Derivation of Human Corneal Keratocytes from ReLEx SMILE Lenticules for Cell Therapy and Tissue Engineering.

Fibroblasts isolated and expanded from ReLEx SMILE lenticules can be a source of human keratocytes. Since corneal keratocytes are quiescent cells, it is difficult to expand them in vitro in suitable numbers for clinical and experimental use. In the present study, this problem was solved by isolating and growing corneal fibroblasts (CFs) with a high proliferative potential and their reversion to keratocytes in a selective serum-free medium. Fibroblasts reversed into keratocytes (rCFs) had a dendritic morphology and ultrastructural signs of activation of protein synthesis and metabolism. The cultivation of CFs in a medium with 10% FCS and their reversion into keratocytes was not accompanied by the induction of myofibroblasts. After reversion, the cells spontaneously formed spheroids and expressed keratocan and lumican markers, but not mesenchymal ones. The rCFs had low proliferative and migratory activity, and their conditioned medium contained a low level of VEGF. CF reversion was not accompanied by a change with the levels of IGF-1, TNF-alpha, SDF-1a, and sICAM-1. In the present study, it has been demonstrated that fibroblasts from ReLEx SMILE lenticules reverse into keratocytes in serum-free KGM, maintaining the morphology and functional properties of primary keratocytes. These keratocytes have a potential for tissue engineering and cell therapy of various corneal pathologies.

Carotid Artery Disease in Subjects with Type 2 Diabetes: Risk Factors and Biomarkers.

Carotid atherosclerosis (CA) and, especially, carotid artery stenosis (CAS), are associated with a high risk of cardiovascular events in subjects with type 2 diabetes (T2D). In this study, we aimed to identify risk factors and biomarkers of subclinical CA and CAS in T2D individuals. High-resolution ultrasonography of carotid arteries was performed in 389 patients. Ninety-five clinical parameters were evaluated, including diabetic complications and comorbidities; antihyperglycemic, hypolipidemic, and antihypertensive therapy; indices of glycemic control and glucose variability (GV); lipid panels; estimated glomerular filtration rate (eGFR); albuminuria; blood cell count; and coagulation. Additionally, serum levels of calponin-1, relaxin, L-citrulline, and matrix metalloproteinase-2 and -3 (MMP-2, -3) were measured by ELISA. In univariate analysis, older age, male sex, diabetes duration, GV, diabetic retinopathy, chronic kidney disease, coronary artery disease, peripheral artery disease, and MMP-3 were associated with subclinical CA. In addition to these factors, long-term arterial hypertension, high daily insulin doses, eGFR, and L-citrulline were associated with CAS. In multivariate logistic regression, age, male sex, BMI, GV, and eGFR predicted CA independently; male sex, BMI, diabetes duration, eGFR, and L-citrulline were predictors of CAS. These results can be used to develop screening and prevention programs for CA and CAS in T2D subjects.

Circulating Wnt1-inducible signaling pathway protein-1 (WISP-1/CCN4) is a novel biomarker of adiposity in subjects with type 2 diabetes.

BACKGROUND: Wnt1-inducible signaling pathway protein 1, or cellular communication network factor 4 (CCN4), a member of CCN family of secreted, extracellular matrix associated signaling proteins, recently was validated as a novel adipose tissue derived cytokine. OBJECTIVE: To assess the relationships between circulating CCN4, adipose tissue distribution and function, and chronic low-grade inflammation in subjects with type 2 diabetes. METHODS: We observed 156 patients with type 2 diabetes and 24 healthy controls. Serum levels of CCN4, hsCRP and alpha1-acid glycoprotein (alpha1-AGP) were measured by ELISA. Serum concentrations of leptin, resistin, visfatin, adipsin, adiponectin, IL-6, IL-8, IL-18 and TNF-alpha were determined by multiplex analysis. Fat mass and distribution was assessed by DEXA. Mean diameter of adipocytes was estimated in samples of subcutaneous adipose tissue. RESULTS: Patients with diabetes had higher levels of circulating CCN4, leptin, resistin, adipsin, visfatin, hsCRP, alpha1-AGP, and IL-6 (all p < 0.02). The CCN4 concentration correlated positively with percentage of fat mass in central abdominal area, as well as with leptin, resistin and visfatin levels; negative correlation was found between CCN4 and mean adipocyte diameter. In multiple regression analysis fat mass in central abdominal area was independent predictor for CCN4 concentration. CONCLUSION: In subjects with type 2 diabetes serum levels of CCN4 are associated with central abdominal fat mass and adipose tissue dysfunction.

A comparative study of the hypolipidaemic effects of a new polysaccharide, mannan Candida albicans serotype A, and atorvastatin in mice with poloxamer 407-induced hyperlipidaemia.

OBJECTIVES: We evaluated the hypolipidaemic effect of mannan Candida albicans serotype A, relative to atorvastatin, in a mouse model of hyperlipidaemia. METHODS: Mannan serotype A was investigated in vitro and in vivo to determine its effects on macrophage proliferation, nitric oxide (NO) production by cultured macrophages, serum and liver lipids, changes in liver morphology and serum chitotriosidase activity and its expression in the liver. KEY FINDINGS: Mannan serotype A stimulates the macrophage proliferation and NO production in murine peritoneal macrophages in vitro. The activity of serum chitotriosidase (an enzyme released from the activated macrophages) was found to be significantly increased in P-407-induced hyperlipidaemic mice pretreated with low-dose mannan compared with mice administered P-407 only. Mannan treatment in mice was shown to significantly increase the chitotriosidase expression in the liver of both non-hyperlipidaemic and P-407-induced hyperlipidaemic mice. Lastly, mice pretreated with mannan before the induction of hyperlipidaemia with P-407 showed a significant reduction in the serum concentration of atherogenic LDL cholesterol, total cholesterol, triglycerides and liver triglycerides. CONCLUSIONS: It is suggested that mannan serotype A, like ß-glucan, may represent another hypolipidaemic agent, which could potentially be used as an adjunctive therapy with conventional antihyperlipidaemic drugs (statins and fibrates) in humans.