marcescens AS-1 produces the N-hexanoylhomoserine lactone (C6HSL) receptor SpnR, a homologue of LuxR from Vibrio fischeri, which activates pig clusters to produce the antibacterial prodigiosin. In this study, we attempted to artificially regulate quorum sensing (QS) by changing the role of SpnR in N-acylhomoserine lactone (AHL)-mediated QS. SpnR was obtained as a fusion protein tagged with maltose-binding protein (MBP) from overexpression in Escherichia coli, and its specific affinity to C6HSL was demonstrated by quartz crystal microbalance analysis and AHL-bioassay with Chromobacterium violaceum CV026. Prodigiosin production was effectively inhibited by externally added MBP-SpnR in both wild-type AS-1 and the AHL synthase-defective mutant AS-1(ΔspnI). For the mutant, the induced amount of prodigiosin was drastically reduced to approximately 4% with the addition of 18 µM MBP-SpnR to the liquid medium, indicating 81% trapping of C6HSL. A system for inhibiting QS can be constructed by adding exogenous AHL receptor to the culture broth to keep the concentration of free AHL low, whereas intracellular SpnR naturally functions as the activator in response to QS.

In vitro analysis of essential binding sites on the promoter of the Serratia marcescens spn operon with the quorum-sensing receptor SpnR.

The N-acylhomoserine lactone (AHL) receptor SpnR is a LuxR family protein that acts as a negative regulator of AHL-dependent quorum sensing (QS). SpnR binds to DNA in Serratia marcescens AS-1 via the spn box; however, the binding affinity of SpnR with the nucleotides on the spn box has not yet been investigated. In this study, we used an spn-box-modified sensor electrode, and quartz crystal microbalance analysis demonstrated a drastic reduction of the uptake of SpnR. The nucleotides G5 and C16 at the AHL-receptor complex-binding site are conserved in Gram-negative bacteria, including the lux box in Vibrio fischeri, the tra box in Agrobacterium tumefaciens, and the spn box in S. marcescens. Indeed, the affinity of SpnR to DNA was reduced to 8% by G5C substitution of the spn box. The affinity of SpnR tagged with maltose-binding protein to the immobilized gene promoter was reduced in the order of C16G and G5C substitutions, which corresponded with previous reports on the lux box. These results suggest that formation of hydrogen bonds at amino acid residues containing guanine at position 5 on a lux-box-like promoter universally contributes to the stability of the receptor complex, whose interaction initiates a sequential QS process in the LuxR family. Biotechnol. Bioeng. 2016;113: 2513-2517. © 2016 Wiley Periodicals, Inc.

[Preoperative Chemoradiotherapy for Stage II or III Esophageal Squamous Cell Carcinoma].

BACKGROUND: The goal of this retrospective study was to investigate the efficacy and safety of preoperative chemoradiotherapy( NACR)in patients with Stage II or Stage III esophageal squamous cell carcinoma(SCC). METHODS: Between 2004 and 2014, a total of 86 patients underwent surgical resection in conjunction with NACR for esophageal SCC at our institute. Thirty-one patients(36.0%)had Stage II disease and 55 patients(64.0%)had Stage III disease. RESULTS: The median age was 64(43-81)years. A total of 78 patients received the full NACR regimen. The most common major Grade 3 hematologic toxic effects of NACR were leukopenia and neutropenia(48 cases), while the most common major Grade 3 non-hematologic toxic effect was anorexia(12 cases). One patient died in the hospital and no patients died within 30 days after surgery. A pathological complete response was achieved in 23 cases. Pathological staging(number of cases)was Stage 0(23), Stage I (8), Stage II (28), Stage III (25), and Stage IV (2). The 5-year overall survival rate(OS)was 51.0%, and was 83.2% in Stage II patients and 29.9% in Stage III patients. CONCLUSION: Preoperative NACR is safe and may improve OS and downstaging rates in patients with esophageal SCC.

Radiochemotherapy for esophageal squamous cell carcinoma in elderly patients.

BACKGROUND/AIMS: The goal of this retrospective study was to investigate the efficacy and safety of curative radiochemotherapy (CRT) in elderly patients with esophageal squamous cell cancer (SCC). METHODOLOGY: Between 1986 and 2011, 38 consecutive patients aged over 75 years with esophageal SCC were initially treated with a CRT regimen comprising radiation therapy at a dose of 60 Gy and two cycles of chemotherapy with 5-fluorouracil/cisplatin. RESULTS: Fourteen patients were aged over 80 and 24 under 79 years. Complete treatment compliance was obtained in 30 (78.9%) of the patients. Chemotherapy was terminated and the dose of radiation reduced in 7 and 1 patients, respectively. The overall incidence of CRT-related complications was 86.8% (33 patients). Grade 3 leukocytopenia was observed in 6 (15.8%) patients, all of whom recovered promptly. A complete response was observed in 17 patients (44.7%). The overall 3-year survival rate was 32.0%. CONCLUSIONS: Curative CRT is safe and may improve overall and progression-free 3-year survival rates in elderly patients aged more than 75 years with esophageal SCC.

