Differentiation of Human Embryonic Stem Cells into Cells with Corneal Keratocyte Phenotype

Corneal transparency depends on a unique extracellular matrix secreted by stromal keratocytes, mesenchymal cells of neural crest lineage. Derivation of keratocytes from human embryonic stem (hES) cells could elucidate the keratocyte developmental pathway and open a potential for cell-based therapy for corneal blindness. This study seeks to identify conditions inducing differentiation of pluripotent hES cells to the keratocyte lineage. Neural differentiation of hES cell line WA01(H1) was induced by co-culture with mouse PA6 fibroblasts. After 6 days of co-culture, hES cells expressing cell-surface NGFR protein (CD271, p75NTR) were isolated by immunoaffinity adsorption, and cultured as a monolayer for one week. Keratocyte phenotype was induced by substratum-independent pellet culture in serum-free medium containing ascorbate. Gene expression, examined by quantitative RT-PCR, found hES cells co-cultured with PA6 cells for 6 days to upregulate expression of neural crest genes including NGFR, SNAI1, NTRK3, SOX9, and MSX1. Isolated NGFR-expressing cells were free of PA6 feeder cells. After expansion as a monolayer, mRNAs typifying adult stromal stem cells were detected, including BMI1, KIT, NES, NOTCH1, and SIX2. When these cells were cultured as substratum-free pellets keratocyte markers AQP1, B3GNT7, PTDGS, and ALDH3A1 were upregulated. mRNA for keratocan (KERA), a cornea-specific proteoglycan, was upregulated more than 10,000 fold. Culture medium from pellets contained high molecular weight keratocan modified with keratan sulfate, a unique molecular component of corneal stroma. These results show hES cells can be induced to differentiate into keratocytes in vitro. Pluripotent stem cells, therefore, may provide a renewable source of material for development of treatment of corneal stromal opacities. © 2013 Chan et al

Atomic force microscopy techniques for nanomechanical characterization : a polymer case study

Atomic force microscopy (AFM) is a versatile tool to perform mechanical characterization of surface samples at the nanoscale. In this work, we review two of such methods, namely contact resonance AFM (CR-AFM) and torsional harmonics AFM (TH-AFM). First, such techniques are illustrated and their applicability on materials with elastic moduli in different ranges are discussed, together with their main advantages and limitations. Then, a case study is presented in which we report the mechanical characterization using both CR-AFM and TH-AFM of polyaniline and polyaniniline doped with nanodiamond particles tablets prepared by a pressing process. We determined the indentation modulus values of their surfaces, which were found in fairly good agreement, thus demonstrating the accuracy of the techniques. Finally, the determined surface elastic moduli have been compared with the bulk ones measured through standard indentation testing. INTRODUCTION In the field of nanotechnology, the development of innovative and nondestructive characterization techniques plays a crucial role. Indeed, the characterization of nanostructured hybrid materials (e.g., thin films and nanocomposites) and devices requires the capability of acquiring maps of the local mechanical properties at the nanoscale. Nanoindentation is the most common method for determining the mechanical properties of thin films. However, its applicability is strictly limited by the thickness of the sample. Furthermore, its poor spatial resolution does not allow the reconstruction of an accurate distribution of the sample surface mechanical properties. For this reason, alternative methods, based on atomic force microscopy (AFM), have been developed. By exploiting the high resolution of the AFM, maps of the surface mechanical properties (i.e., indentation modulus) can be achieved. Among these techniques, AFM nanoindentation1 is the simplest method used to evaluate the local mechanical properties o

Outcomes of pregnancies after kidney transplantation: lessons learned from CKD. A comparison of transplanted, nontransplanted chronic kidney disease patients and low-risk pregnancies: a multicenter nationwide analysis.

BACKGROUND: Kidney transplantation (KT) may restore fertility in CKD. The reasons why materno-foetal outcomes are still inferior to the overall population are only partially known. Comparison with the CKD population may offer some useful insights for management and counselling.Aim of this study was to analyse the outcomes of pregnancy after KT, compared with a large population of non-transplanted CKD patients and with low-risk control pregnancies, observed in Italy the new millennium. METHODS: We selected 121 live-born singletons after KT (Italian study group of kidney in pregnancy, national coverage about 75%), 610 live-born singletons in CKD and 1418 low-risk controls recruited in 2 large Italian Units, in the same period (2000-2014). The following outcomes were considered: maternal and foetal death; malformations; preterm delivery; small for gestational age baby (SGA); need for the neonatal intensive care unit (NICU); doubling of serum creatinine or increase in CKD stage. Data were analysed according to kidney diseases, renal function (staging according to CKD-EPI), hypertension, maternal age, partity, ethnicity. RESULTS: Materno-foetal outcomes are less favourable in CKD and KT as compared with the low-risk population. CKD stage and hypertension are important determinants of results. KT patients with e-GFR >90 have worse outcomes compared with CKD stage 1 patients; the differences level off when only CKD patients affected by glomerulonephritis or systemic diseases ('progressive CKD') are compared with KT. In the multivariate analysis, risk for preterm and early-preterm delivery was linked to CKD stage (2-5 versus 1: RR 3.42 and 3.78) and hypertension (RR 3.68 and 3.16) while no difference was associated with being a KT or a CKD patient. CONCLUSIONS: The materno-foetal outcomes in patients with kidney transplantation are comparable with those of nontransplanted CKD patients with similar levels of kidney function impairment and progressive and/or immunologic kidney diseas

