blue-light-inducible-semi-random-mutagenesis-on-specific-exogenous

Blue Light-inducible Semi-random Mutagenesis on Specific Exogenous DNA in <em>Escherichia coli</em> BL21 Strain

Mutagenesis technologies have been widely used in variation breeding, directed evolution, and protein engineering. However, most methods can only induce ...

Luoyang Central Hospital Affiliated to Zhengzhou University

Nanog, a unique homeobox transcription factor, maintains self-renewal and pluripotency of embryonic stem cells by binding to nuclear factor κB proteins in order to inhibit their transcriptional and prodifferentiation activities. We previously reported that Nanog attenuated inflammatory responses in rat primary microglia cells stimulated by lipopolysaccharide. However, the effects of Nanog on another microglia cell type, BV-2 cells, are still unknown. In this study, we investigated whether Nanog attenuated inflammatory responses in lipopolysaccharide-stimulated BV-2 cells and found that Nanog significantly decreased the release of nitric oxide and the expression of inducible nitric oxide synthase at the mRNA and protein levels. The production of proinflammatory cytokines including tumor necrosis factor-α and interleukin-1β was also significantly inhibited by Nanog. Further, we observed that the transcriptional activity of nuclear factor κB was dramatically reduced by Nanog. These results suggest that Nanog may be a potential anti-inflammatory therapy for neurological diseases caused by persistent microglia activation.

Nanog attenuates lipopolysaccharide-induced inflammatory responses by blocking nuclear factor-κB transcriptional activity in BV-2 cells.

MicroRNAs mediates gene expression in various diseases. Studies have shown that aberrant expression of miRNAs affected cerebral protection. In this study, we have investigated the effects of microRNA-708 (miR-708) on cell survival of oxygen and glucose-deprived reoxygenation (OGD/R) human neuroblastoma cells (SH-SY5Y) and explored whether miR-708 inhibited neuronal death by targeting JAK1. In vitro model of ischemia was used to investigate the neuroprotective functions of miR-708. MiR-708 mimics/siJAK1 transfected SH-SY5Y cells were treated with OGD. After 48h of reoxygenation, cell viability and cell survival were determined by EdU and FITC/PI double staining flow cytometry, respectively. Luciferase activity assay was performed to validate the role of JAK1 as a direct target of miR-708. qRT-PCR and Immunofluorescence assays were used to determine the expression of JAK1, MAP2 and NEUN in miR-708 mimics transfected SH-SY5Y cells. To explore the mechanisms involved in cell growth promotion by JAK1, morphological changes in cells were detected upon knockdown of JAK1, and the expression levels of JAK1, Bax, Bcl-2, cleaved-caspase-3, STAT3 and Mcl-1 were determined by Western blotting. The expression of miR-708 significantly decreased in cells treated with OGD/R. MiR-708 directly targeted JAK1 3'UTR to down-regulate JAK1 mRNA expression, whereas the expression of MAP2 and NEUN was upregulated. Previous studies have demonstrated that the suppression of JAK1 inhibited apoptosis phenocopied function of the miR-708 overexpression in OGD/R SH-SY5Y cells. miR-708 decreased the rate of apoptosis of OGD/R SH-SY5Y cells by suppressing the expression of JAK1.

Inhibition of JAK1 by microRNA-708 promotes SH-SY5Y neuronal cell survival after oxygen and glucose deprivation and reoxygenation.

Cerebral amyloid angiopathy-related inflammation (CAA-ri) is a rare clinical entity, characterized by headaches, seizures, rapidly progressive cognitive decline, behavioral changes and magnetic resonance imaging (MRI) findings underlying the autoimmune and inflammatory reaction at the level of CAA-affected vessel. CAA-ri is likely responsive to corticosteroid. MRI shows asymmetric and multifocal white matter hyperintensity (WMH) lesions and multiple cerebral microbleeds. Apolipoprotein E (ApoE) ε4 homozygosity is associated with CAA-ri strongly [Neurology 68(17):1411-1416, 2007, Ann Neurol 73(4):449-458, 2013, J Alzheimers Dis 44(4):1069-1074, 2015]. SORL1 processes a causal involvement in Alzheimer's disease (AD) as a proposed modulator of the amyloid precursor protein (APP). It is unclear whether SORL1 is involved with CAA-ri or not.

Cerebral amyloid angiopathy-related inflammation: a case report presenting with a rare variant in SORL1 gene.

Lowering homocysteine (HCY) has beneficial effects on vascular events in primary prevention but not in secondary prevention. Research on serum HCY level and middle cerebral artery (MCA) stenosis is lacking. The purpose of the study was to determine the association between these factors and provide more evidence for the prevention of ischemic stroke.

