The immunology of corneal limbal stem cells: the up-to-date approach to stem cell transplantation.

Limbal epithelial stem cells (LSC, LESC) are multipotent cells used as regenerative treatment of the cornea in patients with limbal epithelial stem cell deficiency (LSCD, LESCD). There are different types of stem cell grafting including cultivated limbal epithelial transplantation (CET) and simple limbal epithelial transplantation (SLET). The outcomes of the techniques have been assessed as similar, with differences in the sample size required during the procedures. The most important culture components for stem cell cultivation include 3T3 murine fibroblasts, human amniotic membrane (HAM), fibrin gel, and culture medium. The culture medium may be enriched with serum or not; however, xenobiotic-free materials are preferred because of the low risk of pathogen transmission. Multiple studies have defined molecules important for maintaining the function of LSC including C/EBP δ, Bmi-1, p63 α, interleukins (IL-6), epithelial structural proteins - keratins, and antibodies against epidermal growth factor receptor (EGFR). The cell phenotype of LSC has been described with factors of transplantation success rate such as a high percentage of p63 positive cells. The article emphasizes the role of recipient tissue preparation, modern cultivation techniques and pathophysiological processes in LSC transplantation effectiveness.

Retinal arterial remodeling in patients with pheochromocytoma or paraganglioma and its reversibility following surgical treatment.

OBJECTIVE: Structural abnormalities in resistance arteries are a hallmark of patients with hypertension. In hypertensive patients with pheochromocytoma or paraganglioma (PPGL), it is still a matter of debate whether structural vascular changes are because of elevated blood pressure (BP) or to toxic effects of elevated circulating catecholamines. Hence, the aim of our study was to assess whether catecholamine excess and/or elevated BP affect the structure of small retinal arteries in patients with catecholamine-producing tumors. METHODS: The study included 27 patients with PPGL and 27 hypertensive patients. All patients underwent biochemical tests for catecholamine excess, echocardiography and analyses of scanning-laser-Doppler-flowmetry (SLDF) both at baseline and 12 months following surgical resection of PPGL. RESULTS: Baseline retinal arterial diameter, arterial wall thickness and wall cross sectional area (WCSA) were higher in patients with PPGL as compared with subjects without PPGL (arterial diameter: 110 ±â€Š16.5 vs. 99.5 ±â€Š10.8 µm, wall thickness: 16.3 ±â€Š6.0 vs. 13.5 ±â€Š4.0 µm, WCSA: 4953.9 ±â€Š2472.8 vs. 3784.1 ±â€Š1446.3 µm, P < 0.05). Significant correlations were noted between wall thickness and WCSA and echocardiographic parameters assessing diastolic and systolic function of left ventricle. No correlations between retinal parameters, BP level and plasma concentrations of metanephrines were observed. In patients with PPGL, there were postoperative decreases in wall thickness (16.4 ±â€Š15.8 vs. 14.8 ±â€Š4.7 µm; P = 0.011) and WLR (0.42 ±â€Š0.13 vs. 0.37 ±â€Š0.10; P = 0.003) at 12 months after surgical removal of tumors. CONCLUSION: This is the first study to demonstrate that catecholamine excess is related to thickening of retinal arteries independent of BP and reversible after surgical cure. These data support a role of catecholamines in vascular remodeling in PPGL patients.

The relationship between HDAC6, CXCR3, and SIRT1 genes expression levels with progression of primary open-angle glaucoma.

BACKGROUND: Primary open-angle glaucoma (POAG) belongs to neurodegenerative diseases. Its etiology is not fully understood. However, a lot of reports have indicated that many biochemical molecules are involved in the retinal ganglion cell damage. Therefore, the purpose of this study was to evaluate a relationship between HDAC6, CXCR3, and SIRT1 genes expression levels with the occurrence risk of POAG and its progression. MATERIALS AND METHODS: The study included 34 glaucoma patients and 32 subjects without glaucoma symptoms. RNA was isolated from peripheral blood lymphocytes. Level of mRNA expression was determined by real-time PCR method. RESULTS: Our results have shown significant association of the HDAC6 and SIRT1 expression levels with progression of POAG according to rim area (RA) value, p = 0.041; p = 0.012. Moreover, the analysis of the CXCR3 expression level showed a correlation with progression of POAG based on RA and cup disc ratio (c/d) value, p = 0.006 and p = 0.012, respectively. CONCLUSIONS: The expression level of HDAC6, CXCR3, and SIRT1 genes may be involved in the progression of POAG.

The analysis of single nucleotide polymorphisms of the DGCR8 and XPO5 genes, and their association with the incidence of primary open angle glaucoma. / Analiza polimorfizmów pojedynczych nukleotydów genów DGCR8 i XPO5 oraz ich zwiazek z wystepowaniem jaskry pierwotnej otwartego kata.

