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Journal of Ophthalmic & Vision Research 2018To determine 1) which components of retinal function are impaired after rhegmatogenous retinal detachment, 2) which outer retinal pathways (rod- or cone-driven) are more...
PURPOSE
To determine 1) which components of retinal function are impaired after rhegmatogenous retinal detachment, 2) which outer retinal pathways (rod- or cone-driven) are more severely affected, and 3) whether there is concomitant inner retinal dysfunction.
METHODS
We conducted a prospective observational study in a large academic institution. We performed preoperative electroretinography on eight patients to assess outer and inner retinal function. In all cases, a comparison between the eye with the detached retina and the control fellow eye was made.
RESULTS
Eyes with a detached retina had significantly lower a-wave and b-wave amplitudes with respect to both rod- and cone-dominated testing parameters ( < 0.05) and reduced 30 Hz flicker responses compared to fellow eyes ( < 0.05); the effect size was similar for all significantly reduced parameters (r~0.6). There were no significant differences between eyes with detached retinas and control fellow eyes with respect to b/a-wave ratios, a-wave latencies, or b-wave latencies.
CONCLUSION
Patients with rhegmatogenous retinal detachment have preoperative outer retinal dysfunction equally affecting both rod- and cone-driven pathways, and they have minimal inner retinal dysfunction.
PubMed: 30090179
DOI: 10.4103/jovr.jovr_161_17 -
Journal of Molecular Biology Jan 2019Meters of DNA wrap around histone proteins to form nucleosomes and fit inside the micron-diameter nucleus. For the genetic information encoded in the DNA to become...
Meters of DNA wrap around histone proteins to form nucleosomes and fit inside the micron-diameter nucleus. For the genetic information encoded in the DNA to become available for transcription, replication, and repair, the DNA-histone assembly must be disrupted. Experiment has indicated that the outer stretches of nucleosomal DNA "breathe" by spontaneously detaching from and reattaching to the histone core. Here, we report direct observation of spontaneous DNA breathing in atomistic molecular dynamics simulations, detailing a microscopic mechanism of the DNA breathing process. According to our simulations, the outer stretches of nucleosomal DNA detach in discrete steps involving 5 or 10 base pairs, with the detachment process being orchestrated by the motion of several conserved histone residues. The inner stretches of nucleosomal DNA are found to be more stably associated with the histone core by more abundant nonspecific DNA-protein contacts, providing a microscopic interpretation of nucleosome unraveling experiments. The CG content of nucleosomal DNA is found to anticorrelate with the extent of unwrapping, supporting the possibility that AT-rich segments may signal the start of transcription by forming less stable nucleosomes.
Topics: DNA; Histones; Humans; Molecular Dynamics Simulation; Nucleic Acid Conformation; Nucleosomes
PubMed: 30468737
DOI: 10.1016/j.jmb.2018.11.013 -
Scientific Reports Dec 2022Adherent cell cultures are often dissociated from their culture vessel (and each other) through enzymatic harvesting, where the detachment response is monitored by an...
Adherent cell cultures are often dissociated from their culture vessel (and each other) through enzymatic harvesting, where the detachment response is monitored by an operator. However, this approach is lacking standardisation and reproducibility, and prolonged exposure or too high concentrations can affect the cell's viability and differentiation potential. Quantitative monitoring systems are required to characterise the cell detachment response and objectively determine the optimal time-point to inhibit the enzymatic reaction. State-of-the-art methodologies rely on bulky imaging systems and/or features (e.g. circularity) that lack robustness. In this study, lens-free imaging (LFI) technology was used to develop a novel cell detachment feature. Seven different donors were cultured and subsequently harvested with a (diluted) enzymatic harvesting solution after 3, 5 and 7 days of culture. Cell detachment was captured with the LFI set-up over a period of 20 min (every 20 s) and by optimising the reconstruction of the LFI intensity images, a new feature could be identified. Bright regions in the intensity image were identified as detaching cells and using image analysis, a method was developed to automatically extract this feature, defined as the percentage of detached cell regions. Next, the method was quantitatively and qualitatively validated on a diverse set of images. Average absolute error values of 1.49%, 1.34% and 1.97% were obtained for medium to high density and overconfluent cultures, respectively. The detachment response was quantified for all conditions and the optimal time for enzyme inhibition was reached when approximately 92.5% of the cells were detached. On average, inhibition times of 9.6-11.1 and 16.2-17.2 min were obtained for medium to high density and overconfluent cultures, respectively. In general, overconfluent cultures detached much slower, while their detachment rate was also decreased by the diluted harvesting solution. Moreover, several donors exhibited similar trends in cell detachment behaviour, with two clear outliers. Using the novel feature, measurements can be performed with an increased robustness, while the compact LFI design could pave the way for in situ monitoring in a variety of culture vessels, including bioreactors.
