Papers by Stephen Richard
Methods and Protocols, 2025
Cobalt is a trace element, crucial for red blood cell formation and neurological function. Cobalt... more Cobalt is a trace element, crucial for red blood cell formation and neurological function. Cobalt toxicity is often only diagnosed after severe manifestations, including visual impairment. We aimed to investigate whether optical coherence tomography (OCT) and magnetic resonance imaging (MRI) can effectively detect cobalt-induced ocular toxicity in a murine model. Five wild-type mice (WT, C57Bl6) received daily intraperitoneal cobalt chloride injections for 28 days with a dosage of 12.5 mg/kg. Another 5 WT mice served as controls. After 28 days, all mice underwent manganese contrast-enhanced MRI and OCT examinations. Macroscopic and histological analysis of the enucleated eyes were performed. MRI revealed an increased signal in the optic nerves of injected mice. Anterion OCT provided in vivo visualization of the entire eye, demonstrating incipient cataract formation in the cobalt-injected mice. Both Spectralis domain OCT and Anterion, followed by histological analyses, confirmed preserved retinal structure with decreased thickness in the cobalt-injected group, with only minor neuronal damage and cell loss. Optic nerve analysis demonstrated myelin loss and increased inflammation with high levels of reactive gliosis. This study demonstrates optic neuropathy induced by cobalt toxicity, as shown by increased optic nerve signal on MRI without significant retinopathy. Anterion OCT showed incipient cataracts in the anterior segment.
Investigative Ophthalmology & Visual Science, 2024
PURPOSE. To explore the effect of cobalt toxicity on vision. METHODS. A total of 103 wild-type (W... more PURPOSE. To explore the effect of cobalt toxicity on vision. METHODS. A total of 103 wild-type (WT) mice were injected with cobalt chloride by two routes in different concentrations: single intravenous (IV) high or low doses (total, n = 43); or daily repeated intraperitoneal (IP) high (three days) or low (28 days, 56 days) dose, and low-dose cobalt with added minocycline (56 days) (total, n = 60); 10 WT mice served as a control group. An additional group of 17 immunodeficient NOD scid gamma (NSG) mice were injected IV or IP with cobalt, and 10 NSG mice served as control. Cobalt levels were measured in blood, urine, and tears by particle-induced X-ray emission (PIXE). Macroscopic, immunohistochemical, electroretinography (ERG), and molecular studies were done. RESULTS. PIXE revealed cobalt elimination from the blood by two hours, with increased levels in urine but under the detection limit in tears. In the retina, ERG recordings showed decreased b-wave amplitude. Apoptosis mainly involved the inner retina, with inner retinal inflammatory reaction in both WT and less in the NSG mice. In the optic nerves, an increased microglial and astrocytic activation was noted. CONCLUSIONS. This study demonstrated functional visual impairment with extensive inflammatory reaction secondary to cobalt toxicity in mice.
Cells, 2024
Cobalt toxicity is difficult to detect and therefore often underdiagnosed. The aim of this study ... more Cobalt toxicity is difficult to detect and therefore often underdiagnosed. The aim of this study was to explore the pathophysiology of cobalt-induced oxidative stress in the brain and its impact on structure and function. Thirty-five wild-type C57B16 mice received intraperitoneal cobalt chloride injections: a single high dose with evaluations at 24, 48, and 72 h (n = 5, each) or daily low doses for 28 (n = 5) or 56 days (n = 15). A part of the 56-day group also received minocycline (n = 5), while 10 mice served as controls. Behavioral changes were evaluated, and cobalt levels in tissues were measured with particle-induced X-ray emission. Brain sections underwent magnetic resonance imaging (MRI), electron microscopy, and histological, immunohistochemical, and molecular analyses. High-dose cobalt caused transient illness, whereas chronic daily low-dose administration led to longterm elevations in cobalt levels accompanied by brain inflammation. Significant neurodegeneration was evidenced by demyelination, increased blood-brain barrier permeability, and mitochondrial dysfunction. Treated mice exhibited extended latency periods in the Morris water maze test and heightened anxiety in the open field test. Minocycline partially mitigated brain injury. The observed signs of neurodegeneration were dose-and time-dependent. The neurotoxicity after acute exposure was reversible, but the neurological and functional changes following chronic cobalt administration were not.
