Long-term follow-up results of endorectal balloon-assisted helical tomotherapy for localized prostate cancer patients in the high-risk group of gastrointestinal adverse events

BackgroundEndorectal balloon (ERB) has been shown to reduce rectal radiation dose and late gastrointestinal toxicities in patients with prostate cancer. However, the usefulness of ERBs for patients with prostate cancer whose rectal shape or size is suboptimal has not been investigated. The purpose of this study was to present the long-term follow-up results of ERB-assisted helical tomotherapy for localized prostate cancer patients whose initial radiation treatment planning (RTP) was unacceptable due to suboptimal rectal shape or size.Materials and methodsOf 541 consecutive patients with localized prostate cancer, 10 were included in this study whose RTPs without ERBs did not meet dose constraints due to: 1) Intestinal intrusion, 2) Small rectum; or 3) Unstable rectal shape. We re-planned using ERBs and delivered 76 Gy in 38 fractions, and evaluated the long-term usefulness and safety of ERB-assisted helical tomotherapy.ResultsAt a median follow-up of 109 months, there were no local recurrences of prostate cancer. The overall, cause-specific, and progression-free survivals at 10 years were 90.0%, 100%, and 83%, respectively. Adverse events of grade 3 or higher were not observed during or after ERB-assisted helical tomotherapy.ConclusionsWhen intestinal intrusion, a small rectum, or an unstable rectal shape is an obstacle for administering helical tomotherapy, ERBs might be the solution.     

Cyclooxygenase-2 is required for activated pancreatic stellate cells to respond to proinflammatory cytokines

Cyclooxygenase-2 (COX-2) mediates various inflammatory responses and is expressed in pancreatic tissue from patients with chronic pancreatitis. To examine the role of COX-2 in chronic pancreatitis, we investigated its participation in regulating functions of pancreatic stellate cells (PSCs), using isolated rat PSCs. COX-2 was expressed in culture-activated PSCs but not in freshly isolated quiescent PSCs. TGF-1, IL-1, and IL-6 enhanced COX-2 expression in activated PSCs, concomitantly increasing the expression of -smooth muscle actin (-SMA), a parameter of PSC activation. The COX-2 inhibitor NS-398 blocked culture activation of freshly isolated quiescent PSCs. NS-398 also inhibited the enhancement of -SMA expression by TGF-1, IL-1, and IL-6 in activated PSCs. These data indicate that COX-2 is required for the initiation and promotion of PSC activation. We further investigated the mechanism by which cytokines enhance COX-2 expression in PSCs. Adenovirus-mediated expression of dominant negative Smad2/3 inhibited the increase in expression of COX-2, -SMA, and collagen-1 mediated by TGF-1 in activated PSCs. Moreover, dominant negative Smad2/3 expression attenuated the expression of COX-2 and -SMA enhanced by IL-1 and IL-6. Anti-TGF- neutralizing antibody also attenuated the increase in COX-2 and -SMA expression caused by IL-1 and IL-6. IL-6 as well as IL-1 enhanced TGF-1 secretion from PSCs. These data indicate that Smad2/3-dependent pathway plays a central role in COX-2 induction by TGF-1, IL-1, and IL-6. Furthermore, IL-1 and IL-6 promote PSC activation by enhancing COX-2 expression indirectly through Smad2/3-dependent pathway by increasing TGF-1 secretion from PSCs. transforming growth factor-; interleukin; Smad; autocrine; pancreatic fibrosis     

Occurrence of a nonsulfated chondroitin proteoglycan in the dried saliva of Collocalia swiftlets (edible bird's-nest)

