PurposeImage-guided prostate biopsies are routinely acquired in the diagnosis and treatment monitoring of prostate cancer, yielding useful tissue for identifying metabolic biomarkers and therapeutic targets. We developed an optimized biopsy tissue culture protocol in combination with [1,6-C-13(2)]glucose labeling and quantitative high-resolution NMR to measure glycolysis and tricarboxcylic acid (TCA) cycle activity in freshly acquired living human prostate biopsies. MethodsWe acquired 34 MRI-ultrasound fusion-guided prostate biopsies in vials on ice from 22 previously untreated patients. Within 15 min, biopsies were transferred to rotary tissue culture in 37 degrees C prostate medium containing [1,6-C-13(2)]glucose. Following 24 h of culture, tissue lactate and glutamate pool sizes and fractional enrichments were quantified using quantitative H-1 high resolution magic angle spinning Carr-Purcell-Meiboom-Gill (CPMG) spectroscopy at 1 degrees C with and without C-13 decoupling. Lactate effluxed from the biopsy tissue was quantified in the culture medium using quantitative solution-state high-resolution NMR. ResultsLactate concentration in low-grade cancer (1.15 0.78 nmol/mg) and benign (0.74 +/- 0.15 nmol/mg) biopsies agreed with prior published measurements of snap-frozen biopsies. There was substantial fractional enrichment of [3-C-13]lactate (approximate to 70%) and [4-C-13]glutamate (approximate to 24%) in both low-grade cancer and benign biopsies. Although a significant difference in tissue [3-C-13]lactate fractional enrichment was not observed, lactate efflux was significantly higher (P < 0.05) in low-grade cancer biopsies (0.55 +/- 0.14 nmol/min/mg) versus benign biopsies (0.31 +/- 0.04 nmol/min/mg). ConclusionA protocol was developed for quantification of lactate production-efflux and TCA cycle activity in single living human prostate biopsies, allowing metabolic labeling on a wide spectrum of human tissues (e.g., metastatic, post-non-surgical therapy) from patients not receiving surgery.
Background Pyridoxine (VB6), which acts as a coenzyme in the biosynthesis of niacin, is formulated in pharmaceuticals to treat skin roughness. However, the mechanism of action of VB6 is not known precisely. Objective This study was conducted to clarify the influence of highly oxidative conditions on the expression of skin moisture-related mRNAs and to evaluate the preventive effects of VB6 focusing on antioxidant behaviour. Methods Intracellular levels of reactive oxygen species (ROS) in normal human epidermal keratinocytes (NHEKs) were determined using the 2 ',7 '-dichlorofluorescein diacetate assay. Real-time PCR was employed to investigate the influence of higher oxidative conditions on the expression of mRNAs encoding serine palmitoyl transferase (SPT) and filaggrin, and to characterize the mechanism of the antioxidant effect of VB6. Intracellular glutathione was quantified using an assay based on the glutathione recycling system with 5,5 '-dithiobis (2-nitrobenzoic acid) reagent and glutathione reductase. Carbonylated proteins (CPs) were semi-quantified by detecting aldehyde residues. Results Treatment of NHEKs with BSO increased the level of intracellular CPs by interfering with intracellular glutathione synthesis. Further, treatment with BSO down-regulated the expression level of SPT mRNA, but VB6 restored SPT mRNA expression in BSO-treated NHEKs. VB6 decreased the level of intracellular CPs with or without BSO treatment in a dose-dependent manner. In addition, VB6 increased levels of intracellular NADH/NADPH and glutathione through the activation of nuclear factor E2-related factor 2 (Nrf2) signalling. Conclusion These results suggest that highly oxidative conditions cause an impaired skin barrier function due to the down-regulation of SPT that results in skin roughness. VB6 improved the down-regulation of SPT mRNA expression initiated by highly oxidative conditions by enhancing the intracellular antioxidant system.