Inhibition of quorum sensing in opportunistic pathogen, Serratia marcescens, using cyclodextrin-immobilized, multiple parallel gel filaments fabricated with dynamic flow of polymer blend solution.

Prodigiosin production in the model opportunistic human pathogen, Serratia marcescens AS-1, depends on quorum sensing (QS), the activation of which is induced by an increase in N-hexanoyl homoserine lactone (C6HSL) concentration. When mixed with C6HSL in the broth culture of S. marcescens immobilized cyclodextrin (CD) effectively formed an inclusion complex resulting in suppression of the QS-mediated prodigiosin production. Thousands of parallel gel filament bundles could be fabricated by the dynamic flow of aqueous sodium alginate blended with α-CD or hydroxypropyl-ß-CD (HP-ß-CD) immobilized hydroxypropyl cellulose (HPC) inside a co-axial microfluidic device. Shear-induced fibrous domains of HPC-alginate complex assembly were immediately jacketed by calcium alginate gel formed by a sheath flow containing Ca2+ ions. Immobilized α-CD and HP-ß-CD on the HPC/alginate gel fibers could inhibit the C6HSL-mediated prodigiosin production to approximately 10%. Furthermore, the swarming motility of S. marcescens AS-1 was effectively suppressed in the presence of free or immobilized α-CD or HP-ß-CD. The CD-immobilized gel filament bundle is a novel type of preventive material against bacterial infections.

Dynamic gelation of shear-induced filamentous domains for cellulose ether assemblies due to polyion complexation.

Assemblies of carbohydrate polymers are important in a number of applications and improved methods for their fabrication are increasingly sought after. Herein, we report that an aqueous two-phase system of alginate (Alg) and hydroxypropyl cellulose with poly(methacrylic acid) graft chains (HPC-PMA) facilitated the assembly of Alg/HPC-PMA in both phases. Dynamically formed filamentous domains in a flow field were gelled by rapid complexation with cationic polyethyleneimine (PEI). The fabricated HPC-PMA gel filament morphologies can be switched between the bundled and dissociated gel filaments using a co-flow microfluidic device in response to the amount of supplied PEI crosslinker. Excess complexation of PEI contributes to the fabrication of cationic gel filaments; this contribution results in a dissociated structure due to electrostatic repulsion. In contrast, an appropriate amount of PEI resulted in a bundle structure. The proposed spinning method avoids the risk of nozzle clogging, and enables the one-step spinning of multiple gel filaments.

Structuring of multiple parallel pectin gel filaments by applied shear.

The bundled structure of micron-sized pectin gel filaments was formed by quick shear-induced gelation of the filamentous domains of pectin-polyethylene glycol (PEG) assemblies. Highly concentrated pectin with PEG in a separated pectin-rich phase under aqueous two-phase separation in the pectin/PEG/NaCl system enabled the formation of the pectin-PEG assembly, which was elongated in the flow direction, resulting in the generation of filamentous domains using a microfluidic device. The pectin gel filaments were formed by crosslinking with Ca2+ in the presence of shear-responsive PEG assemblies formed in the PEG-rich phase, because the filamentous PEG assemblies prevented fusion of the pectin filaments to form the seamless cylindrical gel. The shear-dependent elongation applied to the pectin-PEG assembly under the aqueous two-phase separation condition enabled the formation of the biomimetic bundled filamentous structure using bio-safe PEG as a sacrificial polymer, without the requirement of a multi-hole nozzle. Potential applications for gel filaments possessing a bundled structure are matrices in the biomedical field, such as a biodegradable scaffold for cell engineering.

One-step reconstruction of IVC and right hepatic vein using reversed auto IVC and left renal vein graft.

INTRODUCTION: A malignant tumor invading the inferior vena cava (IVC) and concomitant with 3 hepatic venous involvement still remains surgical therapeutic challenge. PRESENTATION OF CASE: A 79-year-old male presented with liver dysfunction and was eventually diagnosed with an intrahepatic huge IHCC originating at the level of confluence of 3 hepatic veins to the IVC, extending to the right hepatic vein (HV). Under extracorporeal circulation, the liver with the IVC, portal vein, hepatic artery and common bile duct were removed en bloc. In the back table, the left liver lobe was resected including the tumor and IVC, and the right HV was reconstructed using the patient's reversed left renal vein (LRV) and IVC graft. Subsequently, the patient's right liver lobe with the reversed LRV/IVC graft was transplanted back into the patient using a partial liver transplant technique. Liver enzymes gradually came down to the value within normal range on postoperative day 13 with the patency of the right HV. However, the patient expired on POD 16 because of sudden septic complications. DISCUSSION: This technique could be one of the options to reconstruct the HV/IVC when a tumor invades all 3 HVs. CONCLUSION: To the best of our knowledge, this is the first report of such HV reconstruction in combination with extracorporeal resection of huge liver cancer in updated world literature.