The evolutionary significance of polyploidy

Polyploidy, or the duplication of entire genomes, has been observed in prokaryotic and eukaryotic organisms, and in somatic and germ cells. The consequences of polyploidization are complex and variable, and they differ greatly between systems (clonal or non-clonal) and species, but the process has often been considered to be an evolutionary 'dead end'. Here, we review the accumulating evidence that correlates polyploidization with environmental change or stress, and that has led to an increased recognition of its short-term adaptive potential. In addition, we discuss how, once polyploidy has been established, the unique retention profile of duplicated genes following whole-genome duplication might explain key longer-term evolutionary transitions and a general increase in biological complexity

Time to surgery and rehabilitation resources affect outcomes in orthogeriatric units

Objective: To compare the pathways of care and clinical results for patients admitted for hip fracture (HF) in 3 orthogeriatric co-managed care centers in order to estimate the effect of system factors on mortality and functional outcome. Design: Prospective inception multicenter cohort study. Setting: Three tertiary Hospitals. Participants: 806 patients consecutively admitted with HF. Measurements: 1-Year mortality, the loss of 1 or more functional abilities in activities of daily living (ADLs), and the recovery/maintenance of independent ambulation at 6 months from the fracture. Results: On the whole sample, 71.1% of patients survived 1 year from the fracture. In one hospital the risk of 1-year mortality was significantly higher even after adjusting for age, sex, comorbidity, prefracture functional status and cognitive impairment (odd ratio (OR) 1.56, 95% confidence interval (CI) 1.15-2.18, p = 0.01). This was principally explained by a longer time to surgery (5.2 days +/- 3.2 vs 2.7 +/- 2.3 and 2.7 +/- 2.2, p < 0.001). The three hospitals also differed in the rate of subjects losing the ability in ADLs after 6 months from the fracture (54.2%, 61%, 43.5%, p = 0.016), while no statistical differences were found in the recovery of independent ambulation. On the basis of multivariate models, a lower access to post-acute rehabilitation could account for lower outcome in functional status. Conclusions: This study suggests that system factors such as time to surgery and rehabilitation resources can affect functional recovery and 1-year mortality in orthogeriatric units and they could explain different outcomes when comparing care models. (C) 2011 Elsevier Ireland Ltd. All rights reserved

Older people with hip fracture and IADL disability require earlier surgery

Background: Hip fractures represent a major challenge for physicians as well as society as a whole. Both poor functional status and delay to surgery are well known risk factors for negative outcomes. We hypothesized that the timing of the operation is more important for frail older people than older people without functional limitations before fracture. Methods: We performed a prospective multicenter cohort study on 806 consecutive patients, 75 years of age or older, admitted with a fragility hip fracture to three hospitals in the Emilia-Romagna Region (Italy). All three hospitals had a comanaged care model, and the patients were under the shared responsibility of an orthopedic surgeon and a geriatrician. Results: Functional status assessed as instrumental activities of daily living was an important predictor of survival after 1 year from fracture. After adjusting for confounders, the hazard ratios per 1 point score of increase from 0 to 8 was 1.30 (95% confidence interval 1.19-1.42, p =. 000). Time to surgery increased 1-year mortality in patients with a low instrumental activities of daily living score (hazard ratios per day of surgical delay 1.14, 95% confidence interval 1.06-1.22, p &lt;. 001) and intermediate instrumental activities of daily living score (hazard ratios 1.21, 95% confidence interval 1.09-1.34, p &lt;. 001) but was an insignificant risk factor in functionally independent patients (hazard ratios 1.05 95% confidence interval 0.79-1.41, p =. 706). Conclusions: Surgery delay is an independent factor for mortality in older patients after hip fracture but only for the frail older people with prefracture functional impairment. If our results are confirmed, a more intensive approach should be adopted for older people with hip fractures who have disabilities. © 2012 The Author