Aldose reductase inhibitor fidarestat counteracts diabetes-associated cataract formation, retinal oxidative-nitrosative stress, glial activation, and apoptosis

This study was aimed at evaluating the potent and specific aldose reductase inhibitor fidarestat, on diabetes-associated cataract formation, and retinal oxidative-nitrosative stress, glial activation, and apoptosis. Control and streptozotocin-diabetic rats were treated with or without fidarestat (16 mg kg(-1)d(-1)) for 10 weeks after an initial 2-week period without treatment. Lens changes were evaluated by indirect ophthalmoscopy and portable slit lamp. Nitrotyrosine, poly(ADP-ribose), and glial fibrillary acidic protein expression were assessed by immunohistochemistry. The rate of apoptosis was quantified in flat-mounted retinas by TUNEL assay with immunoperoxidase staining. To dissect the effects of high glucose exposure in retinal microvascular cells, primary bovine retinal pericytes and endothelial cells were cultured in 5 or 30 mM glucose, with or without fidarestat (10 microM) for 3-14 days. Apoptosis was assessed by TUNEL assay, nitrotyrosine and poly(ADP-ribose) by immunocytochemistry, and Bax and Bcl-2 expression by Western blot analyses. Fidarestat treatment prevented diabetic cataract formation and counteracted retinal nitrosative stress, and poly(ADP-ribose) polymerase activation, as well as glial activation. The number of TUNEL-positive nuclei (mean +/- SEM) was increased approximately 4-fold in diabetic rats vs. controls (207+/-33 vs. 49+/-4, p<0.01), and this increase was partially prevented by fidarestat (106+/-34, p<0.05 vs. untreated diabetic group). The apoptotic cell number increased with the prolongation of exposure of both pericytes and endothelial cells to high glucose levels. Fidarestat counteracted nitrotyrosine and poly(ADP-ribose) accumulation and apoptosis in both cell types. Antiapoptotic effect of fidarestat in high glucose-exposed retinal pericytes was not associated with the inhibition of Bax or increase in Bcl-2 expression. In conclusion, the findings, i) support an important role for aldose reductase in diabetes-associated cataract formation, and retinal oxidative-nitrosative stress, glial activation, and apoptosis, and ii) provide a rationale for the development of aldose reductase inhibitors, and, in particular, fidarestat, for the prevention and treatment of diabetic ocular complications.     

Thymic selection determines gammadelta T cell effector fate: antigen-naive cells make interleukin-17 and antigen-experienced cells make interferon gamma

gammadelta T cells uniquely contribute to host immune defense, but how this is accomplished remains unclear. Here, we analyzed the nonclassical major histocompatibility complex class I T10 and T22-specific gammadelta T cells in mice and found that encountering antigen in the thymus was neither required nor inhibitory for their development. But when triggered through the T cell receptor, ligand-naive lymphoid-gammadelta T cells produced IL-17, whereas ligand-experienced cells made IFN-gamma. Immediately after immunization, a large fraction of IL-17(+) gammadelta T cells were found in the draining lymph nodes days before the appearance of antigen-specific IL-17(+) *beta T cells. Thus, thymic selection determines the effector fate of gammadelta T cells rather than constrains their antigen specificities. The swift IL-17 response mounted by antigen-naive gammadelta T cells suggests a critical role for these cells at the onset of an acute inflammatory response to novel antigens.     

A thermoreversible hydrogel as a biosynthetic bandage for corneal wound repair

Ocular trauma and disorders that lead to corneal blindness account for over 2 million new cases of monocular blindness every year. A popular ocular surface reconstruction therapy, amniotic membrane transplantation, has been shown to aid corneal wound repair. However, the success rates of the procedure are variable. Here, we proposed to bioengineer a novel synthetic material that would serve as a biomimetic corneal bandage. The PLGA-PEG-PLGA triblock copolymer was synthesised via ring-opening polymerisation. Thermoreversible gelation behaviour was investigated at different polymer concentrations (23%, 30%, 35%, 40%, 45%, w/v) at temperatures ranging between 5 and 60 degrees C. Viscoelastic properties were studied in dynamic mechanical analysis with 1 degrees C/min temperature ramp. Cryo-SEM revealed a porous hydrogel with interconnecting networks. No adverse cytotoxicity was observed with an in vitro scratch-wound assay and in in vivo biocompatibility tests. We have demonstrated that the PLGA-PEG-PLGA hydrogel possessed a suitable gelling profile and, for the first time, the biocompatibility properties for this application as a potential bandage for corneal wound repair.     

