Nakayama K, Takebayashi Y, Hata K, Fujiwaki R, Iida K, Fukumoto M, Miyazaki K
Br J Cancer. 2004 Mar 22;90(6):1204–10
Ovarian tumours of low malignant potential (LMP) are intermediate between adenomas and ovarian carcinomas. These tumours are often associated with a significantly better prognosis than ovarian carcinomas. However, a subset of these tumours can progress and become lethal. In order to seek sensitive diagnostic tools for monitoring patients after surgical operation, we performed a genome-wide scan for loss of heterozygosity (LOH) in 41mucinous LMPs using 91 polymorphic microsatellite markers at an average interval of 50 cM across all of the human chromosomes and 25 LOH markers reportedly associated with ovarian carcinoma. In addition, we assessed whether clinicopathological parameters, microvessel density, Ki-67 labeling index, apoptotic index or p53 overexpression would be useful for predicting the postoperative outcome of LMP patients. Of the 116 markers examined, 19q12 and Xq11-12 showed significant correlation between postoperative progression-free survival time and LOH status (P<0.05).Patients with a high Ki-67 labeling index had a significantly poorer progression-free survival time than those with lower levels (P=0.042). Other clinicopathological factors and immunohistochemical analysis had no correlation with progression-free survival time in this series of patients. When the combination of LOH at 19q12 and/or Xq11-12 was assessed using Cox's regression analysis, patients with tumours that showed LOH at these positions were at greatest risk of progression (P=0.0073). These findings suggest that the identification of LOH at 19q12 and/or Xq11-12 in formermucinous LMP sites should alert the clinician to the presence of a potentially aggressive lesion in the coelomic epithelium, even if a distinction between second primary tumours or recurrence could not be determined.
Nigmatullin R, Lovitt R, Wright C, Linder M, Nakari-Setälä T, Gama M
Colloids Surf B Biointerfaces. 2004 May 15;35(2):125–35
Colloidal probe microscopy has been used to study the interaction between model cellulose surfaces and the role of cellulose binding domain (CBD), peptides specifically binding to cellulose, in interfacial interaction of cellulose surfaces modified with CBDs. The interaction between pure cellulosesurfaces in aqueous electrolyte solution is dominated by double layer repulsive forces with the range and magnitude of the net force dependent on electrolyte concentration. AFM imaging reveals agglomeration of CBD adsorbed on cellulose surface. Despite an increase in surface charge owing to CBD binding to cellulose surface, force profiles are less repulsive for interactions involving, at least, one modified surface. Such changes are attributed to irregularity of the topography of protein surface and non-uniform distribution of surface charges on the surface of modified cellulose.Binding double CBD hybrid protein to cellulose surfaces causes adhesive forces at retraction, whereas separation curves obtained with cellulosemodified with single CBD show small adhesion only at high ionic strength. This is possibly caused by the formation of the cross-links betweencellulose surfaces in the case of double CBD.
Do K, Baker K, Praetorius M, Staecker H
International Congress Series. 2004 Nov;1273:167–70.
Objective: To develop and validate a mouse model of implantation trauma. Study design: Adult CBA mice were anesthetized and a basal turn cochleostomy created. Using a micromanipulator varying amounts of sterile phosphate-buffered saline were injected into the cochleostomy. The cochleostomy was sealed and hearing was then tested by ABR. Animals were also treated with cell death inhibitors prior to injection. Hearing was tested after 24 h allowing the elucidation of cell death pathways after cochlear trauma. Results: Volume changes under 4% of the perilymph volume of the mouse cochlea were well tolerated. Greater volume injections resulted in progressively larger hearing losses with profound hearing loss occurring at 30% of perilymph volume injection. There was a correlation between volume delivered, hearing loss, and histological findings with higher volumes resulting in tears of Reisner's membrane and blood in the perilymph. Addition of caspase inhibitors to the injection mix significantly improved hearing outcomes in high volume injections. Conclusion: This animal model allows evaluation of a cochlear injury in a quantitative fashion. Use of cell death pathway inhibitors significantly improved hearing outcome, suggesting that these substances may play a role in cases where hearing preservation is planned in implant surgery. This mouse model of hydraulic trauma may be used to test a variety of potential protective substances.