Dermal papillae ( DP ) play key roles in hair growth and regeneration by regulating follicular cell activity. Owing to the established roles of exosomes (Exos) in the regulation of cell functions, we investigated whether DP ‐derived Exos, especially those from three‐dimensional (3D)‐cultured DP cells, affect hair growth, cycling and regeneration. Exos derived from 3D DP (3D DP ‐Exos) promoted the proliferation of DP cells and outer root sheath ( ORS ) cells and increased the expression of growth factors ( IGF ‐1, KGF and HGF ) in DP cells. 3D DP ‐Exo treatment also increased hair shaft elongation in cultured human hair follicles. In addition, local injections of 3D DP ‐Exos induced anagen from telogen and also prolonged anagen in mice. Moreover, Exo treatment in human DP spheres augmented hair follicle neogenesis when the DP spheres were implanted with mouse epidermal cells. Similar results were obtained using Exos derived from 2D‐cultured DP cells (2D DP ‐Exo). Collectively, our data strongly suggest that Exos derived from DP cells promote hair growth and hair regeneration by regulating the activity of follicular dermal and epidermal cells; accordingly, these findings have implications for the development of therapeutic strategies for hair loss.
Cultured meat may be a novel food that would overcome the limits of conventional meat production. This paper assesses the willingness to try, buy and pay for cultured meat among a sample of Italian consumers, unveiling the attitudes towards an engineered food on the part of a consumer oriented in favour of the Mediterranean diet. A survey was conducted by submitting a questionnaire to 525 Italian consumers. Consumers showed higher agreement with the statements concerning positive externalities than the intrinsic characteristics of cultured meat, and more than half of the respondents (54%) stated that they would be willing to try cultured meat. The profile for a potential consumer of cultured meat was young, highly educated, somewhat familiar with cultured meat, a meat consumer and willing to reduce meat consumption. However, the survey findings may be biased by the unavailability of the product on the market and the information provided to the respondents focused on the potential benefits of cultured meat.
Objective Evaluating sexual function and quality of life (QoL) in patients treated with a modified Abbe-McIndoe technique using in vitro cultured autologous vaginal mucosa. Design Descriptive study. Setting Policlinico Umberto I, Sapienza University of Rome. Population From 2006 to 2016, 39 women affected by Mayer-Rokitansky-Kuster-Hauser syndrome (MRKHS) underwent vaginoplasty at our centre using a modified Abbe-McIndoe technique with in vitro cultured autologous vaginal tissue. Methods For each patient, vaginal tissue was obtained by full-thickness biopsy of the vaginal vestibule. Following enzymatic dissociation, cells were cultured for 2-3 weeks before the transplant. Main outcome measures Each patient completed two validated questionnaires to quantify sexual function and QoL: the Female Sexual Function Index (FSFI), administered at 12, 36, and 60 months, and the Psychological General Well Being Index (PGWBI) administered at 0, 6, and 36 months after surgery. Results Twelve months after surgery, 29 patients were engaging in regular sexual activity. The FSFI test results showed a satisfactory sexual function compared to the general population, with median values of 25.85 (range 4.6-30.5) at 12 months, 27.2 (range 4.4-33.6) at 36 months, and 29.6 (range 23.9-33.6) at 60 months. The PGWBI questionnaire showed a median score of 420.5 (range 108-540) before surgery, and 459 (range 252-533) at the 60-month follow-up. Conclusions Vaginoplasty performed with the use of autologous vaginal tissue, besides ensuring a long-term satisfying sex life, helps in achieving an improvement in QoL that is maintained over time.
n‐3 PUFAs decrease ASC‐mediated IL‐17 A secretion, through ICAM‐1 inhibition of expression . ASCs isolated from obese, but not lean adipose tissue (AT) are previously shown to promote Th17 cells, activate IL‐1β and IL‐6 secretion, and inhibit Th1 cells. Here, it is demonstrated that n‐3PUFAs inhibit ICAM‐1‐mediated cell interactions between ASCs and mononuclear cells, which results in inhibition of IL‐17A secretion.
