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Although the analysis of genetic interactions and networks is a powerful approach to understanding biology, it has not been applied widely to the pathogenic yeast Candida albicans Here, we describe the use of both screening and directed genetic interaction studies based on complex haploinsufficiency to probe the function of the R: egulation of A: ce2 and M: orphogenesis (RAM) pathway in C. albicans A library of 5200 Tn7-mutagenized derivatives of a parental strain heterozygous at CBK1, the key kinase in the RAM pathway, was screened for alterations in serum-induced filamentation. Following confirmation of phenotypes and identification of insertion sites by sequencing, a set of 36 unique double heterozygous strains showing complex haploinsufficiency was obtained. In addition to a large set of genes regulated by the RAM transcription factor Ace2, genes related to cell wall biosynthesis, cell cycle, polarity, oxidative stress, and nitrogen utilization were identified. Follow-up analysis led to the first demonstration that the RAM pathway is required for oxidative stress tolerance in a manner related to the two-component-regulated kinase Chk1 and revealed a potential direct connection between the RAM pathway and the essential Mps1 spindle pole-related kinase. In addition, genetic interactions with CDC42-related genes MSB1, a putative scaffold protein, and RGD3, a putative Rho GTPase-activating protein (GAP) were identified. We also provide evidence that Rgd3 is a GAP for Cdc42 and show that its localization and phosphorylation are dependent on Cbk1., (Copyright © 2016 by the Genetics Society of America.)

Purpose: To describe the morphological distribution of host tissue and microbial biofilms on the intraocular surfaces of Boston type I keratoprostheses (KPros) explanted because of corneal melt., Methods: Retrospective study of scanning electron microscopy (SEM) images from 4 explanted Boston type I KPros composed of polymethylmethacrylate and titanium. SEM images of KPro-associated ocular surfaces were reviewed for the presence of inflammatory cells, microbes, and/or biofilm formation. One sterile type I KPro was also imaged to serve as a (device only) control., Results: All 4 KPros were explanted because of culture-negative, clinically "sterile" donor corneal melt with impending KPro extrusion. In all cases, the rough, irregular surfaces of the device harbored more adherent corneal epithelium and stromacytes, inflammatory cells, and bacteria than the smooth, polished surface of the KPro optic. Two KPros showed not only evidence of prior bacterial colonization but marked biofilm formation., Conclusions: SEM images of explanted KPros explanted for "sterile" corneal melt demonstrated evidence of biofilm formation despite negative donor corneal cultures and the absence of clinical suspicion for infection. These results suggest that "sterile" corneal melt may be due to inflammatory host responses to low microbial burdens as seen in biofilms and/or released antigens after antibiotic-induced lysis. There was increased adherence of host tissue cells and microbial biofilms on the nonpolished surfaces of the KPro. Polishing the intraocular polymethylmethacrylate and titanium KPro surfaces may decrease microbial adhesion and biofilm formation in human subjects with KPros, but what impact this will have on rates of postoperative endophthalmitis is unknown.

Purpose: Tissues and biomaterials used for corneal surface repair require fulfilling specific optical standards prior to implantation in the patient. However, there is not a feasible evaluation method to be applied in clinical or Good Manufacturing Practice settings. In this study, we describe and assess an innovative easy-applied photographic-based method (PBM) for measuring functional optical blurring and transparency in corneal surface grafts., Methods: Plastic compressed collagen scaffolds (PCCS) and multilayered amniotic membranes (AM) samples were optically and histologically evaluated. Transparency and image blurring measures were obtained by PBM, analyzing photographic images of a standardized band pattern taken through the samples. These measures were compared and correlated to those obtained applying the Inverse Adding-Doubling (IAD) technique, which is the gold standard method., Results: All the samples used for optical evaluation by PBM or IAD were histological suitable. PCCS samples presented transmittance values higher than 60%, values that increased with increasing wavelength as determined by IAD. The PBM indicated that PCCS had a transparency ratio (TR) value of 80.3 ± 2.8%, with a blurring index (BI) of 50.6 ± 4.2%. TR and BI obtained from the PBM showed a high correlation (ρ>|0.6|) with the diffuse transmittance and the diffuse reflectance, both determined using the IAD (p<0.005). The AM optical properties showed that there was a largely linear relationship between the blurring and the number of amnion layers, with more layers producing greater blurring., Conclusions: This innovative proposed method represents an easy-applied technique for evaluating transparency and blurriness of tissues and biomaterials used for corneal surface repair.