Phase II Study of Neoadjuvant Chemotherapy With S-1 and CDDP in Patients With Lymph Node Metastatic Stage II or III Gastric Cancer.

OBJECTIVES: We conducted a phase II trial to investigate the efficacy and safety of neoadjuvant chemotherapy (NAC) comprising S-1 and cisplatin (CDDP) followed by extensive resection in the management of resectable locally advanced gastric cancer with lymph node (LN) metastases. METHODS: Patients with LN metastases from stage II or III gastric cancer received S-1 and CDDP, as NAC criteria for LN metastases were the involvement of ≥4 nodes <2 cm or ≥1 nodes ≥2 cm as confirmed by a total body computed tomography scan. All patients underwent extensive resection including D2 gastrectomy. The primary endpoint was complete resection rate and the secondary endpoints were 3-year relapse-free and overall survival. RESULTS: Fifty patients were assessable for the analysis. The complete resection rate was 87.8%. Three-year relapse-free survival was 44.9% and 3-year overall survival rate was 48.0%. CONCLUSIONS: NAC with S-1 and CDDP is safe and may improve the complete resection rate in patients with metastatic LN gastric cancer. This suggests that LN metastases would provide good target lesions in future clinical trials of NAC.

Effect of preoperative colonic drainage for obstructing colorectal cancer.

Obstructing colorectal cancer (OCRC) is believed to indicate poorer long-term survival. The purpose of this study was to compare retrospectively perioperative safety and long-term results in patients undergoing surgery for OCRC following preoperative colonic decompression with that in those undergoing elective surgery alone for nonobstructing colorectal cancer (CRC). A total of 656 consecutive CRC patients undergoing colectomy between 2001 and 2011 at our institute were eligible for inclusion in the study. The patients were divided into an OCRC group, which included 104 patients undergoing colectomy with preoperative colonic decompression, and a CRC group, which included 552 patients undergoing colectomy alone. Morbidity, mortality, and prognosis were assessed. In the OCRC group, decompression was performed by nasointestinal tube in 42 patients (40.4%), transanal tube in 15 (14.4%), and colostomy in 47 (45.2%). The mortality rate was 0% in the OCRC group and 0.4% in the CRC group (2 of 552 patients). The morbidity rate was 44.8% in the OCRC group (48 of 104 patients) and 36.6% in the CRC group (202 of 552 patients). The 5-year overall survival rate was 69.5% in the OCRC group and 72.9% in the CRC group [hazard ratio 0.76; 95% confidence interval, 0.35 to 1.16; P = 0.48)]. No statistically significant difference in survival was observed between the 2 groups in stage II, III, or IV, or overall. No difference was observed in safety or survival between advanced OCRC patients undergoing preoperative colonic decompression and advanced non-obstructing CRC patients undergoing surgery alone.

D2 gastrectomy with versus without bursectomy for gastric cancer.

OBJECTIVES: The purpose of this study was to determine the survival benefit of bursectomy by retrospectively comparing the prognosis in patients undergoing D2 lymphadenectomy and gastrectomy (D2 gastrectomy) with bursectomy for gastric cancer with that in patients undergoing D2 gastrectomy alone. METHODS: A total of 254 consecutive stage IA to IIIC gastric cancer patients undergoing curative intent surgery between 2004 and 2009 were enrolled. The patients were divided into 2 groups: a bursectomy group, which included patients undergoing curative D2 gastrectomy with bursectomy by one surgeon, and a nonbursectomy group, which included those undergoing curative D2 gastrectomy alone by other surgeons. RESULTS: No statistically significant difference was observed in the number of metastatic nodes or penetration of the serosa between the 2 groups. The overall incidence of surgery-related complications was 24.0% in the bursectomy group (29 of 121 patients) and 25.6% in the nonbursectomy group (34 of 133 patients). The 5-year overall survival rate was 85.8% in the bursectomy group and 80.8% in the nonbursectomy group (hazard ratio 0.82; 95% confidence interval, 0.37-1.74; P=0.60). CONCLUSIONS: The results of this retrospective study indicate no survival benefit for bursectomy plus D2 gastrectomy over D2 gastrectomy alone.

A case of complete response to S-1 plus CDDP in early-stage mucosal esophageal cancer.

We report a case of early-stage mucosal esophageal cancer, showing a complete response to S-1 and cis-diamminedichloplatinum (CDDP). The patient was a 67-year-old man with synchronous double primary early-stage mucosal esophageal and advanced gastric cancer. We planned neoadjuvant chemotherapy with S-1 and CDDP for the advanced gastric cancer and endoscopic mucosal resection for the early-stage esophageal cancer. After the first course of chemotherapy, the endoscopy revealed that the esophageal cancer had become a normal mucosal lesion, and the biopsy was negative for cancer. We diagnosed a complete response to S-1 and CDDP in early-stage esophageal cancer. After two courses of chemotherapy, distal gastrectomy was performed. The patient is still alive with no sign of recurrence at 16 months after the disappearance of the original tumor. These results suggest that chemotherapy with S-1 plus CDDP may be effective in early-stage esophageal cancer.