No abstract

Tear osmolarity can now be considered a test suitable to be performed in a clinical setting. It showed a good performance in dry eye diagnosis, higher than the other tests considered, mainly in severe dry eye. Tear osmolarity values should be interpreted as an indicator of DED evolutionary process to severity.

Purpose: To analyze tear protein variations in patients suffering from dry eye symptoms in the presence of tear film instability but without epithelial defects. Methods: Five microlitres of non-stimulated tears from 60 patients, suffering from evaporative dry eye (EDE) with a break-up time (BUT) o10 s, and from 30 healthy subjects as control (no symptoms, BUT 410 s) were collected. Tear proteins were separated by mono and bi-dimensional SDS-PAGE electrophoresis and characterized by immunoblotting and enzymatic digestion. Digested peptides were analyzed by liquid chromatography coupled to electrospray ionization quadrupole-time of flight mass spectrometry followed by comparative data analysis into Swiss-Prot human protein database using Mascot. Statistical analysis were performed by applying a t-test for independent data and a MannWhitney test for unpaired data (Po0.05). Results: In EDE patients vs controls, a significant decrease in levels of lactoferrin (data in % ± SD): 20.15 ± 2.64 vs 24.56 ± 3.46 (P ¼ 0.001), lipocalin-1: 14.98±2.70 vs 17.73±2.96 (P ¼ 0.0001), and lipophilin A-C: 2.89±1.06 vs 3.63±1.37 (P ¼ 0.006) was revealed, while a significant increase was observed for serum albumin: 9.45 ± 1.87 vs 3.46 ± 1.87 (P ¼ 0.0001). No changes for lysozyme and zinc a-2 glycoprotein (P ¼ 0.07 and 0.7, respectively) were shown. Proteomic analysis showed a downregulation of lipophilin A and C and lipocalin-1 in patients, which is suggested to be associated with post-translational modifications. Conclusions: Data show that tear protein changes anticipate the onset of more extensive clinical signs in early stage dry eye disease.

Viral-based techniques are the most efficient systems to deliver DNA into stem cells because they show high gene transduction and transgene expression in many cellular models. However, the use of viral vectors has several disadvantages mainly involving safety risks. Conversely, nonviral methods are rather inefficient for most primary cells. The Nucleofector technology, a new nonviral electroporation-based gene transfer technique, has proved to be an efficient tool for transfecting hard-to-transfect cell lines and primary cells. However, little is known about the capacity of this technique to transfect adult stem cells. In this study, we applied the Nucleofector technology to engineer human bone marrow-derived mesenchymal stem cells (hMSCs). Using a green fluorescent protein reporter vector, we demonstrated a high transgene expression level using U-23 and C-17 pulsing programs: 73.7% ؎ 2.9% and 42.5% ؎ 3.4%, respectively. Cell recoveries and viabilities were 38.7% ؎ 2.9%, 44.5% ؎ 3.9% and 91.4% ؎ 1.3%, 94.31% ؎ 0.9% for U-23 and C-17, respectively. Overall, the transfection efficiencies were 27.4% ؎ 2.9% (U-23) and 16.6% ؎ 1.4% (C-17) compared with 3.6% ؎ 2.4% and 5.4% ؎ 3.4% of other nonviral transfection systems, such as FUGENE6 and DOTAP, respectively (p < .005 for all comparisons). Nucleofection did not affect the immunophenotype of hMSCs, their normal differentiation potential, or ability to inhibit T-cell alloreactivity. Moreover, the interleukin-12 gene could be successfully transfected into hMSCs, and the immunomodulatory cytokine was produced in great amount for at least 3 weeks without impairment of its biological activity. In conclusion, nucleofection is an efficient nonviral transfection technique for hMSCs, which then may be used as cellular vehicles for the delivery of biological agents. STEM CELLS 2006;24:454 -461

Neurotrophic keratitis (NK) is a degenerative corneal disease caused by damage of trigeminal corneal innervation, which leads to spontaneous epithelial breakdown and corneal ulceration. The impairment of corneal sensory innervation causes the reduction of both protective reflexes and trophic neuromodulators that are essential for the vitality, metabolism, and wound healing of ocular surface tissues. A wide range of ocular and systemic conditions, including herpetic keratitis, ocular chemical burns, corneal surgery, diabetes, multiple sclerosis, and neurosurgical procedures, can cause NK by damaging trigeminal innervation. Diagnosis of NK requires careful investigation of any ocular and systemic condition associated with the disease, complete ocular surface examination, and quantitative measurement of corneal sensitivity. The clinical stages of NK range from corneal epithelial alterations (stage 1) to persistent epithelial defect (stage 2) and ulcer (stage 3), which may progress to corneal perforation. Management of NK is based on clinical severity, and the aim of the therapy is to halt the progression of corneal damage and promote epithelial healing. Although several medical and surgical treatments have been proposed, no therapies are currently available to restore corneal sensitivity, and thus, NK remains difficult and challenging to treat. The purpose of this review is to summarize available evidence on the pathogenesis, diagnosis, and treatment of NK. Novel medical and surgical therapies including the topical administration of nerve growth factor and corneal neurotization are also described.

Discomfort eye syndrome (DES) comprises a series of ‘minor’ subjective symptoms in patients where no relevant clinical signs are observed suggesting ocular disease. Our study includes 100 DES patients, excluding video terminal users, selected from the First Aid Service of our Department over two peak periods in both winter and summer time. The Schirmer test I, ferning test, breakup time and conjunctival cytology (scraping and imprint) were performed and data were related to sex, age and air pollution indexes, recorded in the patients’ living zones. Our results demonstrate that: (i) the ocular surface cytology and the analysis of tear film changes provide significant information in those patients where no other clinical signs are evident; (ii) DES symptoms are more frequent in women than in men (ratio about 2:1), both with ages over 51 years; (iii) DES is significantly associated with ocular surface inflammation, as detected by cytological methods, and (iv) ocular surface subclinical inflammation and ocular dryness are related to high concentrations of atmospheric polluters, in both sexes.

No abstract

Heterologous CBS-based eye drops represent a promising therapeutic approach in the healing of severely injured corneal epithelium and in subjective symptom relief. These drops can be obtained as readily available and quality-controlled blood derivative from cord blood banks on a routine basis.