Unlabelled: We describe a technique for image-guided femtosecond laser-assisted cataract surgery in a case of Peters anomaly type 2. The femtosecond laser technology enabled reliable construction of a complete capsulotomy despite central corneal opacification and a tented anterior capsule. Use of an image-guided femtosecond laser for cataract extraction in Peters anomaly type 2 is a safe method for removing these complex cataracts while preserving capsular and corneal integrity., Financial Disclosure: Dr. De la Cruz is a consultant to Alcon Surgical, Inc. No other author has a financial or proprietary interest in any material or method mentioned., (Copyright © 2015 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.)

Drought severely limits forage productivity of C4 grasses across the tropics. The avoidance of water deficit by increasing the capacity for water uptake or by controlling water loss are common responses in forage C4 grasses. Napier grass (Pennisetum purpureum) and Brachiaria hybrid cv. Mulato II are tropical C4 grasses used for livestock production due to their reputed resistance to drought conditions. However, there is scant information on the mechanisms used by these grasses to overcome water-limited conditions. Therefore, assessments of cumulative transpired water, shoot growth, leaf rolling, leaf gas exchange, dry mass production and a number of morpho-physiological traits were recorded over a period of 21 days under well-watered or drought conditions. Drought reduced shoot dry mass of both grasses by 35 %, yet each grass exhibited contrasting strategies to cope with water shortage. Napier grass transpired most available water by the end of the drought treatment, whereas a significant amount of water was still available for Mulato II. Napier grass maintained carbon assimilation until the soil was fairly dry, whereas Mulato II restricted water loss by early stomatal closure at relatively wet soil conditions. Our results suggest that Napier grass exhibits a 'water-spending' behaviour that might be targeted to areas with intermittent drought stress, whereas Mulato II displays a 'water-saving' nature that could be directed to areas with longer dry periods., (Published by Oxford University Press on behalf of the Annals of Botany Company.)

Background: Being able to estimate (predict) the final spectrum of reflectance of a biomaterial, especially when the final color and appearance are fundamental for their clinical success (as is the case of dental resin composites), could be a very useful tool for the industrial development of these type of materials. The main objective of this study was the development of predictive models which enable the determination of the reflectance spectrum of experimental dental resin composites based on type and quantity of pigments used in their chemical formulation., Methods: 49 types of experimental dental resin composites were formulated as a mixture of organic matrix, inorganic filler, photo activator and other components in minor quantities (accelerator, inhibitor, fluorescent agent and 4 types of pigments). Spectral reflectance of all samples were measured, before and after artificial chromatic aging, using a spectroradiometer. A Multiple Nonlinear Regression Model (MNLR) was used to predict the values of the Reflectance Factors values in the visible range (380 nm-780 nm), before and after aging, from % Pigment (%P1, %P2, %P3 and %P4) within the formulation., Results: The average value of the prediction error of the model was 3.46% (SD: 1.82) across all wavelengths for samples before aging and 3.54% (SD: 1.17) for samples after aging. The differences found between the predicted and measured values of the chromatic coordinates are smaller than the acceptability threshold and, in some cases, are even below the perceptibility threshold., Conclusions: Within the framework of this pilot study, the nonlinear predictive models developed allow the prediction, with a high degree of accuracy, of the reflectance spectrum of the experimental dental resin composites.

Waterlogging is one of the major factors limiting the productivity of pastures in the humid tropics. Brachiaria humidicola is a forage grass commonly used in zones prone to temporary waterlogging. Brachiaria humidicola accessions adapt to waterlogging by increasing aerenchyma in nodal roots above constitutive levels to improve oxygenation of root tissues. In some accessions, waterlogging reduces the number of lateral roots developed from main root axes. Waterlogging-induced reduction of lateral roots could be of adaptive value as lateral roots consume oxygen supplied from above ground via their parent root. However, a reduction in lateral root development could also be detrimental by decreasing the surface area for nutrient and water absorption. To examine the impact of waterlogging on lateral root development, an outdoor study was conducted to test differences in vertical root distribution (in terms of dry mass and length) and the proportion of lateral roots to the total root system (sum of nodal and lateral roots) down the soil profile under drained or waterlogged soil conditions. Plant material consisted of 12 B. humidicola accessions from the gene bank of the International Center for Tropical Agriculture, Colombia. Rooting depth was restricted by 21 days of waterlogging and confined to the first 30 cm below the soil surface. Although waterlogging reduced the overall proportion of lateral roots, its proportion significantly increased in the top 10 cm of the soil. This suggests that soil flooding increases lateral root proliferation of B. humidicola in the upper soil layers. This may compensate for the reduction of root surface area brought about by the restriction of root growth at depths below 30 cm. Further work is needed to test the relative efficiency of nodal and lateral roots for nutrient and water uptake under waterlogged soil conditions., (Published by Oxford University Press on behalf of the Annals of Botany Company.)