Unrelated bone marrow transplantation with a reduced toxicity myeloablative conditioning regimen in Wiskott-Aldrich syndrome

Wiskott-Aldrich syndrome (WAS) is an X-linked congenital immune-deficiency syndrome, and bone marrow transplantation (BMT) has become a curative modality. However, the transplant with the alternative donor needed more intensive conditioning with increased treatment-related toxicities. Recently, fludarabine-based reduced toxicity myeloablative conditioning regimens have been developed for adult myeloid malignancies with promising results of good engraftment and low treatment-related toxicities. To increase the engraftment potential without serious complications, a boy with WAS received successful unrelated BMT with a reduced toxicity myeloablative conditioning regimen composed of fludarabine (40 mg/m2) on days -8, -7, -6, -5, -4, -3), busulfan (0.8 mg/kg i. v. q 6 hr on days -6, -5, -4, -3), and thymoglobulin (2.5 mg/kg on days -4, -3, -2). This novel conditioning regimen could improve the outcome of allogeneic transplantation for other non-malignant diseases such as congenital immune-deficiency syndromes or metabolic storage diseases.     

Mutation of Glu78 of the AVP-NPII gene impairs neurophysin as a carrier protein for arginine vasopressin in a family with neurohypophyseal diabetes insipidus

Familial neurohypophyseal diabetes insipidus (FNDI; OMIM 192340) is a rare inherited disorder with an autosomal dominant inheritance pattern. It is characterized by persistent polydipsia and polyuria induced by deficient or absent secretion of arginine vasopressin (AVP). We report a Korean kindred in whom FNDI is associated with a novel deletion mutation in exon 2 of the AVP-NPII gene encoding the neurophysin II moiety. An 18-yr-old man with polyuria and polydipsia was shown to have central diabetes insipidus by using the water deprivation test. Four family members were suspected to have symptomatic vasopressin-deficient diabetes insipidus. Direct sequencing of the AVP-NPII gene showed a heterozygous GAG deletion mutation in exon 2, which results in in-frame deletion of glutamic acid (c.232_234delGAG; p.Glu78del). The mutation was predicted to yield an abnormal AVP precursor lacking Glu78 (E78) in its neurophysin II moiety. Because Glu78 is essential for neurophysin II molecules to form a salt bridge with AVP, the function of neurophysin as a carrier protein for AVP would be impaired. The proband's mother and sister have the same mutation. Presence of this mutation suggests that the portion of the neurophysin peptide encoded by this sequence is important for the appropriate expression of vasopressin.     

Autosomal glycogenosis of liver and muscle due to phosphorylase kinase deficiency is caused by mutations in the phosphorylase kinase beta subunit (PHKB)

Glycogen storage disease due to phosphorylase kinase deficiency occurs in several variants that differ in mode of inheritance and tissue- specificity. This heterogeneity is suspected to be largely due to mutations affecting different subunits and isoforms of phosphorylase kinase. The gene of the ubiquitously expressed beta subunit, PHKB, was a candidate for involvement in autosomally transmitted phosphorylase kinase deficiency of liver and muscle. To identify such mutations, the complete PHKB coding sequence was amplified by RT-PCR of RNA isolated from blood samples of patients and analyzed by direct sequencing of PCR products. The characterization of mutations was complemented by PCR of genomic DNA. In one female and four male patients, we identified five independent nonsense mutations (Y418ter; R428ter; Y974H+E975ter; Q656ter in two cases), one single-base insertion in codon N421, one splice-site mutation affecting exon 31, and a large deletion involving the loss of exon 8. Although these severe translation-disrupting mutations occur in constitutively expressed sequences of the only known beta subunit gene of phosphorylase kinase, PHKB, they are associated with a surprisingly mild clinical phenotype, affecting virtually only the liver, and relatively high residual enzyme activity of approximately 10%.      