Topics: Reproducibility of Results; Cell Culture Techniques; Lens, Crystalline; Lenses; Diagnostic Imaging
PubMed: 36564377
DOI: 10.1038/s41598-022-22561-x -
Cornea Nov 2023The aim of this study was to explore types of Descemet membrane detachment (DMD) after ocular surface burns by anterior segment optical coherence tomography. (Observational Study)
Observational Study
PURPOSE
The aim of this study was to explore types of Descemet membrane detachment (DMD) after ocular surface burns by anterior segment optical coherence tomography.
METHODS
This is a pilot, case series, observational study. Patients with DMD after ocular surface burns were enrolled. Ophthalmologic examinations were performed in all patients including slit-lamp photography and anterior segment optical coherence tomography.
RESULTS
Three types of DMDs in 9 eyes of 9 patients with ocular surface burns were identified depending on the detachment components involved with the pre-Descemet layer (PDL). Type A was referred as a taut chord that the PDL and Descemet membrane (DM) detached simultaneously but were remained attached to each other, while type B was identified as a wavy line separated from the stroma by a dark slit that demonstrated the detachment of DM from the PDL and stroma. Type C was defined as the DM detached with or without PDL but they were separated from each other. We found that DM and PDL were detached simultaneously in most condition, with type A in 4 cases, type C in 5 cases, and type B in only 1 case.
CONCLUSIONS
Our study demonstrated 3 types of DMDs after ocular surface burns and revealed that the limbal involvement and retrocorneal exudations may give clues to DMD in the corresponding areas. DMDs may be neglected for long in patients with extensive limbal involvement in early stages and also play an important role in unstable ocular surface condition until the late stages of conjunctivalization after ocular surface burns.
Topics: Humans; Burns; Corneal Diseases; Descemet Membrane; Face; Tomography, Optical Coherence; Pilot Projects
PubMed: 36729715
DOI: 10.1097/ICO.0000000000003210 -
Indian Journal of Ophthalmology Nov 2023Aggressive retinopathy of prematurity (AROP) is a severe and progressive variant of retinopathy of prematurity (ROP) rapidly forming fibrous tissue extending from the...
INTRODUCTION
Aggressive retinopathy of prematurity (AROP) is a severe and progressive variant of retinopathy of prematurity (ROP) rapidly forming fibrous tissue extending from the disc toward the posterior lens surface progressing to Stage 5 disease without traversing the classical course that includes Stages 1 to 3. Since AROP behaves differently from type 1 ROP, this study was undertaken to evaluate the surgical outcome of AROP-related detachments.
METHODS
Retrospective analysis of data from electronic medical records of babies diagnosed with AROP-related detachments who underwent micro-incision vitrectomy surgery (MIVS) was included. The demographic data, details of primary intervention (laser and/or intravitreal bevacizumab), and surgery were noted. In a subset of patients, surgical intervention was planned early at the onset of fibrovascular tissue.
RESULTS
43 eyes of 26 babies with median birth weight 1175 g and median gestational age of 29 weeks were analyzed. 42/43 eyes underwent primary intervention in form of laser and/or anti-VEGF injection before surgery. 41.8%, 25.5%, and 32.5% eyes progressed to stages 4A, 4B, and 5, respectively, requiring surgical intervention. 66% eyes underwent lensectomy and vitrectomy (LV), and 44% eyes underwent lens sparring vitrectomy (LSV). 58% eyes had attached macula. 44% eyes that had a relatively less vascular diseases had better anatomical outcome (P = 0.019). At final follow-up, 53.4% eyes followed or at least had light fixation, and 77.7% eyes undergoing LSV fixated and/or followed light compared to 33% for LV (P = 0.04).
CONCLUSION
Challenges in AROP include rapid progression to advanced stages of ROP requiring close monitoring and multiple interventions. Surgeries for AROP have a favorable anatomical and functional outcome in 58% and 53%, respectively. Eyes undergoing lens sparing vitrectomy had better visual outcomes.
Topics: Infant, Newborn; Infant; Humans; Retinal Detachment; Retinopathy of Prematurity; Follow-Up Studies; Treatment Outcome; Retrospective Studies; Vitrectomy; Gestational Age
PubMed: 37870006
DOI: 10.4103/IJO.IJO_2999_22 -
Expert Review of Ophthalmology 2019Retinopathy of prematurity (ROP) is a leading cause of childhood blindness worldwide.
