Cellular characterisation of Candida tropicalis presenting fluconazole-related trailing growth

We assessed fluconazole susceptibility in 52 Candida tropicalis clinical strains using seven antifungal susceptibility methods, including broth microdilution (BMD) [standard M27 A3 (with neutral and acid pH), ATB Fungus 3, Vitek 2 system and flow cytometric analysis] and agar-based methods (disk diffusion and E-test). Trailing growth, detection of cell-associated secreted aspartic proteases (Saps) and morphological and ultrastructural traits of these clinical strains were also examined. The ranges of fluconazole 24 h-minimum inhibitory concentration (MIC) values were similar among all methods. The essential agreement among the methods used for MIC determinations was excellent and all methods categorised all strains as susceptible, except for one strain that showed a minor error. The presence of the trailing effect was assessed by six methods. Trailing positivity was observed for 86.5-100% of the strains. The exception was the BMD-Ac method where trailing growth was not observed. Morphological and ultrastructural alterations were detected in C. tropicalis trailing cells, including mitochondrial swelling and cell walls with irregular shapes. We tested the production of Saps in 13 C. tropicalis strains expressing trailing growth through flow cytometry. Our results showed that all of the C. tropicalis strains up-regulated surface Sap expression after 24 h or 48 h of exposure to fluconazole, which was not observed in untreated yeast strains. We concluded that C. tropicalis strains expressing trailing growth presented some particular features on both biological and ultrastructural levels.     

Fat pad-derived mesenchymal stem cells as a potential source for cell-based adipose tissue repair strategies

Background: Mesenchymal stem cells are able to undergo adipogenic differentiation and present a possible alternative cell source for regeneration and replacement of adipose tissue. The human infrapatellar fat pad is a promising source of mesenchymal stem cells with many source advantages over from bone marrow. It is important to determine whether a potential mesenchymal stem‐cell exhibits tri‐lineage differentiation potential and is able to maintain its proliferation potential and cell‐surface characterization on expansion in tissue culture. We have previously shown that mesenchymal stem cells derived from the fat pad can undergo chondrogenic and osteogenic differentiation, and we characterized these cells at early passage. In the study described here, proliferation potential and characterization of fat pad‐derived mesenchymal stem cells were assessed at higher passages, and cells were allowed to undergo adipogenic differentiation. Materials and methods: Infrapatellar fat pad tissue was obtained from six patients undergoing total knee replacement. Cells isolated were expanded to passage 18 and proliferation rates were measured. Passage 10 and 18 cells were characterized for cell‐surface epitopes using a range of markers. Passage 2 cells were allowed to undergo differentiation in adipogenic medium. Results: The cells maintained their population doubling rates up to passage 18. Cells at passage 10 and passage 18 had cell‐surface epitope expression similar to other mesenchymal stem cells previously described. By staining it was revealed that they highly expressed CD13, CD29, CD44, CD90 and CD105, and did not express CD34 or CD56, they were also negative for LNGFR and STRO1. 3G5 positive cells were noted in cells from both passages. These fat pad‐derived cells had adipogenic differentiation when assessed using gene expression for peroxisome proliferator‐activated receptor γ2 and lipoprotein lipase, and oil red O staining. Discussion: These results indicate that the cells maintained their proliferation rate, and continued expressing mesenchymal stem‐cell markers and pericyte marker 3G5 at late passages. These results also show that the cells were capable of adipogenic differentiation and thus could be a promising source for regeneration and replacement of adipose tissue in reconstructive surgery.     

Differential hypogeous sporocarp production from Nothofagus dombeyi and N. pumilio forests in southern Argentina

Mycorrhizal fungi that form hypogeous sporocarps are an important component of the temperate forest soil community. In many regions, such as the Nothofagus forest in the Patagonian Andes, this group of fungi has been poorly studied. Here we examined the spring and autumn community composition of "sequestrate fungi", based on sporocarp production in pure forests of Nothofagus dombeyi (evergreen) and N. pumilio (deciduous). We investigated the possible relationships between these communities and environmental factors over 2 y. The rarefaction curves and the minimal richness estimates converged at nearly the same level for each forest type, and the asymptotes suggested that the sampling effort was sufficient to capture most of the hypogeous sporocarp richness in these forest stands. In total 27 species were recovered. Basidiomycota, Ascomycota and Glomeromycota respectively accounted for nine, two and one genera. Species richness of hypogeous sporocarps varied in relation to forest type but not to season (fall and spring), whereas sporocarp biomass varied according to an interaction between season and forest type. Species richness and sporocarp biomass were positively correlated with rainfall and negatively correlated with altitude. In addition sporocarp species richness was positively related to number of trees per transect. We found that two different forest stands, each dominated by different species of Nothofagus, exhibited different hypogeous sporocarp communities.     

Fabrication and Application of Rose Bengal-chitosan Films in Laser Tissue Repair

Photochemical tissue bonding (PTB) is a sutureless technique for tissue repair, which is achieved by applying a solution of rose bengal (RB) between two tissue edges^1,2. These are then irradiated by a laser that is selectively absorbed by the RB. The resulting photochemical reactions supposedly crosslink the collagen fibers in the tissue with minimal heat production³. In this report, RB has been incorporated in thin chitosan films to fabricate a novel tissue adhesive that is laser-activated. Adhesive films, based on chitosan and containing ~0.1 wt% RB, are fabricated and bonded to calf intestine and rat tibial nerves by a solid state laser (λ=532 nm, Fluence~110 J/cm², spot size~0.5 cm). A single-column tensiometer, interfaced with a personal computer, is used to test the bonding strength. The RB-chitosan adhesive bonds firmly to the intestine with a strength of 15 ± 6 kPa, (n=30). The adhesion strength drops to 2 ± 2 kPa (n=30) when the laser is not applied to the adhesive. The anastomosis of tibial nerves can be also completed without the use of sutures. A novel chitosan adhesive has been fabricated that bonds photochemically to tissue and does not require sutures.     