Purpose: To analyse the single nucleotide polymorphisms of DGCR8 and XPO5 genes, involved in miRNA processing pathway, in relation to the incidence of primary open-angle glaucoma. Material and methods: Blood samples as the biological material used for the experiment were voluntarily donated by patients with known primary open-angle glaucoma and age-matched healthy controls. The two control groups ­ rs3757 DGCR8 and rs11077 XPO5 ­ consisted of 135 and 140 volunteers, respectively. The two study groups ­ rs3757 DGCR8 and rs11077 XPO5 ­ consisted of 137 and 138 subjects, respectively. The polymorphic variant frequencies of rs3757 and rs1107 were determined using DNA isolated from the peripheral blood lymphocytes in TaqMan® SNP Genotyping Assays. Results: The statistical analysis revealed that the genotype AG of DGCR8 rs3757 occurred more frequently in healthy individuals (P = 0.001), while homozygote GG was present mostly in people affected by primary open-angle glaucoma (P = 0.003). No association between the risk of primary open angle glaucoma and AC/CC genotypes of XPO5 was found. Conclusions: Many reports suggest the association between the miRNA alteration and the pathogenesis of glaucoma. The single nucleotide polymorphisms in DGCR8 and XPO5 genes, involved in microRNA biogenesis, may be the key factor in this process. Our experiment showed that genotype AG in rs3757 DGCR8 exhibits protective effect, decreasing the risk of primary open angle glaucoma, while the homozygote GG is probably associated with increased risk of glaucoma. The analysis of polymorphic variants of the genes involved in miRNA biogenesis could enable identification of glaucoma high-risk groups.

Altered Expression Levels of MMP1, MMP9, MMP12, TIMP1, and IL-1ß as a Risk Factor for the Elevated IOP and Optic Nerve Head Damage in the Primary Open-Angle Glaucoma Patients.

The aim of presented work was to analyze the impact of particular polymorphic changes in the promoter regions of the -1607 1G/2G MMP1, -1562 C/T MMP9, -82 A/G MMP12, -511 C/T IL-1ß, and 372 T/C TIMP1 genes on their expression level in POAG patients. Blood and aqueous humor samples acquired from 50 patients with POAG and 50 control subjects were used for QPCR and protein levels analysis by ELISA. In vivo promoter activity assays were carried on HTM cells using dual luciferase assay. All studied subjects underwent ophthalmic examination, including BCVA, intraocular pressure, slit-lamp examination, gonioscopy, HRT, and OCT scans. Patients with POAG are characterized by an increased mRNA expression of MMP1, MMP9, MMP12, and IL-1ß genes as compared to the control group (P < 0.001). Aqueous humor acquired from patients with POAG displayed increased protein expression of MMP1, MMP9, MMP12, and IL-1ß compared to the control group (P < 0.001). Allele -1607 1G of MMP1 gene possesses only 42,91% of the -1607 2G allele transcriptional activity and allele -1562 C of MMP9 gene possesses only 21,86% of the -1562 T allele. Increased expression levels of metalloproteinases can be considered as a risk factor for the development of POAG.

The role of base excision repair in the development of primary open angle glaucoma in the Polish population.

Glaucoma is a leading cause of irreversible blindness in developing countries. Previous data have shown that progressive loss of human TM cells may be connected with chronic exposure to oxidative stress. This hypothesis may suggest a role of the base excision repair (BER) pathway of oxidative DNA damage in primary open angle glaucoma (POAG) patients. The aim of our study was to evaluate an association of BER gene polymorphism with a risk of POAG. Moreover, an association of clinical parameters was examined including cup disk ratio (c/d), rim area (RA) and retinal nerve fiber layer (RNFL) with glaucoma progression according to BER gene polymorphisms. Our research included 412 patients with POAG and 454 healthy controls. Gene polymorphisms were analyzed by PCR-RFLP. Heidelberg Retinal Tomography (HRT) clinical parameters were also analyzed. The 399 Arg/Gln genotype of the XRCC1 gene (OR 1.38; 95% CI 1.02-1.89 p = 0.03) was associated with an increased risk of POAG occurrence. It was indicated that the 399 Gln/Gln XRCC1 genotype might increase the risk of POAG progression according to the c/d ratio (OR 1.67; 95% CI 1.07-2.61 P = 0.02) clinical parameter. Moreover, the association of VF factor with 148 Asp/Glu of APE1 genotype distribution and POAG progression (OR 2.25; 95% CI 1.30-3.89) was also found. Additionally, the analysis of the 324 Gln/His MUTYH polymorphism gene distribution in the patient group according to RNFL factor showed that it might decrease the progression of POAG (OR 0.47; 95% CI 0.30-0.82 P = 0.005). We suggest that the 399 Arg/Gln polymorphism of the XRCC1 gene may serve as a predictive risk factor of POAG.

Analysis of the expression and polymorphism of APOE, HSP, BDNF, and GRIN2B genes associated with the neurodegeneration process in the pathogenesis of primary open angle glaucoma.

Glaucoma is characterized by optic neuropathy of the RGC or retinal nerve fiber. The aim of this study was to evaluate a relationship between the neurodegenerative genes' polymorphisms of the APOE (rs449647), BDNF (rs2030324), GRIN2B (rs3764028), and HSP70-1 (rs1043618) and the occurrence risk of POAG and to investigate its effect on allele-specific gene expression. Genomic DNA was extracted from peripheral blood. Analysis of the genes' polymorphisms was performed using PCR-RFLP. The level of mRNA expression was determined by QRT-PCR. We showed a statistically significant association of BDNF and APOE genes' polymorphisms with a risk of POAG occurrence. There was a statistically significant association of the rs2030324 polymorphism with progression of POAG based on cup disc ratio value and rs1043618 polymorphism based on nerve fiber index and rim area. Furthermore, we found that mean HSP70-1 mRNA expression was significantly lower in the case of individuals with the G/G genotype than in the case of minor allele carriers, that is, G/C and C/C. We also found that BDNF and HSP70-1 expression level are associated with the progression of POAG based on rim area value. In conclusion, our results suggest that BDNF, APOE, and HSP70-1 genes might be associated with a risk of POAG occurrence in the Polish population.