INTRODUCTION
Retinopathy of prematurity (ROP) is a leading cause of childhood blindness worldwide.
AREAS COVERED
Recent methods to identify and manage treatment-warranted vascularly active ROP are recognized and being compared to standard care by laser treatment in prospective large-scale clinical studies. Pharmacologic anti-angiogenic (anti-VEGF) treatment has changed the natural history of vascularly active ROP by reducing stage 3 intravitreal neovascularization and extending physiologic retinal vascularization in many infants. Tractional retinal detachments in stage 4 ROP after treatment with anti-VEGF agents show additional fibrovascular complexity compared to eyes treated with laser only. We review current management and outcomes for vascularly active and fibrovascular retinal detachment in ROP (stages 3, 4, 5 ROP), highlighting the evidence from recent clinical studies. Included are technical details important in surgery for retinal detachment in ROP. Literature searches were employed through PubMed.
EXPERT OPINION
Methods in pediatric imaging, safer pharmacologic treatments, and surgical techniques continue to advance to improve future ROP outcomes.
PubMed: 31762784
DOI: 10.1080/17469899.2019.1596026 -
Journal of Cell Science Apr 2021Rab5 and Rab7a are the main determinants of early and late endosomes and are important regulators of endosomal progression. The transport from early endosomes to late...
Rab5 and Rab7a are the main determinants of early and late endosomes and are important regulators of endosomal progression. The transport from early endosomes to late endosome seems to be regulated through an endosomal maturation switch, where Rab5 is gradually exchanged by Rab7a on the same endosome. Here, we provide new insight into the mechanism of endosomal maturation, for which we have discovered a stepwise Rab5 detachment, sequentially regulated by Rab7a. The initial detachment of Rab5 is Rab7a independent and demonstrates a diffusion-like first-phase exchange between the cytosol and the endosomal membrane, and a second phase, in which Rab5 converges into specific domains that detach as a Rab5 indigenous endosome. Consequently, we show that early endosomal maturation regulated through the Rab5-to-Rab7a switch induces the formation of new fully functional Rab5-positive early endosomes. Progression through stepwise early endosomal maturation regulates the direction of transport and, concomitantly, the homeostasis of early endosomes.
Topics: Endosomes; Transport Vesicles; rab5 GTP-Binding Proteins
PubMed: 33737317
DOI: 10.1242/jcs.254185 -
Biomolecules Mar 2021The aim of this study was to identify a relation between the clinical characteristics and differences in lipid peroxidation in the subretinal fluid (SRF) of...
The aim of this study was to identify a relation between the clinical characteristics and differences in lipid peroxidation in the subretinal fluid (SRF) of rhegmatogenous retinal detached patients by malondialdehyde (MDA) quantification. We collected 65 SRF samples from consecutive patients during scleral buckling surgery in rhegmatogenous retinal detachment (RRD) eyes. In addition to a complete ophthalmic evaluation, we studied the refractive status, evolution time, and the number of detached retinal quadrants to establish the extension of RRD. We studied the clinical aspects and oxidative stress and compared the characteristics among groups. We found that neither the evolution time of RRD nor the patients' age correlated with the MDA concentration in the SRF. The MDA and the protein content of the SRF increased in the patients with high myopia and with more extended RRD. Our results suggest that oxidative imbalance was important in more extended retinal detachment (RD) and in myopic eyes and should be taken into account in the managing of these cases.
Topics: Adult; Aged; Aged, 80 and over; Biomarkers; Female; Humans; Lipid Peroxidation; Male; Malondialdehyde; Middle Aged; Prognosis; Retinal Detachment; Subretinal Fluid
PubMed: 33808427
DOI: 10.3390/biom11040514 -
MSphere May 2024Biofilm formation is an important virulence factor for methicillin-resistant (MRSA). The extracellular matrix of MRSA biofilms contains significant amounts of...