Despite their wide occurrence, proteoglycans (PGs) have never been isolated from the saliva of higher animals. We found that the Collocalia glycoproteins isolated from edible birds'-nests (the dried forms of regurgitated saliva of male Collocalia swiftlets) were rich in a PG containing nonsulfated chondroitin glycosaminoglycans (GAGs). We have devised a method to isolate a PG from the water extract of the white nest built by Aerodramus fuciphagus (white nest swiftlets) with a yield of 2-mg PG per gram nest. This PG contained 83% of carbohydrates, of which 79% were GalNAc and GlcUA (D-glucuronic acid) in an equimolar ratio. By using chondroitin AC lyase, the structure of GAGs in this PG was established to be chondroitin ( --> 4GlcUAbeta1 --> 3GalNAcbeta1 --> )(n) chains. The average molecular mass of the chondroitin chain was estimated to be 49 kDa by gel filtration. We have isolated a linkage region hexasaccharide, DeltaHexUAalpha1 --> 3GalNAcbeta1 --> 4GlcUAbeta1 --> 3Galbeta1 --> 3Galbeta1 --> 4Xyl, from this PG by chondroitinase ABC digestion to show that the GAGs in this PG are also linked to the core protein through the common tetrasaccharide linker, GlcUAbeta1 --> 3Galbeta1 --> 3Galbeta1 --> 4Xyl, found in various PGs. As water was not effective in extracting uronic acid-containing glycoconjugates from the black nest built by black nest swiftlets (A. maximus), we used 4 M guanidium chloride and anion-exchange chromatography in the presence of urea to extract and isolate about 30 mg of a chondroitin PG preparation from 10 g of the desialylated black nest. As the biological significance of chondroitin is still not well understood, bird's nest should become a convenient source for preparing this unique GAG to study its biological functions.     

Epithelial Na+ channel and ion transport in human nasal polyp and paranasal sinus mucosa

The purpose of the present study is to characterize the ENaC-mediated Na+ absorption in human upper airway epithelia, nasal cavity, and paranasal sinus. To perform the purpose, we obtained epithelial cells from human nasal polyp (NP) and paranasal sinus mucosa (PSM) by endoscopic surgery. We measured the short-circuit current (I(sc)) sensitive to benzamil (a specific ENaC blocker). The benzamil-sensitive I(sc) (Na+ absorption) in NP was larger than that in PSM. The mRNA expression of three subunits of ENaC was as follows: alpha>beta>gamma in both tissue, NP and MS. The mRNA expression of gamma subunit of ENaC in NP was larger than that in PSM, but no difference of mRNA expression of alpha or beta ENaC subunit between NP and PSM was detected. We found correlation of the Na+ absorption to mRNA expression of gamma ENaC in NP and PSM. Forskolin diminished the Na+ absorption associated with an increase in Cl- secretion. These observations suggest that: (1) human NP absorbs more ENaC-mediated Na+ than PSM, (2) expression of gamma ENaC in plays a key role in the ENaC-mediated Na+ absorption in NP and PSM, and (3) cAMP diminishes the ENaC-mediated Na+ absorption by stimulating Cl- secretion (diminution of driving force for Na+ absorption) in NP and PSM.     

Preparation and preliminary evaluation of a biotin-targeted, lectin-targeted dendrimer-based probe for dual-modality magnetic resonance and fluorescence imaging

A novel approach for the preparation of a biotinylated dendrimer-based MRI agent 5 is described, in which a unique disulfide bond in the core of the Gd(III)-1B4M-DTPA chelated G2 PAMAM dendrimer was reduced and then attached to a maleimide-functionalized biotin. The new MRI agent 5 features a well-defined dendron structure and a unique biotin functionality. Immobilization of up to four copies of biotinylated dendrimer 5 to fluorescently labeled avidin yields a supramolecular avidin-biotin-dendrimer-Gd(III) complex. Validation of the complex in mice bearing ovarian cancer tumors demonstrates that the avidin-biotin-dendrimer targeting system efficiently targets and delivers sufficient amounts of chelated Gd(III) and fluorophores (e.g., Rhodamine green) to ovarian tumors to produce visible changes in the tumors by both MRI and optical imaging, respectively. Thus, the avidin-biotin-dendrimer complex may be used as a tumor-targeted probe for dual-modality magnetic resonance and fluorescence imaging.     