Metformin (dimethyl-biguanide) induced DNA damage in mammalian cells

Metformin (dimethyl-biguanide) is an insulin-sensitizing agent that lowers fasting plasma-insulin concentration, wherefore it's wide use for patients with a variety of insulin-resistant and prediabetic states, including impaired glucose tolerance. During pregnancy it is a further resource for reducing first-trimester pregnancy loss in women with the polycystic ovary syndrome. We tested metformin genotoxicity in cells of Chinese hamster ovary, CHO-K1 (chromosome aberrations; comet assays) and in mice (micronucleus assays). Concentrations of 114.4 μg/mL and 572 μg/mL were used in in vitro tests, and 95.4 mg/kg, 190.8 mg/kg and 333.9 mg/kg in assaying. Although the in vitro tests revealed no chromosome aberrations in metaphase cells, DNA damage was detected by comet assaying after 24 h of incubation at both concentrations. The frequency of DNA damage was higher at concentrations of 114.4 μg/mL. Furthermore, although mortality was not observed in in vitro tests, the highest dose of metformin suppressed bone marrow cells. However, no statistically significant differences were noted in micronuclei frequencies between treatments. In vitro results indicate that chronic metformin exposure may be potentially genotoxic. Thus, pregnant woman undergoing treatment with metformin should be properly evaluated beforehand, as regards vulnerability to DNA damage.     

Effects of self-selected music on strength, explosiveness, and mood

There has been much investigation into the use of music as an ergogenic aid to facilitate physical performance. However, previous studies have primarily focused on predetermined music and aerobic exercise. The purpose of this study was to investigate the effects of self-selected music (SSM) vs. those of no music (NM) on the mood and performance of the athletes performing bench press and squat jump. Twenty resistance trained collegiate men completed 2 experimental conditions, one while listening to SSM and the other with NM. The subjects reported their profile of mood states (POMS) and rating of perceived exertion (RPE) before and after performing 3 sets to failure of the bench press at 75% 1 repetition maximum (1RM) and 3 reps of the squat jump at 30% 1RM. Statistical analyses revealed no differences in squat jump height or relative ground reaction force, but the takeoff velocity (SSM-2.06 ± 0.17 m·s(-1); NM-1.99 ± 0.18 m·s(-1)), rate of velocity development (SSM-5.92 ± 1.46 m·s(-2); NM-5.63 ± 1.70 m·s(-2)), and rate of force development (SSM-3175.61 ± 1792.37 N·s(-1); NM-2519.12 ± 1470.32 N·s(-1)) were greater with SSM, whereas RPE (SSM-5.71 ± 1.37; NM-6.36 ± 1.61) was greater with NM. Bench press reps to failure and RPE were not different between conditions. The POMS scores of vigor (SSM-20.15 ± 5.58; NM-17.45 ± 5.84), tension (SSM-8.40 ± 3.99; NM-6.07 ± 3.26), and fatigue (SSM-8.65 ± 4.49; NM-7.40 ± 4.38) were greater with SSM. This study demonstrated increased performance during an explosive exercise and an altered mood state when listening to SSM. Therefore, listening to SSM might be beneficial for acute power performance.     

Imaging characteristics, tissue distribution, and spread of a novel oncolytic vaccinia virus carrying the human sodium iodide symporter

Introduction

Oncolytic viruses show promise for treating cancer. However, to assess therapy and potential toxicity, a noninvasive imaging modality is needed. This study aims to determine the in vivo biodistribution, and imaging and timing characteristics of a vaccinia virus, GLV-1h153, encoding the human sodium iodide symporter (hNIS.

Methods

GLV-1h153 was modified from GLV-1h68 to encode the hNIS gene. Timing of cellular uptake of radioiodide ¹³¹I in human pancreatic carcinoma cells PANC-1 was assessed using radiouptake assays. Viral biodistribution was determined in nude mice bearing PANC-1 xenografts, and infection in tumors confirmed histologically and optically via Green Fluorescent Protein (GFP) and bioluminescence. Timing characteristics of enhanced radiouptake in xenografts were assessed via ¹²⁴I-positron emission tomography (PET). Detection of systemic administration of virus was investigated with both ¹²⁴I-PET and 99m-technecium gamma-scintigraphy.

Results

GLV-1h153 successfully facilitated time-dependent intracellular uptake of ¹³¹I in PANC-1 cells with a maximum uptake at 24 hours postinfection (P<0.05). In vivo, biodistribution profiles revealed persistence of virus in tumors 5 weeks postinjection at 10⁹ plaque-forming unit (PFU)/gm tissue, with the virus mainly cleared from all other major organs. Tumor infection by GLV-1h153 was confirmed via optical imaging and histology. GLV-1h153 facilitated imaging virus replication in tumors via PET even at 8 hours post radiotracer injection, with a mean %ID/gm of 3.82±0.46 (P<0.05) 2 days after intratumoral administration of virus, confirmed via tissue radiouptake assays. One week post systemic administration, GLV-1h153-infected tumors were detected via ¹²⁴I-PET and 99m-technecium-scintigraphy.

Conclusion

GLV-1h153 is a promising oncolytic agent against pancreatic cancer with a promising biosafety profile. GLV-1h153 facilitated time-dependent hNIS-specific radiouptake in pancreatic cancer cells, facilitating detection by PET with both intratumoral and systemic administration. Therefore, GLV-1h153 is a promising candidate for the noninvasive imaging of virotherapy and warrants further study into longterm monitoring of virotherapy and potential radiocombination therapies with this treatment and imaging modality.