Praetorius M, Baker K, Brough DE, Plinkert P, Staecker H
Hear Res. 2007 May;227(1-2):53–8
Recent studies have demonstrated that delivery of genes to the inner ear can achieve a variety of effects ranging from support of auditory neuron survival to protection and restoration of hair cells, demonstrating the utility of vector based gene delivery. Translation of these findings to useful experimental systems or even clinical applications requires a detailed understanding of the pharmacokinetics of gene delivery in the inner ear. Ideal gene delivery systems will employ a well tolerated vector which efficiently transduces the appropriate target cells within a tissue, but spare non-target structures. Adenovectors based on serotype 5 (Ad 5) are commonly used vectors, are easy to construct and have a long track record of efficacious gene transfer in the inner ear. In this study we demonstrate that distribution of Ad5 vector occurs in a basal to apical gradient with rapiddistribution of vector to the vestibule after delivery via a round window cochleostomy. Transduction of the vector and expression of the delivered transgene occurs by 10 min post vector delivery. At 24 h post delivery only 16% of vector that was initially detectable within the inner ear by quantitative PCR remained. Perilymph sampling was used to determine that vector concentrations in perilymph peaked at 30 min post delivery and then declined rapidly. Understanding these basic distribution patterns and parameters for delivery are important for the design of gene delivery vectors and vital for modeling dose responses to achieve safe efficacious delivery of a therapeutic agent.
Huan Z1, Nakayama K, Nakayama N, Ishibashi M, Yeasmin S, Katagiri A, Purwana IN, Iida K, Maruyama R, Fukumoto M, Miyazaki K.
Oncol Rep. 2008 Mar;19(3):775–81
Ovarian carcinomas can progress through two pathways of genomic instability: chromosomal instability (CIN) and microsatellite instability (MSI). However, it is unknown whether these two mechanisms could be distinguished from each other in the molecular characteristics in ovariancarcinomas. We hypothesized that these two pathways are not always independent in ovarian carcinomas. We classified 51 ovarian carcinomasbased on their MSI and CIN status using microsatellite analysis and assessed whether these carcinogenic pathways affect the clinicopathologicalfeatures and patient survival. Of the 51 cases, 77.4% of the tumors were microsatellite stable (MSS), 5.9% were MSI-Low (MSI-L) whilst, 16.7% were MSI-High (MSI-H). Overall, 56.8% of the tumors had at least one loss of heterozygosity (LOH) event, i.e., 56.8% CIN. Notably, we identified a significant degree of overlap between the MSI and CIN pathways. Of the 34 tumors with LOH events (CIN), 5 (14.7%) were MSI-H. In addition, of the 7 tumors that were MSI-H, 5 (71.4%) had one or more LOH events (CIN). We also identified a group of 29.4% of all tumors that did not demonstrate any evidence of either of the two pathways of genomic instability as they were MSS/MSI-L with no evidence of LOH events (CIN negative). Furthermore, patients with CIN with MSS/MSI-L have a significantly shorter overall survival compared to those in other genetic categories (P=0.019). Cox regression analysis revealed that tumors with CIN with MSS/MSI-L exhibit a poor prognostic outcome after adjustment for FIGO stage and grade. These findings suggest that some ovarian carcinomas have a significant degree of overlap between the two pathways of genomic instability and that the genetic classification using microsatellite markers may represent a potential new biomarker of risk prediction in ovarian carcinoma.
Llanos J, Williams PM, Cheng S, Rogers D, Wright C, Pérez A, Cañizares P
Water Res. 2010 Jun;44(11):3522–30
In the present study, the atomic force microscopy (AFM) technique has been used to characterize a Carbosep M5 ceramic membrane(MWCO=10kDa, TiO(2)-ZrO(2) active layer). This membrane was previously used in a polymer supported ultrafiltration (PSU) process to recover copper, using partially ethoxylated polyethylenimine as the water-soluble polymer. The membrane was characterized in four different operationalstates: new, new and cleaned, fouled in a PSU stage and cleaned after a PSU process. The influence of the membrane state on pore opening size distribution and roughness was studied, finding a 16% decrease in the former and a 20% increase in the latter due to foulant deposition upon themembrane active layer. Phase angle distribution was also analyzed to indicate the foulant spreading on the membrane surface. These phase angle measurements can be related to pore opening size and roughness, concluding that the cleaning procedure is not totally effective and that foulant presence on the membrane active layer is not remarkable. Finally, AFM was used to measure the influence of pH on adhesion forces between a silica probe and the membrane active layer. These results can be related to the flux evolution vs pH in PSU experiments, finding both lowest adhesion and highest flux at pH 6.