3,3'-Diindolylmethane (DIM), a digestive metabolite originating from cruciferous vegetables, has dietary potential for the treatment of various human intestinal diseases. Although intestinal permeability dysfunction is closely related to the initiation and progression of human intestinal inflammatory diseases (IBDs), the effect of DIM on intestinal permeability is unclear. We evaluated the effect of DIM on the intestinal permeability of human intestinal cell monolayers and the animal model Caenorhabditis elegans, which were treated with IL-1 beta and Pseudomonas aeruginosa, respectively, to mimic IBD conditions. DIM substantially restored the intestinal permeability of differentiated Caco-2 cells by enhancing the expression of tight junction proteins (including occludin and ZO-1). Compared to the IL-1 beta single treatment (551.0 +/- 49.0 Omega.cm(2)), DIM (10 mu M) significantly increased the transepithelial electrical resistance (TEER) of Caco-2 cell monolayers (919.0 +/- 66.4 Omega.m(2), p < 0.001). DIM also ameliorated the impaired intestinal permeability and extended the lifespan of C. elegans fed P. aeruginosa. The mean lifespan of DIM-treated worms (10.8 +/- 1.3 days) was higher than that of control-treated worms (9.7 +/- 1.1 days, p < 0.01). Thus, DIM is a potential nutraceutical candidate for the treatment of leaky gut syndrome by improving intestinal permeability.
Since sturgeons do not show clear sexual dimorphism particularly when are small in size, attempts were made to determine the best methods to identify early sex in farmed beluga sturgeon ( Huso huso ). The present study describes the ultrasonography, small surgery and plasma steroid hormone methods to determine gender at 18‐month fish, which no research has been conducted yet into the fish at such small ages. Twenty one cultured beluga sturgeon's gonad were imaged using an ultrasonograph unit with a 9–13 MHz linear transducer. Overall accuracy of sex determination using ultrasonography was 80.95%. Plasma testosterone (T) levels were significantly higher in males than in females whereas 17α,20βOH‐P levels were significantly higher in females than in males. Testosterone (T) and 17α,20βOH‐P were not correlated with morphometric parameters (TL, SL, W, CF) in 18‐month beluga sturgeon. Results of this study indicated that sex could be identified by each of ultrasonography, small surgery and analysis of blood plasma in such a small size (18‐month). Although direct observation was more efficient than the other methods, ultrasonography was the simplest and cost‐effective tool in sturgeon's sex determination compared to other methods, and the least invasive.
Vibrio rotiferianus is an important marine pathogen of various aquatic organisms and can be found widely distributed in the marine environment. To further characterize this pathogen, the pathogenic properties and genome of V. rotiferianus SSVR1601 isolated from Sebastes schlegelii with skin ulcer were analysed. SSVR1601 was shown to be short rod‐shaped cell with a single polar flagellum. Different degrees of pathological changes in fish kidney, intestine, gills and liver were observed after SSVR1601 challenge. The SSVR1601 genome consists of two chromosomes and two plasmids with a total of 5,717,113 bp, 42.04%–44.93% GC content, 5,269 predicted CDSs, 134 tRNAs and 40 rRNAs. The common virulence factors including OMPs, haemolysin, flagellin, DNase, entF , algU , tcpI , acfB and rfaD were found in strain SSVR1601. Furthermore, factors responsible for iron uptake ( fur , fepC and ccmC ) and types II, IV and VI secretion systems were detected, which are likely responsible for the pathogenicity of SSVR1601. The antimicrobial resistance genes, bacA , tet34 and norM, were detected based on Antibiotic Resistance Genes Database. The phylogenetic analysis revealed SSVR1601 to be most closely related to V. rotiferianus strains CAIM577 and B64D1.