Purpose: To analyze iris behavior and angle status using serial anterior segment optical coherence tomography (AS-OCT) after Boston keratoprosthesis (KPro)., Methods: A prospective cases series consisted of 11 eyes with implanted type 1 KPro at a tertiary care institution. The patients underwent preoperative and serial postoperative AS-OCT imaging. The main outcome measures included anterior chamber angle (ACA) at representative meridians (0 degrees, 45 degrees, 90 degrees, 135 degrees, 180 degrees, 225 degrees, 270 degrees, and 315 degrees), grading of total ACA as open (>10 degrees), shallow (≥1 degrees and ≤10 degrees) or closed (<1 degrees), preoperative anterior chamber depth (ACD), postoperative anatomical and functional ACD, and the presence of iris-back plate touch and peripheral anterior synechiae., Results: The mean follow-up with serial AS-OCT was 13.1 months. Preoperatively, 54.5%, 27.3%, and 18.2% of the eyes had open, shallow, and closed angles, respectively. The mean change in the effective ACD decreased postoperatively by 1.61 mm. At 0 degree and 180 degree meridians, the ACA decreased by a mean change of 6.95 degrees and 8.40 degrees, with a mean change of 8.12 degrees for all meridians. Eight (72.7%) eyes had synechiae with 7.3 and 6.7 clock hours of peripheral anterior synechiae and iris-back plate touch. At the last follow-up, 7 (63.6%) eyes had considerable progression of angle closure (change in grading of total angle), and 18.2%, 36.4%, and 45.5% had open, shallow, and closed angles, respectively., Conclusions: KPro implantation induced progressive angle closure, shallowing of the anterior chamber, and synechiae formation that is not visible on clinical examination. Serial AS-OCT plays an important role in the detection and monitoring of progressive angle closure, and clinical correlation is needed to assess the association with glaucoma development or progression.

Advances in the development of cornea substitutes by tissue engineering techniques have focused on the use of decellularized tissue scaffolds. In this work, we evaluated different chemical and physical decellularization methods on small intestine tissues to determine the most appropriate decellularization protocols for corneal applications. Our results revealed that the most efficient decellularization agents were the SDS and triton X-100 detergents, which were able to efficiently remove most cell nuclei and residual DNA. Histological and histochemical analyses revealed that collagen fibers were preserved upon decellularization with triton X-100, NaCl and sonication, whereas reticular fibers were properly preserved by decellularization with UV exposure. Extracellular matrix glycoproteins were preserved after decellularization with SDS, triton X-100 and sonication, whereas proteoglycans were not affected by any of the decellularization protocols. Tissue transparency was significantly higher than control non-decellularized tissues for all protocols, although the best light transmittance results were found in tissues decellularized with SDS and triton X-100. In conclusion, our results suggest that decellularized intestinal grafts could be used as biological scaffolds for cornea tissue engineering. Decellularization with triton X-100 was able to efficiently remove all cells from the tissues while preserving tissue structure and most fibrillar and non-fibrillar extracellular matrix components, suggesting that this specific decellularization agent could be safely used for efficient decellularization of SI tissues for cornea TE applications.

