[Impact of preservatives in topicals on the cutaneous microbiota]. / Einfluss von Konservierungsmitteln in Topika auf die kutane Mikrobiota.

Preservatives are used to stabilize topical preparations and protect the user from the influence of pathogenic microbes. After the application of a topical preparation, the matrix undergoes a metamorphosis, and by proportional evaporation of the hydrophilic phase the preservative may accumulate on the skin surface. This is believed to lead to antiseptic effects and may influence the diversity of the cutaneous microbiota. The regulation of the cutaneous microbiome and the associated influencing factors is a complex system that results in highly individualized conditions. Therefore, investigations on the influence of defined interventions are methodologically difficult. In the present proof-of-concept study, potential antiseptic effects of preservatives were investigated in a combination of in vitro and in vivo methods using microbiological culture tests. In addition, the investigations served to develop a clinical study design to answer further questions and use of an extended range of methods. The results support the hypothesis of an antiseptic effect of the tested preservatives (methyl-4-hydroxybenzoate and propyl-4-hydroxybenzoate, potassium sorbate and propylene glycol) on prominent reference bacteria, which could also be observed in clinical settings.

Isocitrate dehydrogenase 1 mutation drives leukemogenesis by PDGFRA activation due to insulator disruption in acute myeloid leukemia (AML).

Acute myeloid leukemia (AML) is characterized by complex molecular alterations and driver mutations. Elderly patients show increased frequencies of IDH mutations with high chemoresistance and relapse rates despite recent therapeutic advances. Besides being associated with global promoter hypermethylation, IDH1 mutation facilitated changes in 3D DNA-conformation by CTCF-anchor methylation and upregulated oncogene expression in glioma, correlating with poor prognosis. Here, we investigated the role of IDH1 p.R132H mutation in altering 3D DNA-architecture and subsequent oncogene activation in AML. Using public RNA-Seq data, we identified upregulation of tyrosine kinase PDGFRA in IDH1-mutant patients, correlating with poor prognosis. DNA methylation analysis identified CpG hypermethylation within a CTCF-anchor upstream of PDGFRA in IDH1-mutant patients. Increased PDGFRA expression, PDGFRA-CTCF methylation and decreased CTCF binding were confirmed in AML CRISPR cells with heterozygous IDH1 p.R132H mutation and upon exogenous 2-HG treatment. IDH1-mutant cells showed higher sensitivity to tyrosine kinase inhibitor dasatinib, which was supported by reduced blast count in a patient with refractory IDH1-mutant AML after dasatinib treatment. Our data illustrate that IDH1 p.R132H mutation leads to CTCF hypermethylation, disrupting DNA-looping and insulation of PDGFRA, resulting in PDGFRA upregulation in IDH1-mutant AML. Treatment with dasatinib may offer a novel treatment strategy for IDH1-mutant AML.

Contact Lens Fitting in Patients with Keratoconus - A Retrospective Assessment of 200 Patients. / Verlauf der Kontaktlinsenanpassung bei Keratokonus ­ eine retrospektive Untersuchung bei 200 Patienten.

BACKGROUND AND PURPOSE: Nowadays, keratoconus (KC) is very well treatable in a stage-oriented manner. A wide range of designs and materials of contact lenses (CL) are available for the treatment of KC. The aim of this study was to evaluate the possibilities, the possible challenges and the visual outcome of lens fitting in KC eyes. PATIENTS AND METHODS: This retrospective study includes data from 200 patients who received a lens fitting trial in our contact lens service between 2006 and 2016. We documented ophthalmological parameters, the type of prescribed CL, the number of required trial lenses and possible causes of the failure of the lens fitting. RESULTS: The mean age at initial lens fitting was 33.9 ± 12.5 years. In 98.8% of the cases, the fitting was performed with rigid gas permeable lenses, in 90.1% with four-curve lenses. Of the total number of aspheric lenses prescribed, 87.5% were fitted in keratoconus stages "1" to "2" (topographic keratoconus classification; Oculus Keratograph). Back surface toric lenses or bitoric lenses were fitted to 61.7% in keratoconus stages "2 - 3" to "4". Before patients received their final CL, a median of 2 trial lenses were required (max. 16). Mean visual acuity with lens correction was 0.8 ± 0.2 at the initial fitting, mean visual acuity with glasses correction was 0.5 ± 0.3. In 7.7% of the eyes, the KC lens fitting was discontinued due to the advanced stage of keratoconus, requiring a corneal transplant. Reasons for discontinuing contact lens fitting included lens intolerance (2.3%), application problems (0.3%) or acute corneal hydrops (0.3%). Discontinuation of lens wearing due to incompatibilities or application problems occurred in only four cases (1.1%) in the further course after lens fitting. CONCLUSIONS: The use of contact lenses is an integral part of the stage-appropriate therapy of keratoconus. Good visual acuity can be achieved in all stages of keratoconus with a low drop-out rate. In most cases, the adjustment is carried out with rigid gas permeable lenses with a four-curve geometry. In initial stages, aspherical lenses may be sufficient. Toric lenses can be fitted in advanced stages when rotationally symmetrical lenses cannot achieve a satisfactory fit. If contact lenses have been successfully fitted, there are only a few cases in which patients abandon their contact lenses because of intolerance.

