Association of genetic variants rs641153 (CFB), rs2230199 (C3), and rs1410996 (CFH) with age-related macular degeneration in a Brazilian population

This study aimed to investigate the association among genetic variants of the complement pathway CFB R32Q (rs641153), C3 R102G (rs2230199), and CFH (rs1410996) with age-related macular degeneration (AMD) in a sample of the Brazilian population. In a case-control study, 484 AMD patients were classified according to the clinical age-related maculopathy grading system (CARMS) and compared to 479 unrelated controls. The genetic variants rs1410996 of complement H (CFH), rs641153 of complement factor B (CFB), and rs2230199 of complement 3 (C3) were evaluated through polymerase chain reaction (PCR) and direct sequencing. The associations between single nucleotide polymorphisms (SNPs) and AMD, adjusted by age, were assessed by using logistic regression models. A statistically significant association was observed between AMD risk and rs2230199 variant with an OR of 2.01 (P  = 0.0002) for CG individuals compared to CC individuals. Regarding the comparison of advanced AMD versus the control group, the OR was 2.12 (P = 0.0036) for GG versus AA genotypes for rs1410996 variant. Similarly, the OR for rs2230199 polymorphism was 2.3034 (P  = 5.47^e-05) when comparing CG individuals to CC carriers. In contrast, the rs641153 variant showed a significant protective effect against advanced AMD for GA versus GG genotype (OR = 0.4406; P  = 0.0019). When comparing wet AMD versus controls, a significant association was detected for rs1410996 variant (OR = 2.16; P  = 0.0039) comparing carriers of the homozygous GG versus AA genotype, as well as in the comparisons of GG (OR = 3.0713; P  = 0.0046) and CG genotypes (OR = 2.2249; P  = 0.0002) versus CC genotype for rs2230199 variant, respectively. The rs641153 variant granted a significant protective effect against wet AMD for GA versus GG genotypes (OR = 0.4601; P  = 0.0044). Our study confirmed the risk association between rs2230199 and rs1410996 variants and AMD, and the protective role against AMD for rs641153 variant.     

Nitric oxide induces transient Ca2+ changes in endothelial cells independent of cGMP

Ca²⁺ changes induced by nitric oxide (NO^·) were investigated in cultured human endothelial cells. Sodium nitroprusside (SNP) (1–100 μmol/L) and S-Nitroso-N-acetylpenicillamine (SNAP) (100 μmol/L) were used as NO^· donors. The cytoplasmatic Ca²⁺ concentration was calculated using ratiometric FURA2 fluorescence measurements. Both NO^· donors caused transient oscillatory Ca²⁺ changes, which were not detectable in the presence of oxyhemoglobin (50 μmol/L). Digital ratio imaging revealed initiation sites within cells where Ca²⁺ increases started spreading, which indicates that nonuniformly distributed targets might be involved in these reactions. Calcium was released from intracellular stores as indicated by experiments performed in Ca²⁺-free buffer. L-type Ca²⁺-channel blocker diltiazem (100 μmol/L) was not able to block these responses. NO^·-induced Ca²⁺ release from intracellular stores caused capacitative Ca²⁺ entry. Both thapsigargin (1 μmol/L) and cyclopiazonic acid (10 μmol/L) inhibited the SNP response completely, whereas neither ryanodine (up to 100 μmol/L) nor dantrolene (100 μmol/L) was able to inhibit Ca²⁺ changes induced by SNP, indicating that primarily inositol 1,4,5-triphosphate (IP₃)-dependent stores are released upon stimulation with NO^·. A small inhibitory effect of ATP- and SNP-induced peak [Ca²⁺]_i increase was measured in the presence of both caffeine (20 mmol/L) and procaine (1 mmol/L). Evidence is presented that cGMP is not involved in NO^·-induced Ca²⁺ signals, as neither inhibitors of guanylate cyclase (methylene blue and LY (83583) nor cell permeant analogues of cGMP altered or simulated [Ca²⁺]_i changes. An inhibitor of cGMP-dependent protein kinase was also ineffective. We therefore propose that endothelial cells have specific targets proximal or at IP₃ receptors to induce Ca²⁺ changes in endothelial cells stimulated with NO^·. J. Cell. Physiol. 172:296–305, 1997. © 1997 Wiley-Liss, Inc.     