Objectives: Evaluate color and translucency of both non-colored and colored zirconia ceramics and compare them with corresponding properties of human dentine., Methods: 0.5mm thick samples of human (MSHD) and bovine (MSBD) dentine were obtained from 5 extracted human anterior maxillary teeth and 5 bovine incisors, respectively. The zirconia systems evaluated (0.5mm samples) were: (1) IPS e.max(®) ZirCAD sintered samples: (a) regular (ZC1); (b) colored - coloring liquid CL2 (ZC2); (2) LAVA™ Zirconia sintered samples: (a) regular (LV1); (b) colored - coloring liquid FS3 (LV2). Translucency parameter (TP) and ΔE(ab)(*) and ΔE(00) color differences (with respect to the MSHD) were calculated. Spectral reflectance curves were compared using the VAF coefficient., Results: Reflectance spectral behaviour of MSBD and MSHD were similar (VAF=99.3%). All zirconia samples showed higher spectral reflectance values than the MSHD. Lightness values obtained for all zirconia ceramics (especially LV) were higher than MSHD and MSBD. The range of color differences, with respect to MSHD, was 6.9-20.5 for ΔE(ab)(*) and 4.9-15.6 for ΔE(00). Colored samples showed lower color differences with respect to MSHD. The highest TP values were found for IPS e.max(®) ZirCAD system and the lowest for LAVA™ Zirconia system. In terms of translucency, MSHD, MSBD and zirconia ceramics showed no statistically significant differences., Conclusions: In terms of translucency, the studied zirconia systems could satisfactorily replace the human dentine within a dental restoration but, in order to produce a clinically acceptable match, it is necessary to carefully adjust the color of these systems., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Objective: To evaluate whether bilayer chitosan scaffolding (BChS) can provide a watertight dural closure and permit regeneration by fibroblasts in an experimental in vivo model., Methods: In the in vitro phase, BChS was elaborated and the following characteristics were evaluated: pore size, thickness, water absorption capacity, tensile strength, strain, and toughness. In the second in vivo phase, 27 durectomized New Zealand rabbits were randomly assigned into three duraplasty groups with autologous dura, collagen matrix (CM), or BChS. In all groups, fluid leakage pressure was measured at 10, 21, or 180 days. Histology response to regeneration was evaluated through hematoxylin and eosin stain., Results: BChS was standardized to obtain bilayer scaffoldings with a nonporous layer and a porous layer. The pore size was 10 μm, total thickness was 400 μm, strain was 57.8%, and tensile strength was 5.5 gr/mm(2). The physical characteristics of BChS allowed dural closure without cerebrospinal fluid (CSF) leak. There were no differences in fluid leakage pressures between the BChS, dura, and CM groups. Histologic analysis showed fibroblast migration with adequate dural regeneration., Conclusions: BChS is an ideal alternative for a watertight dural closure because it can be sutured, and it induces organized regeneration with fibroblasts without evidence of fibrosis., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Purpose: To examine the transparency characteristics at different times of development in the culture of 2 different types of human corneal stroma substitutes generated by tissue engineering using human fibrin or human fibrin and 0.1% agarose, with human keratocytes entrapped within., Methods: The transparency of these artificial corneal stromas was analyzed after 1, 7, 14, 21, and 28 days of development in culture by determining the scattering and absorption coefficients from the spectral reflectance data of each sample using the Kubelka-Munk equations., Results: The scattering coefficient of both types of bioengineered tissues tended to increase with culture time and wavelength until 550 nm, whereby a slight decrease was observed for longer wavelengths. In general, the spectral distribution of the Kubelka-Munk scattering coefficient of the fibrin-agarose corneal constructs was more stable than that of the fibrin constructs. The absorption coefficient of the human fibrin and fibrin-agarose corneal substitutes tended to decrease with increasing wavelength, and their absolute values were higher for fibrin-agarose than for fibrin scaffolds, especially for short wavelengths. In addition, the spectral transmittance behavior of both types of tissue analyzed was similar to the ones of the theoretical and control corneas, with absolute values above 90% for all wavelengths at 28 days of development., Conclusions: The transparency, scattering, and absorption of both fibrin and fibrin-agarose corneal stroma substitutes indicate that these new biomaterials could be adequate for clinical use.

Chiari disease (or malformation) is in general a congenital condition characterized by an anatomic defect of the base of the skull, in which the cerebellum and brain stem herniate through the foramen magnum into the cervical spinal canal. The onset of Chiari syndrome symptoms usually occurs in the second or third decade (age 25 to 45 years). Symptoms may vary between periods of exacerbation and remission. The diagnosis of Chiari type I malformation in patients with or without symptoms is established with neuroimaging techniques. The most effective therapy for patients with Chiari type I malformation/syringomyelia is surgical decompression of the foramen magnum, however there are non-surgical therapy to relieve neuropathic pain: either pharmacological and non-pharmacological. Pharmacological therapy use drugs that act on different components of pain. Non-pharmacological therapies are primarily based on spinal or peripheral electrical stimulation. It is important to determine the needs of the patients in terms of health-care, social, educational, occupational, and relationship issues, in addition to those derived from information aspects, particularly at onset of symptoms. Currently, there is no consensus among the specialists regarding the etiology of the disease or how to approach, monitor, follow-up, and treat the condition. It is necessary that the physicians involved in the care of people with this condition comprehensively approach the management and follow-up of the patients, and that they organize interdisciplinary teams including all the professionals that can help to increase the quality of life of patients.