The recombination mediator RAD51D promotes geminiviral infection.

To study a possible role for homologous recombination in geminivirus replication, we challenged Arabidopsis recombination gene knockouts by Euphorbia yellow mosaic virus infection. Our results show that the RAD51 paralog RAD51D, rather than RAD51 itself, promotes viral replication at early stages of infection. Blot hybridization analyses of replicative intermediates using one- and two-dimensional gels and deep sequencing point to an unexpected facet of recombination-dependent replication, the repair by single-strand annealing (SSA) during complementary strand replication. A significant decrease of both intramolecular, yielding defective DNAs and intermolecular recombinant molecules between the two geminiviral DNA components (A, B) were observed in the absence of RAD51D. By contrast, DNA A and B reacted differentially with the generation of inversions. A model to implicate single-strand annealing recombination in geminiviral recombination-dependent replication is proposed.

The contribution of translesion synthesis polymerases on geminiviral replication.

Geminiviruses multiply primarily in the plant phloem, but never in meristems. Their Rep protein can activate DNA synthesis in differentiated cells. However, when their single-stranded DNA is injected into the phloem by insects, no Rep is present for inducing initial complementary strand replication. Considering a contribution of translesion synthesis (TLS) polymerases in plants, four of them (Polη, Polζ, Polκ, Rev1) are highly and constitutively expressed in differentiated tissues like the phloem. Two geminiviruses (Euphorbia yellow mosaic virus, Cleome leaf crumple virus), inoculated either biolistically or by whiteflies, replicated in Arabidopsis thaliana mutant lines of these genes to the same extent as in wild type plants. Comparative deep sequencing of geminiviral DNAs, however, showed a high exchange rate (10(-4)-10(-3)) similar to the phylogenetic variation described before and a significant difference in nucleotide substation rates if Polη and Polζ were absent, with a differential response to the viral DNA components.

KU80, a key factor for non-homologous end-joining, retards geminivirus multiplication.

KU80 is well-known as a key component of the non-homologous end-joining pathway used to repair DNA double-strand breaks. In addition, the KU80-containing DNA-dependent protein kinase complex in mammals can act as a cytoplasmic sensor for viral DNA to activate innate immune response. We have now, to our knowledge for the first time, demonstrated that the speed of a systemic infection with a plant DNA geminivirus in Arabidopsis thaliana is KU80-dependent. The early emergence of Euphorbia yellow mosaic virus DNA was significantly increased in ku80 knockout mutants compared with wild-type sibling controls. The possible impact of KU80 on geminivirus multiplication by generating non-productive viral DNAs or its role as a pattern-recognition receptor against DNA virus infection is discussed.

Rad54 is not essential for any geminiviral replication mode in planta.

The circular single-stranded DNA of phytopathogenic geminiviruses is propagated by three modes: complementary strand replication (CSR), rolling circle replication (RCR) and recombination-dependent replication (RDR), which need host plant factors to be carried out. In addition to necessary host polymerases, proteins of the homologous recombination repair pathway may be considered essential, since geminiviruses are particularly prone to recombination. Among several others, Rad54 was suggested to be necessary for the RCR of Mungbean yellow mosaic India virus. This enzyme is a double-stranded DNA-dependent ATPase and chromatin remodeller and was found to bind and modulate the viral replication-initiator protein in vitro and in Saccharomyces cerevisiae. In contrast to the previous report, we scrutinized the requirement of Rad54 in planta for two distinct fully infectious geminiviruses with respect to the three replication modes. Euphorbia yellow mosaic virus and Cleome leaf crumple virus were inoculated into Rad54-deficient and wildtype Arabidopsis thaliana plant lines to compare the occurrence of viral DNA forms. Replication intermediates were displayed in the time course of infection by one and two-dimensional agarose gel electrophoresis and Southern hybridization. The experiments showed that Rad54 was neither essential for CSR, RCR nor RDR, and it had no significant influence on virus titers during systemic infection.