Lack of association between serum homocysteine level and middle cerebral artery stenosis.

Multiple sclerosis (MS) is one of the commonest neurologic disorders globally. LncRNA OIP5-AS1 has been found to be implicated in the etiology of MS. This study was to explore the roles and molecular mechanisms of OIP5-AS1 in the development of MS.

OIP5-AS1 facilitates Th17 differentiation and EAE severity by targeting miR-140-5p to regulate RhoA/ROCK2 signaling pathway.

In recent years, more attention has been paid to the biological effects of short-chain chlorinated paraffin (SCCP). Studies have shown that SCCPs exposure could cause metabolic damage and lipid metabolic damage. In the present work, based on E. coli membrane damage experiments and molecular dynamics (MD) simulation, the effects of SCCPs on the membrane structure and membrane properties were studied to explore the possible toxic damage effects of SCCPs on cell membrane. Experiments results showed that SCCPs had a significant inhibitory effect on E. coli. The E. coli cell membrane of the bacteria was broken and the macromolecules of the cell flowed out when exposed to SCCPs. SCCPs would lead to the decrease and depolarization of cell membrane potential, and then affect the integrity and permeability of cell membrane. The further molecular dynamic simulation revealed that SCCP molecules can easily enter the lipid DPPC membranes from the aqueous phase and tended to aggregate inside bilayer stably. The bound of SCCPs could lead to significant variations in DPPC bilayer with a less dense, more disorder and rougher layer, which thus made the damage of cell membrane. In a word, although the overall toxicity of SCCPs to cell was relatively weak, the damage to the cell membrane may be one of the mechanisms of its toxicity. MAIN FINDING OF THE WORK: The exposure of SCCPs could cause structural change of cell membrane in E. coli, which verified the damage to the cell membrane may be one of the mechanisms of its toxicity.

Effect of short-chain chlorinated paraffins (SCCPs) on lipid membranes: Combination of molecular dynamics and membrane damage experiments.

 Hereditary spastic paraplegia (HSP) caused by mutations in  have been reported as spastic paraplegia 9 (SPG9), with autosomal dominant and autosomal recessive transmission (SPG9A and SPG9B). SPG9 is rare and has shown phenotypic and genotypic heterogeneity in previous reports.  This study screened  mutations in autosomal recessive HSP patients using combined whole exome sequencing and RNA splicing analysis. We conducted  investigations, co-segregation analysis, and ELISA-based analysis of P5CS (Δ1-pyrroline-5-carboxylate synthetase; encoded by ) concentration to validate the pathogenicity of the detected  variants. All previously reported bi-allelic  mutations and cases were reviewed to summarize the genetic and clinical features of -related HSP.  A novel missense mutation c.880T>C, p.S294P and an intronic splicing mutation c.-28-13A>G were both detected in  in an autosomal recessive family presenting with a complicated form HSP. ELISA assays revealed significantly decreased P5CS concentration in the proband's plasma compared with that in the healthy controls. Moreover, review of previously reported recessive cases showed that SPG9B patients in our cohort presented with milder symptoms, i.e., later age at onset and without cognitive impairment.  The present study expands the genetic and clinical spectrum of SPG9B caused by  mutation. Our work defines new genetic variants to facilitate future diagnoses, in addition to demonstrating the highly informative value of splicing mutation prediction in the characterization of disease-related intronic variants.

Novel Compound Missense and Intronic Splicing Mutation in  Causes Autosomal Recessive Spastic Paraplegia.

As an important pollutant, perfluorooctane acid (PFOA) has been widely concerned and reported by thousands of times, while less is known about the concentration-response pathway of PFOA. The aim of the present work was to reveal the concentration-response mechanism of PFOA in human cells. Omics results showed that calcium-related pathways play key roles in PFOA injury mechanisms. The results of GO and KEGG analyses showed that the cAMP signaling pathway was presented as the top one in all of the regulatory patterns and concentrations groups of PFOA. In the cAMP signaling pathway, the adenosine A receptor (ADORA1) recognized the low concentration of PFOA and induced pathway "G-cAMP-PKA" to decrease the concentration of cAMP. This indicated that the low concentration of PFOA may promote breast hyperplasia and inhibit lactation. While adenosine A receptor (ADORA2A) recognized the high concentration of PFOA and induced pathway "G-AC-cAMP-RKA" to increase the concentration of cAMP, induce cell damage and may lead to the deterioration of breast cancer. The results of molecular dynamics simulation showed that PFOA could bind to ADORA1 and ADORA2A, thus cause subsequent signal transduction. Furthermore, considering the strong binding ability of PFOA with ADORA1, PFOA tends to bind to ADORA1 at a low concentration. On the other side, PFOA at high concentration will continue to bind to another receptor protein, ADORA2A, and activate subsequent signaling pathways. Combined analyses of transcriptomic and proteomic revealed that different concentrations of PFOA regulate cellular calcium-related pathways. The cAMP pathway showed a concentration-response effect of PFOA. After treatment with different concentrations of PFOA, ADORA1 and ADORA2A were activated respectively, showing opposite cellular effects, leading to kinds of breast lesions. In the nervous system, PFOA might induce a variety of nervous system diseases. The present work was an exploration on the toxicological mechanism of PFOA, providing important information on the health impacts of PFOA in humans.