Electric Stimulus Opens Intercellular Spaces in Skin

Iontophoresis is a technology for transdermal delivery of ionic small medicines by faint electricity. Since iontophoresis can noninvasively deliver charged molecules into the skin, this technology could be a useful administration method that may enhance patient comfort. Previously, we succeeded in the transdermal penetration of positively charged liposomes (diameters: 200–400 nm) encapsulating insulin by iontophoresis (Kajimoto, K., Yamamoto, M., Watanabe, M., Kigasawa, K., Kanamura, K., Harashima, H., and Kogure, K. (2011) Int. J. Pharm. 403, 57–65). However, the mechanism by which these liposomes penetrated the skin was difficult to define based on general knowledge of principles such as electro-repulsion and electro-osmosis. In the present study, we confirmed that rigid nanoparticles could penetrate into the epidermis by iontophoresis. We further found that levels of the gap junction protein connexin 43 protein significantly decreased after faint electric stimulus (ES) treatment, although occludin, CLD-4, and ZO-1 levels were unchanged. Moreover, connexin 43 phosphorylation and filamentous actin depolymerization in vivo and in vitro were observed when permeation of charged liposomes through intercellular spaces was induced by ES. Ca²⁺ inflow into cells was promoted by ES with charged liposomes, while a protein kinase C inhibitor prevented ES-induced permeation of macromolecules. Consequently, we demonstrate that ES treatment with charged liposomes induced dissociation of intercellular junctions via cell signaling pathways. These findings suggest that ES could be used to regulate skin physiology.     

Interferon-α Acts on the S/G2/M Phases to Induce Apoptosis in the G1 Phase of an IFNAR2-expressing Hepatocellular Carcinoma Cell Line

Interferon-α (IFN-α) is used clinically to treat hepatocellular carcinoma (HCC), although the detailed therapeutic mechanisms remain elusive. In particular, IFN-α has long been implicated in control of the cell cycle, but its actual point of action has not been clarified. Here, using time lapse imaging analyses of the human HCC cell line HuH7 carrying a fluorescence ubiquitination-based cell cycle indicator (Fucci), we found that IFN-α induced cell cycle arrest in the G₀/G₁ phases, leading to apoptosis through an IFN-α type-2 receptor (IFNAR2)-dependent signaling pathway. Detailed analyses by time lapse imaging and biochemical assays demonstrated that the IFN-α/IFNAR2 axis sensitizes cells to apoptosis in the S/G₂/M phases in preparation for cell death in the G₀/G₁ phases. In summary, this study is the first to demonstrate the detailed mechanism of IFN-α as an anticancer drug, using Fucci-based time lapse imaging, which will be informative for treating HCC with IFN-α in clinical practice.     

Development of Nanoparticles Incorporating a Novel Liposomal Membrane Destabilization Peptide for Efficient Release of Cargos into Cancer Cells

In anti-cancer therapy mediated by a nanoparticle-based drug delivery system (DDS), overall efficacy depends on the release efficiency of cargos from the nanoparticles in the cancer cells as well as the specificity of delivery to tumor tissue. However, conventional liposome-based DDS have no mechanism for specifically releasing the encapsulated cargos inside the cancer cells. To overcome this barrier, we developed nanoparticles containing a novel liposomal membrane destabilization peptide (LMDP) that can destabilize membranes by cleavage with intramembranous proteases on/in cancer cells. Calcein encapsulated in liposomes modified with LMDP (LMDP-lipo) was effectively released in the presence of a membrane fraction containing an LMDP-cleavable protease. The release was inhibited by a protease inhibitor, suggesting that LMDP-lipo could effectively release its cargo into cells in response to a cancer-specific protease. Moreover, when LMDP-lipo contained fusogenic lipids, the release of cargo was accelerated, suggesting that the fusion of LMDP-lipo with cellular membranes was the initial step in the intracellular delivery. Time-lapse microscopic observations showed that the release of cargo from LMDP-lipo occurred immediately after association of LMDP-lipo with target cells. Consequently, LMDP-lipo could be a useful nanoparticle capable of effective release of cargos specifically into targeted cancer cells.