Children, 2023
PurposeTo report the rate of ocular torsion in children with horizontal strabismus or orthophoria... more PurposeTo report the rate of ocular torsion in children with horizontal strabismus or orthophoria.Methods A retrospective study design was used. Nineteen children were included in the study, including 7 girls, aged 4-16 years. All patients were examined for strabismus and 12 were scheduled for surgical intervention. All participants had digital fundus photos (DRSplus, Padova, Italy) of both eyes at presentation, and 5 of 12 also had fundus photos following the strabismus operation. Patient files were reviewed for age, demographic data, type of strabismus, clinical symptoms and signs, orthoptic exams, subjective and objective reports of torsion, inferior oblique overaction, and V pattern. Fundus photos were analyzed for torsion by ImageJ software [ ImageJ 1.54f, National Institute of Health, USA]. The disc-foveal angle was calculated for ocular torsion. Discfoveal angle was defined as the angle formed between a line passing through the center of the optic disc to the fovea and another horizontal line passing through the center of the optic disc, using fundus photographs. Results Of the 19 children, 18 had horizontal strabismus, 9 with exotropia, and 9 with esotropia. One child was orthophoric with torsional strabismus. Inferior oblique overaction was detected in all but 3 children, while V pattern was documented in 10. Visual acuity was reduced (under 6/12) in 4 eyes of 4 children. None were symptomatic for ocular torsion. Although extorsion was documented clinically in 3 of 19 children, it was measurable on fundus photos in all patients before surgery with a mean of 8.7+8.5 degrees and 8.5+9.7 degrees in the right and left eyes, respectively. The mean extorsion in both eyes was 19.7+10.1 degrees and improved to a mean of 15.3+7.9 degrees in the children who were operated on and had documented postoperative fundus photographs.Conclusions Extorsion was detected on fundus photos at a significantly higher rate than in clinical examination. Notably, inferior oblique overaction was mainly associated with torsion. This study demonstrated that torsion is underdiagnosed in clinical examinations, as the children are often asymptomatic, but fundus photos which are easily obtained can improve its detection.
International Journal of Molecular Sciences
This study evaluated the potential neuroprotective effect of azithromycin (AZ) intraperitoneal in... more This study evaluated the potential neuroprotective effect of azithromycin (AZ) intraperitoneal injections in male C57Bl/6 (wild type, WT) and female NOD scid gamma (NSG) mice subjected to optic nerve crush (ONC) as a model for optic neuropathy. Histologically, reduced apoptosis and improved retinal ganglion cell (RGC) preservation were noted in the AZ-treated mice as shown by TUNEL staining—in the WT mice more than in the NSG mice. The increased microglial activation following ONC was reduced with the AZ treatment. In the molecular analysis of WT and NSG mice, similar trends were detected regarding apoptosis, as well as stress-related and inflammatory markers examining BCL2-associated X (Bax), heme oxygenase 1 (Ho-1), interleukin 1 beta (Il1β), superoxide dismutase 1 (Sod1), and nuclear factor-kappa B (Nfkb) levels. In the optic nerve, AZ increased the levels of expression of Sod1 and Nfkb only in the WT mice and decreased them in the NSG mice. In the retinas of the WT and NSG mice,...
Cell Reports, 2021
Rpb4/7 binds RNA polymerase II (RNA Pol II) transcripts co-transcriptionally and accompanies them... more Rpb4/7 binds RNA polymerase II (RNA Pol II) transcripts co-transcriptionally and accompanies them throughout their lives. By virtue of its capacity to interact with key regulators (e.g., RNA Pol II, eIF3, and Pat1) temporally and spatially, Rpb4/7 regulates the major stages of the mRNA life cycle. Here we show that Rpb4/7 can undergo more than 100 combinations of post-translational modifications (PTMs). Remarkably, the Rpb4/7 PTM repertoire changes as the mRNA/Rpb4/7 complex progresses from one stage to the next. These temporal PTMs regulate Rpb4 interactions with key regulators of gene expression that control transcriptional and post-transcriptional stages. Moreover, one mutant type specifically affects mRNA synthesis, whereas the other affects mRNA synthesis and decay; both types disrupt the balance between mRNA synthesis and decay (“mRNA buffering”) and the cell’s capacity to respond to the environment. We propose that temporal Rpb4/7 PTMs mediate the cross-talk among the various stages of the mRNA life cycle.
Future microbiology, Jan 13, 2017
The ssrA mutants were found to be more sensitive to mitomycin C (MMC) and our aim was to study th... more The ssrA mutants were found to be more sensitive to mitomycin C (MMC) and our aim was to study this phenomenon in detail. Strains were constructed by P1 transduction. pssrA(+) plasmid was constructed by PCR-based cloning and transformation was done by CaCl2 method. Relative viability analyses were done to assess the extent of viability of strains in relevant conditions. Gram staining was used for microscopic analysis. ssrA mutants become sensitive specifically to MMC, that too in a strain-specific manner. Precise tagging function of SsrA is necessary for conferring resistance to MMC. sulA::kan restored the viability of ssrA::cat mutants in a strain-specific manner. This study for the first time implicates SsrA in progression of efficient cell division and resistance to MMC.
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Papers by Stephen Richard