PFOA regulate adenosine receptors and downstream concentration-response cAMP-PKA pathway revealed by integrated omics and molecular dynamics analyses.

Ionizing radiation, especially heavy ion (HI) beams, has been widely used in biology and medicine. However, the mechanism of membrane damage by such radiation remains primarily uncharacterized.

LsrB, the hub of ABC transporters involved in the membrane damage mechanisms of heavy ion irradiation in .

Chemoresistance is a major obstacle faced by oesophageal cancer patients and is synonymous with a poor prognosis. MCL1 is a pivotal member of the anti-apoptotic Bcl-2 protein family, which has been found to play an important role in cell survival, proliferation, differentiation and chemoresistance. Thus, it might be an ideal target for treating oesophageal cancer patients. Although it is known that MCL1 is degraded via the ubiquitin-proteasome system, the deubiquitylating enzyme (DUB) responsible for stabilizing MCL1 remains elusive to date. Herein, we demonstrate that Ubiquitin-Specific Protease 20 (USP20) is a novel regulator of the apoptotic signaling pathway. Moreover, USP20 could regulate the deubiquitination of MCL1 to, in turn, regulate its stability. Increased expression of USP20 was correlated with increased levels of MCL1 protein in human patient samples. In addition, depletion of USP20 could increase the polyubiquitination of MCL1, thereby increasing the conversion rate of MCL1 and the sensitivity of cells to chemotherapy. Overall, our findings indicate that the USP20-MCL1 axis might play a key role in the apoptotic signaling pathway.

The deubiquitinating enzyme USP20 regulates the stability of the MCL1 protein.

 berry (Ningxia Gouqi, , goji berry, or wolfberry), as a traditional Chinese herb, was recorded beneficial for longevity in traditional Chinese medical scriptures and currently is a natural dietary supplement worldwide. However, under modern experimental conditions, the longevity effect of  berry and the underlying mechanisms have been less studied. Here, we reported that total water extracts of  berry (LBE), which contains 22% polysaccharides and other components, such as anthocyanins, extended the lifespan of  without side effects on worm fertility and pharyngeal pumping. Interestingly, we found that the lifespan extension effect was more prominent in worms with shorter mean lifespan as compared to those with longer mean lifespan. Furthermore, we showed that the lifespan extension effect of LBE depended on deacetylase . Remarkably, LBE rescued heat shock transcription factor-1 () deficiency in wild-type worms with different mean lifespans, and this effect also depended on . In addition, we found that LBE extended lifespan and alleviated toxic protein aggregation in neurodegenerative worms with  deficiency. Our study suggested that LBE may be a potential antiaging natural dietary supplement especially to individuals with malnutrition or chronic diseases and a potential therapeutic agent for neurodegenerative diseases characterized by  deficiency.

 Extracts Extend Lifespan and Alleviate Proteotoxicity in .

Myasthenia gravis (MG) is an autoimmune disorder caused by autoantibodies directed against the postsynaptic membrane at the neuromuscular junction. Perturbation of gut microbiota is thought to contribute to the development of MG, as reflected by fecal metabolomic signatures in humans, but there have been few studies on the relationship between oral microbiota profile and MG. The current study evaluated the correlation between oral microbiota composition and diversity and anti-acetylcholinereceptor (AChR) antibody-positive MG by comparing oral microbiota communities of patients  = 20) and healthy controls (HCs;  = 20) by 16S rRNA gene sequencing. Principal coordinate analysis and Adonis analysis revealed significant differences in oral microflora profile between the twogroups. Compared to HCs, the abundance of the phyla  and  and genera , and  was significantly increased whereas that of phyla  and and genera , and  was significantly decreased in MG patients. The Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the biosynthesis of ansamycins and amino acid metabolism pathways were altered in MG. These results indicate that oral microbiota composition is perturbed in patients with anti-AChR antibody-positive MG, providing new potential avenues for targeted therapeutic interventions.