Evidence for conformationally different states of interleukin‐10: binding of a neutralizing antibody enhances accessibility of a hidden epitope

We present the mapping of two anti‐human interleukin‐10 (hIL‐10) antibodies (CB/RS/2 and CB/RS/11) which have been described as binding their antigen cooperatively. The epitopes were identified using hIL‐10‐derived overlapping peptide scans prepared by spot synthesis. To identify residues essential for binding within the two epitopes, each position was replaced by all other L ‐amino acids. The epitope‐derived peptides were further characterized with respect to antibody affinity and their inhibition of the antibody–hIL‐10 interaction. One antibody (CB/RS/11) binds to residues which are completely buried in the X‐ray structure of IL‐10. Accessibility of this hidden epitope is enhanced upon binding of the antibody CB/RS/2, which recognizes a discontinuous epitope located nearby. The recognition of the hidden CB/RS/11 epitope, as well as the cooperative binding behaviour of the two antibodies, provides evidence that IL‐10 can adopt a conformational state other than that observed in the crystal structure. Copyright © 1999 John Wiley & Sons, Ltd.     

Therapeutic Targeting of MLL Degradation Pathways in MLL-Rearranged Leukemia

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Cuticular wax composition of Salix varieties in relation to biomass productivity

The leaf cuticular waxes of six Salix clones (one Salix miyabeana, one Salix dasyclados, one Salix eriocephala, two Salix purpurea, and one interspecific hybrid of Salix eriocephala x interior) with different biomass productivities were characterized by gas chromatography-mass spectrometry. Total wax content ranged from 6.3 to 16.8 microg cm(-2), and two distinct patterns of wax were measured. The wax from leaves of S. dasyclados 'SV1' differed from all other clones and was dominated by fatty acids (42%), high concentrations of n-alkanes (25%) and n-alcohols (28%), with low n-aldehyde content (4%). All other clones produced cuticular wax dominated by n-alcohols (32-51%), particularly 1-hexacosanol, with fatty acids (14-37%) and n-aldehydes (19-26%) present in lower abundances. Clones of Salix grown under identical environmental conditions produce noticeably different amounts of cuticular wax. In contrast to previous studies of Salix, total wax content was independent of biomass productivity, measured as basal area, suggesting that wax production is not directly linked with woody biomass production by shrub willows under these site conditions.     

Effects of combined ozone and nitrogen deposition on the in situ properties of eleven key plant species of a subalpine pasture

Tropospheric O₃ and deposition of reactive N threaten the composition and function of natural and semi-natural vegetation even in remote regions. However, little is known about effects of these pollutants individually or in combination on plant species in alpine habitats. We analyzed 11 frequent plant species of a subalpine Geo-Montani-Nardetum pasture exposed at 2,000 m a.s.l. in the Swiss Alps during 3 years using a factorial free-air exposure system with three concentrations of O₃ and five rates of N application. The aim was to detect subtle effects on leaf chlorophyll and N concentrations, leaf weight, specific leaf area (SLA), and δ¹⁸O and δ¹³C as proxies for gas exchange. We expected that the species’ responsiveness to O₃ and N would be related to their functional traits and that N-induced changes in these traits would modify the species’ response to O₃ via increased growth and higher leaf conductance (g _s). Most species reacted to N supply with the accumulation of N and chlorophyll, but with no change in SLA, g _s, and growth, except Carex sempervirens which showed increased water use efficiency and leaf weight. Elevated O₃ reduced g _s in most species, but this was not related to a reduction in leaf weight, which was recorded in half of the species. Contrary to our expectation, the magnitude of the response to both O₃ and N was not related to species-specific traits such as SLA or g _s. No pronounced O₃ × N interactions were observed. In conclusion, since for most species neither N nor gas exchange limited growth, their short-term response to O₃ and N and to their combination was small. O₃ × N interactive effects are expected to be more pronounced in habitats where species are more responsive to N due to favorable growth conditions in terms of nutrient availability and temperature. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Genetic diversity and population structure in Malus sieversii, a wild progenitor species of domesticated apple

