A Possible Mechanism of Zika Virus Associated Microcephaly: Imperative Role of Retinoic Acid Response Element (RARE) Consensus Sequence Repeats in the Viral Genome.

Owing to the reports of microcephaly as a consistent outcome in the foetuses of pregnant women infected with ZIKV in Brazil, Zika virus (ZIKV) - microcephaly etiomechanistic relationship has recently been implicated. Researchers, however, are still struggling to establish an embryological basis for this interesting causal handcuff. The present study reveals robust evidence in favour of a plausible ZIKV-microcephaly cause-effect liaison. The rationale is based on: (1) sequence homology between ZIKV genome and the response element of an early neural tube developmental marker ‘retinoic acid’ in human DNA and (2) comprehensive similarities between the details of brain defects in ZIKV-microcephaly and retinoic acid embryopathy. Retinoic acid is considered as the earliest factor for regulating anteroposterior axis of neural tube and positioning of structures in developing brain through retinoic acid response elements (RARE) consensus sequence (5′–AGGTCA–3′) in promoter regions of retinoic acid-dependent genes. We screened genomic sequences of already reported virulent ZIKV strains (including those linked to microcephaly) and other viruses available in National Institute of Health genetic sequence database (GenBank) for the RARE consensus repeats and obtained results strongly bolstering our hypothesis that ZIKV strains associated with microcephaly may act through precipitation of dysregulation in retinoic acid-dependent genes by introducing extra stretches of RARE consensus sequence repeats in the genome of developing brain cells. Additional support to our hypothesis comes from our findings that screening of other viruses for RARE consensus sequence repeats is positive only for those known to display neurotropism and cause foetal brain defects (for which maternal-foetal transmission during developing stage may be required). The numbers of RARE sequence repeats appeared to match with the virulence of screened positive viruses. Although bioinformatic evidence and embryological features are in favour of our hypothesis, additional studies including animal models are warranted to validate our proposition. Such studies are likely to unfold ZIKV-microcephaly association and may help in devising methods to combat it

In vivo imaging of structural, metabolic and functional brain changes in glaucoma

Glaucoma, the world’s leading cause of irreversible blindness, is a condition for which elevated intraocular pressure is currently the only modifiable risk factor. However, the disorder can continue to progress even at reduced intraocular pressure. This indicates additional key factors that contribute to the etiopathogenesis. There has been a growing amount of literature suggesting glaucoma as a neurodegenerative disease of the visual system. However, it remains debatable whether the observed pathophysiological conditions are causes or consequences. This review summarizes recent in vivo imaging studies that helped advance the understanding of early glaucoma involvements and disease progression in the brains of humans and experimental animal models. In particular, we focused on the non-invasive detection of early structural and functional brain changes before substantial clinical visual field loss in glaucoma patients; the eye-brain interactions across disease severity; the metabolic changes occurring in the brain’s visual system in glaucoma; and, the widespread brain involvements beyond the visual pathway as well as the potential behavioral relevance. If the mechanisms of glaucomatous brain changes are reliably identified, novel neurotherapeutics that target parameters beyond intraocular pressure lowering can be the promise of the near future, which would lead to reduced prevalence of this irreversible but preventable disease

Advanced Diffusion MRI of the Visual System in Glaucoma: From Experimental Animal Models to Humans

Glaucoma is a group of ophthalmologic conditions characterized by progressive retinal ganglion cell death, optic nerve degeneration, and irreversible vision loss. While intraocular pressure is the only clinically modifiable risk factor, glaucoma may continue to progress at controlled intraocular pressure, indicating other major factors in contributing to the disease mechanisms. Recent studies demonstrated the feasibility of advanced diffusion magnetic resonance imaging (dMRI) in visualizing the microstructural integrity of the visual system, opening new possibilities for non-invasive characterization of glaucomatous brain changes for guiding earlier and targeted intervention besides intraocular pressure lowering. In this review, we discuss dMRI methods currently used in visual system investigations, focusing on the eye, optic nerve, optic tract, subcortical visual brain nuclei, optic radiations, and visual cortex. We evaluate how conventional diffusion tensor imaging, higher-order diffusion kurtosis imaging, and other extended dMRI techniques can assess the neuronal and glial integrity of the visual system in both humans and experimental animal models of glaucoma, among other optic neuropathies or neurodegenerative diseases. We also compare the pros and cons of these methods against other imaging modalities. A growing body of dMRI research indicates that this modality holds promise in characterizing early glaucomatous changes in the visual system, determining the disease severity, and identifying potential neurotherapeutic targets, offering more options to slow glaucoma progression and to reduce the prevalence of this world’s leading cause of irreversible but preventable blindness

A Novel Methodology for Enhanced and Consistent Heterologous Expression of Unmodified Human Cytochrome P450 1B1 (CYP1B1)

<div><p>Cytochrome P450 1B1 (CYP1B1) is a universal cancer marker and is implicated in many other disorders. Mutations in CYP1B1 are also associated with childhood blindness due to primary congenital glaucoma (PCG). To understand the CYP1B1 mediated etiopathology of PCG and pathomechanism of various cancers, it is important to carry out its functional studies. Heterologous expression of CYP1B1 in prokaryotes is imperative because bacteria yield a higher amount of heterologous proteins in lesser time and so the expressed protein is ideal for functional studies. In such expression system there is no interference by other eukaryotic proteins. But the story is not that simple as expression of heterologous CYP1B1 poses many technical difficulties. Investigators have employed various modifications/deletions of CYP N-terminus to improve CYP1B1 expression. However, the drawback of these studies is that it changes the original protein and, as a result, invalidates functional studies. The present study examines the role of various conditions and reagents in successful and consistent expression of sufficient quantities of unmodified/native human CYP1B1 in <i>E. coli</i>. We aimed at expressing CYP1B1 in various strains of <i>E. coli</i> and in the course developed a protocol that results in high expression of unmodified protein sufficient for functional/biophysical studies. We examined CYP1B1 expression with respect to different expression vectors, bacterial strains, types of culture media, time, Isopropyl β-D-1-thiogalactopyranoside concentrations, temperatures, rotations per minute, conditioning reagents and the efficacy of a newly described technique called double colony selection. We report a protocol that is simple, easy and can be carried out in any laboratory without the requirement of a fermentor. Though employed for CYP1B1 expression, this protocol can ideally be used to express any eukaryotic membrane protein.</p></div

Composition of the trace element solution and the various combinations of terrific broth used in the experimental setup.

<p>Composition of the trace element solution and the various combinations of terrific broth used in the experimental setup.</p