Oral Microbiota Profile in a Group of Anti-AChR Antibody-Positive Myasthenia Gravis Patients.

 leaves are a kind of vegetable, and modern nutrition studies have found that they have an anti-aging function. Our study aims to investigate the anti-aging effects of  leaf flavonoid (LBLF) extracts and its underlying molecular mechanism. LBLFs were purified using D101 and polyamide resin, characterized by ultraperformance liquid chromatography coupled with mass spectrometry, and administered to hydrogen peroxide (HO)-treated human umbilical vein endothelial cells (HUVECs) and . Appropriate enrichment conditions were optimized through dynamic adsorption and desorption experiments, the content of flavonoids reached 909.84 mg/g, rutin and kaempferol being the main ones. LBLFs attenuated HO-induced HUVEC apoptosis, decreased reactive oxygen species and malondialdehyde production levels, increased superoxide dismutase, glutathione peroxidase and catalase activities. Furthermore, pre-treatment with LBLF increased mRNA expression of erythropoietin (EPO) and heme oxygenase-1 (HO-1) via the mitogen-activated protein kinase (MAPK) signaling pathway in HUVECs. Compared with 100 µM rutin monomer, LBLF prolonged the lifespan of , enhanced their mobility in middle life stages and upregulated expression of ,  and  genes, which indicated that the anti-aging effects of LBLF were due to its redox-modulation.

Flavonoids from  Leaves Exhibit Anti-Aging Effects through the Redox-Modulation.

Transthyretin protein-related familial amyloidosis polyneuropathy (TTR-FAP) is an autosomal dominant genetic disease caused by mutations in the TTR gene. The disease is characterized primarily by peripheral and autonomic nerve damage. Disease progression is associated with frequent involvement of the heart, lungs, kidneys, eyes, and other organs. The most common TTR mutation is c.148G>A (p.Val50Met), although the FAP resulting from the mutation rarely involves the spinal cord.

Article	Year
Nanog attenuates lipopolysaccharide-induced inflammatory responses by blocking nuclear factor-κB transcriptional activity in BV-2 cells. Neuroreport\| PubMed ID: 23924954	2013
Inhibition of JAK1 by microRNA-708 promotes SH-SY5Y neuronal cell survival after oxygen and glucose deprivation and reoxygenation. Neuroscience letters\| PubMed ID: 29128627	2018
Cerebral amyloid angiopathy-related inflammation: a case report presenting with a rare variant in SORL1 gene. BMC neurology\| PubMed ID: 31092209	2019
Lack of association between serum homocysteine level and middle cerebral artery stenosis. Brain and behavior\| PubMed ID: 31225691	2019
OIP5-AS1 facilitates Th17 differentiation and EAE severity by targeting miR-140-5p to regulate RhoA/ROCK2 signaling pathway. Life sciences\| PubMed ID: 33515560	2021
Effect of short-chain chlorinated paraffins (SCCPs) on lipid membranes: Combination of molecular dynamics and membrane damage experiments. The Science of the total environment\| PubMed ID: 33631584	2021
Novel Compound Missense and Intronic Splicing Mutation in Causes Autosomal Recessive Spastic Paraplegia. Frontiers in neurology\| PubMed ID: 34093392	2021
PFOA regulate adenosine receptors and downstream concentration-response cAMP-PKA pathway revealed by integrated omics and molecular dynamics analyses. The Science of the total environment\| PubMed ID: 34500266	2022
LsrB, the hub of ABC transporters involved in the membrane damage mechanisms of heavy ion irradiation in . International journal of radiation biology\| PubMed ID: 34597255	2021
The deubiquitinating enzyme USP20 regulates the stability of the MCL1 protein. Biochemical and biophysical research communications\| PubMed ID: 35063767	2022
Extracts Extend Lifespan and Alleviate Proteotoxicity in . Frontiers in nutrition\| PubMed ID: 35096951	2021
Oral Microbiota Profile in a Group of Anti-AChR Antibody-Positive Myasthenia Gravis Patients. Frontiers in neurology\| PubMed ID: 35873770	2022
Flavonoids from Leaves Exhibit Anti-Aging Effects through the Redox-Modulation. Molecules (Basel, Switzerland)\| PubMed ID: 35956901	2022
Transthyretin-Related Familial Amyloidosis Polyneuropathy with Spinal Cord Damage: A Case Report. International medical case reports journal\| PubMed ID: 39790491	2025

Haitao Zhou

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