Malus sieversii (Lebed.) M. Roem. is a wild progenitor species of the domesticated apple. It is found across a mountainous region of central Asia and has been the focus of several collection expeditions by the USDA-ARS-National Plant Germplasm System. This study used microsatellite variation at seven loci to estimate diversity and differentiation within M. sieversii using several complimentary approaches. Multilocus genotypes were amplified from 949 individuals representing seedling trees from 88 half-sib families from eight M. sieversii populations collected in Kazakhstan. Apportioning of genetic variation was estimated at both the family and site level. Analyses using a hierarchical model to estimate F _st showed that differentiation among individual families is more than three times greater than differentiation among sites. In addition, average gene diversity and allelic richness varied significantly among sites. A rendering of a genetic network among all sites showed that differentiation is largely congruent with geographical location. In addition, nonhierarchical Bayesian assignment methods were used to infer genetic clusters across the collection area. We detected four genetic clusters in the data set. The quality of these assignments was evaluated over multiple Markov Chain Monte Carlo runs using both posterior likelihood and stability of the assignments. The spatial pattern of genetic assignments among the eight collection sites shows two broadly distributed and two narrowly distributed clusters. These data indicate that the southwestern collection sites are more admixed and more diverse than the northern sites.     

No difference in between-country variability in use of newly approved orphan and non- orphan medicinal products - a pilot study

Background

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease, which rapidly leads to chronic respiratory failure requiring mechanical ventilation. Currently, forced vital capacity (FVC) < 50% is considered as physiologic marker for admitting patients to Noninvasive Positive Pressure Ventilation (NPPV) intervention, although it has been recently shown the median survival of patients with baseline FVC < 75% much shorter than median survival of patients with baseline FVC > 75%, independently by any treatment.

Aim

To assess the role of NPPV in improving outcome of ALS, a retrospective analysis was performed to investigate 1 year survival of ALS patients with FVC < 75% and nocturnal respiratory insufficiency, treated with NPPV, compared to a well-matched population of ALS patients, who refused or was intolerant to NPPV.

Methods

We investigated seventy-two consecutive ALS patients who underwent pulmonary function test. Forty-four presented a FVC > 75% and served as control group. Twenty-eight patients presented a FVC < 75% and showed, at polysomnography analysis, nocturnal respiratory insufficiency, requiring NPPV; sixteen were treated with NPPV, while twelve refused or were intolerant.

Results

Increased survival rate at 1 year in patients with FVC < 75% treated with NPPV, as compared to those who refused or could not tolerate NPPV (p = 0.02), was observed. The median rate of decline in FVC% was slower in NPPV patients than in patients who did not use NPPV (95% CI: 0.72 to 1.85; p < 0.0001).

Conclusion

This report demonstrates that early treatment with NPPV prolongs survival and reduces decline of FVC% in ALS.     

A homozygous <Emphasis Type="Italic">RAB3GAP2</Emphasis> mutation causes Warburg Micro syndrome

Warburg Micro syndrome and Martsolf syndrome are clinically overlapping autosomal recessive conditions characterized by congenital cataracts, microphthalmia, postnatal microcephaly, and developmental delay. The neurodevelopmental and ophthalmological phenotype is more severe in Warburg Micro syndrome in which cerebral malformations and severe motor and mental retardation are common. While biallelic loss-of-function mutations in RAB3GAP1 are present in the majority of patients with Warburg Micro syndrome; a hypomorphic homozygous splicing mutation of RAB3GAP2 has been reported in a single family with Martsolf syndrome. Here, we report a novel homozygous RAB3GAP2 small in-frame deletion, c.499_507delTTCTACACT (p.Phe167_Thr169del) that causes Warburg Micro syndrome in a girl from a consanguineous Turkish family presenting with congenital cataracts, microphthalmia, absent visually evoked potentials, microcephaly, polymicrogyria, hypoplasia of the corpus callosum, and severe developmental delay. No RAB3GAP2 mutations were detected in ten additional unrelated patients with RAB3GAP1-negative Warburg Micro syndrome, consistent with further genetic heterogeneity. In conclusion, we provide evidence that RAB3GAP2 mutations are not specific to Martsolf syndrome. Rather, our findings suggest that loss-of-function mutations of RAB3GAP1 as well as functionally severe RAB3GAP2 mutations cause Warburg Micro syndrome while hypomorphic RAB3GAP2 mutations can result in the milder Martsolf phenotype. Thus, a phenotypic severity gradient may exist in the RAB3GAP-associated disease continuum (the “Warburg–Martsolf syndrome”) which is presumably determined by the mutant gene and the nature of the mutation.