Paraoxonase Gene Polymorphism in South-western Korean Population

Paraoxonase (PON) has anti-atherogenic activity. Considering the important role of polymorphism in the genetic susceptibility to cardiovascular disease and the variability of its allele frequencies in different ethnic groups, the distribution of genotypes and allele frequencies of PON1M55L, PON1Q192R, PON2A148G, and PON2S311C polymorphisms was analyzed in a total 988 South-western Koreans and determined their effects on lipid parameters. The genotype distribution of PON1 at position 55 was LL=0.886, LM=0.114; and at position 192 was QQ=0.406, QR=0.594. The frequencies of the PON1 55L allele and the PON1 192R allele were similar to those seen in Chinese populations and Western populations, respectively. The genetic distribution of PON2 at position 148 was AA=0.619, AG=0.345, GG=0.035; and at position 311 was CC=0.035, SC=0.345, SS=0.619. The frequencies of the PON2 148G and 311S alleles were similar to those seen in Chinese populations. The concentrations of LDL and ApoB were significantly different between the PON2A148G (P<0.05) and PON2 S311C polymorphisms (P<0.01). PON polymorphisms and allele frequencies were described in Koreans living south-western part of Korea. These ethnic variations are considered important in the interpretation of diseases associated with PON polymorphisms.     

Comparison of the Prevalence of Chronic Obstructive Pulmonary Disease Diagnosed by Lower Limit of Normal and Fixed Ratio Criteria

The Global Initiative of Chronic Obstructive Lung Disease (GOLD) guidelines define chronic obstructive pulmonary disease (COPD) in subjects with FEV₁/FVC <0.7. However, the use of this fixed ratio may result in over-diagnosis of COPD in the elderly, especially with mild degree of COPD. The lower limit of normal (LLN) can be used to minimize the potential misclassification. The aim of this study was to evaluate the impact of different definitions of airflow obstruction (LLN or fixed ratio of FEV₁/FVC) on the estimated prevalence of COPD in a population-based sample. We compared the prevalence of COPD and its difference diagnosed by different methods using either fixed ratio (FEV₁/FVC <0.7) or LLN criterion (FEV₁/FVC below LLN). Among the 4,816 subjects who had performed spirometry, 2,728 subjects met new ATS/ERS spirometry criteria for acceptability and repeatability. The prevalence of COPD was 10.9% (14.7% in men, 7.2% in women) by LLN criterion and 15.5% (21.8% in men, 9.1% in women) by fixed ratio of FEV₁/FVC among subjects older than 45 yr. The difference of prevalence between LLN and fixed ratio of FEV₁/FVC was even higher among subjects with age ≥65, 14.9% and 31.1%, respectively. In conclusion, the prevalence of COPD by LLN criterion was significantly lower in elderly compared to fixed ratio of FEV₁/FVC. Implementing LLN criterion instead of fixed ratio of FEV₁/FVC may reduce the risk of over-diagnosis of COPD in elderly people.     

Association of Cytotoxic T Lymphocyte Antigen-4 Gene Polymorphisms and HLA Class II Alleles with the Development of Type 1 Diabetes in Korean Children and Adolescents

We studied the association of cytotoxic T lymphocyte antigen-4 gene (CTLA4) polymorphisms with the development of type 1 diabetes (T1D) in Korean children and adolescents. A total of 176 Korean subjects (92 females and 84 males) with childhood-onset T1D were studied. The A/G polymorphism at position 49 in CTLA4 exon 1 and the C/T polymorphism at position -318 in the CTLA4 promoter were analyzed by PCR-RFLP methods. The genotype and allele frequencies of the CTLA4 polymorphisms in the T1D patients were not different from those in the controls. These polymorphisms were not associated with the clinical characteristics or the development of autoimmune thyroid disease in the T1D patients. The frequency of the A allele was significantly higher in the patients that did not have two out of the three susceptible HLA-DRB1 alleles, which were DRB1*0301, *0405 and *09012, compared to the controls (P<0.05). These results suggest that CTLA4 polymorphisms do not directly confer any susceptibility to T1D. However, a CTLA4-mediated susceptibility effect on the development of T1D might be significant in children and adolescents that do not have susceptible HLA class II alleles.