Biofilm formation is an important virulence factor for methicillin-resistant (MRSA). The extracellular matrix of MRSA biofilms contains significant amounts of double-stranded DNA that hold the biofilm together. MRSA cells secrete micrococcal nuclease (Nuc1), which degrades double-stranded DNA. In this study, we used standard methodologies to investigate the role of Nuc1 in MRSA biofilm formation and dispersal. We quantified biofilm formation and extracellular DNA (eDNA) levels in broth and agar cultures. In some experiments, cultures were supplemented with sub-MIC amoxicillin to induce biofilm formation. Biofilm erosion was quantitated by culturing biofilms on rods and enumerating detached colony-forming units (CFUs), and biofilm sloughing was investigated by perfusing biofilms cultured in glass tubes with fresh broth and measuring the sizes of the detached cell aggregates. We found that an MRSA mutant strain produced significantly more biofilm and more eDNA than a wild-type strain, both in the absence and presence of sub-MIC amoxicillin. mutant biofilms grown on rods detached significantly less than wild-type biofilms. Detachment was restored by exogenous DNase or complementing the mutant. In the sloughing assay, mutant biofilms released cell aggregates that were significantly larger than those released by wild-type biofilms. Our results suggest that Nuc1 modulates biofilm formation, biofilm detachment, and the sizes of detached cell aggregates. These processes may play a role in the spread and subsequent survival of MRSA biofilms during biofilm-related infections.IMPORTANCEInfections caused by antibiotic-resistant bacteria known as methicillin-resistant (MRSA) are a significant problem in hospitals. MRSA forms adherent biofilms on implanted medical devices such as catheters and breathing tubes. Bacteria can detach from biofilms on these devices and spread to other parts of the body such as the blood or lungs, where they can cause life-threatening infections. In this article, researchers show that MRSA secretes an enzyme known as thermonuclease that causes bacteria to detach from the biofilm. This is important because understanding the mechanism by which MRSA detaches from biofilms could lead to the development of procedures to mitigate the problem.
Topics: Biofilms; Methicillin-Resistant Staphylococcus aureus; Micrococcal Nuclease; Anti-Bacterial Agents; Bacterial Proteins; DNA, Bacterial; Virulence Factors; Microbial Sensitivity Tests; Amoxicillin
PubMed: 38695568
DOI: 10.1128/msphere.00126-24 -
The Journal of Nutritional Biochemistry Nov 2022An emerging hallmark of cancer is cellular metabolic reprogramming to adapt to varying cellular environments. Throughout the process of metastasis cancer cells gain...
An emerging hallmark of cancer is cellular metabolic reprogramming to adapt to varying cellular environments. Throughout the process of metastasis cancer cells gain anchorage independence which confers survival characteristics when detached from the extracellular matrix (ECM). Previous work demonstrates that the bioactive metabolite of vitamin D, 1α,25-dihydroxyvitamin D (1,25[OH]D), suppresses cancer progression, potentially by suppressing the ability of cells to metabolically adapt to varying cellular environments such as ECM detachment. The purpose of the present study was to determine the mechanistic bases of the effects of 1,25(OH)D on cell survival in ECM-detached conditions. Pretreatment of MCF10A-ras breast cancer cells for 3 d with 1,25(OH)D reduced the viability of cells in subsequent detached conditions by 11%. Enrichment of C-glutamine was reduced in glutamate (21%), malate (30%), and aspartate (23%) in detached compared to attached MCF10A-ras cells. Pretreatment with 1,25(OH)D further reduced glutamine flux into downstream metabolites glutamate (5%), malate (6%), and aspartate (10%) compared to detached vehicle treated cells. Compared to attached cells, detachment increased pyruvate carboxylase (PC) mRNA abundance and protein expression by 95% and 190%, respectively. Consistent with these results, C-glucose derived M+3 labelling was shown to preferentially replenish malate and aspartate, but not citrate pools, demonstrating increased PC activity in detached cells. In contrast, 1,25(OH)D pretreatment of detached cells reduced PC mRNA abundance and protein expression by 63% and 56%, respectively, and reduced PC activity as determined by decreased C-glucose derived M+3 labeling in citrate (8%) and aspartate (50%) pools, relative to vehicle-treated detached cells. While depletion of PC with doxycycline-inducible shRNA reduced detached cell viability, PC knockdown in combination with 1,25(OH)D treatment did not additionally affect the viability of detached cells. Further, PC overexpression improved detached cell viability, and inhibited the effect of 1,25(OH)D on detached cell survival, suggesting that 1,25(OH)D mediates its effects in detachment through regulation of PC expression. These results suggest that inhibition of PC by 1,25(OH)D suppresses cancer cell anchorage independence.
Topics: Aspartic Acid; Cell Survival; Doxycycline; Extracellular Matrix; Glucose; Glutamic Acid; Glutamine; Malates; Pyruvate Carboxylase; RNA, Messenger; RNA, Small Interfering; Vitamin D
PubMed: 35934270
DOI: 10.1016/j.jnutbio.2022.109116