Walker GP, Medina Ortega KJ
Entomologia Experimentalis et Applicata. 2012 Sep 1;144(3):326–35
Immediately after their stylets penetrate a phloem sieve element, aphids inject saliva into the sieve element for approximately 30–60 s before they begin to ingest phloem sap. This salivation period is recorded as waveform E1 in electrical penetration graph (EPG) monitoring of aphid feeding behavior. It has been hypothesized that the function of this initial period of phloem salivation is to reverse or prevent plugging of the sieve element by one of the plant's phloem defenses: formation of P-protein plugs or callose synthesis in the sieve pores that connect adjacent sieve elements. This hypothesis was tested using the pea aphid, Acyrthosiphon pisum (Harris) (Hemiptera: Aphididae), and faba bean, Vicia faba L. (Fabaceae), as a model system, and the results do not support the hypothesis. In legumes, such as faba bean, P-protein plugs in sieve elements are formed by dispersal of proteinaceous bodies called forisomes. Contrary to the hypothesis, the great majority of sieve element penetrations by pea aphid stylets do not trigger forisome dispersal. Thirteen sieve elements were cryofixed early in phloem phase before the aphids could complete their salivation period and the forisomes were not dispersed in any of the 13 samples. However, in these samples, the aphids completed on average a little over half of their normal E1 salivation period before they were cryofixed. Thus, it is possible that sieve element penetration triggered forisome dispersal in these samples but the abbreviated period of salivation was still sufficient to reverse dispersal. To rule out this possibility, 17 sieve elements were cryofixed during R-pds, which are an EPG waveform associated with sieve element penetration but without the characteristic E1 salivation that occurs during phloem phase. In 16 of the 17 samples, the forisomes were not dispersed. Thus, faba bean sieve elements usually do not form P-protein plugs in response to penetration by pea aphid stylets. Consequently, the characteristic E1 salivation that occurs at the start of each phloem phase does not seem to be necessary to prevent a plugging response because penetration of sieve elements during R-pds does not trigger forisome dispersal despite the absence of E1 salivation. Furthermore, as P-protein plugs do not normally form in response to sieve element penetration, E1 salivation that occurs at the start of each phloem phase is not a response to development of a P-protein plug. Thus, the E1 salivation period at the beginning of the phloem phase appears to have function(s) unrelated to phloem sealing.
Kitade Y, Sumita T, Izumitsu K, Tanaka C
Mycoscience. 2015 Mar;56(2):150–8
In this study, we identified a gene, Ste7, in the genome of Bipolaris maydis encoding a mitogen-activated protein kinase (MAPK) kinase homologous to yeast Ste7. A Δste7 strain was defective in conidiation and conidial germination, and was severely impaired in lesion formation, mainly due to the loss of appressoria, whereas the effect on infectious hyphal development was not significant. The Δste7 strain did not have a maternal ability for sexual reproduction, mainly in protothecium formation. These phenotypes of Δste7were also observed in Δchk1 and Δste11, null mutant strains of MAPK and MAPK kinase kinase, respectively. These observations strongly suggested that Ste7 is a component of the Chk1 MAPK cascade, and is involved in the regulation of various developmental and morphogenetic processes in the asexual and sexual lifecycles of B. maydis.
Ceriotti A, Colman A
Methods Mol Biol. 1995;37:151-78
The large size, resilience, and high translational capacity of the fullgrown Xenopus oocyte has made it a widely used system for the translation of microinjected natural and synthetic mRNAs. The injected and endogenous mRNAs compete for the oocyte translational machinery in a process that normally results in the translation of at least a fraction of the injected mRNA and in a concomitant decrease in the synthesis of endogenous proteins (1). The injected oocytes can survive in vitro for long periods in simple salt media, and a single oocyte can synthesize nanogram amounts of foreign protein/hour, This often allows the analysis to be performed on the material obtained from one or few oocytes. Additionally, the heterologous polypeptides can be subjected to various and posttranslational processing steps (including glycosylation, subunit assembly, movement along the secretory pathway, proteolytic processing, phosphorylation, acetylation, hydroxylation, amidation) many of which do not occur in the standard cell-free translation systems. Because of these and other characteristics, Xenopus oocytes have been successfully used in situations when more than the mere translation of the mRNA was required.
Lacal JC
Methods Mol Biol. 1998;84:139-52
The oocytes of several organisms—most frequently those of the African clawed toad Xenopus laevis—have been used for many years as an excellent system to study regulation of transcription, translation, protein modification processes, secretion, and protein compartmentalization, as well as the expression of heterologous-membrane receptors and their association to specific signaling cascades. Full-grown oocytes are large cells (over 1.2 mm in diameter) that are arrested in late-G2 phase of the first meiosis (Meiosis I), and must progress after physiologrcal stimulus by progesterone to the second meiotrc metaphase (Meiosis II) before fertilization takes place. This process, called oocyte maturation or germinal vesicle breakdown (GVBD), can be easily visualized by the appearance of a small white spot in the animal pole, a consequence of the dissociation of the nuclear envelope. After GVBD is completed, if the oocytes have been fertilized, DNA synthesis takes places with the consequent initiation of the Meiosis II.