Proteomics technology reveals the marker proteins, potential pathogenesis, and intervention targets after noise-induced hearing loss. To study the differences in cochlea protein expression before and after noise exposure using proteomics to reveal the pathological mechanism of noise-induced hearing loss (NIHL). A guinea pig NIHL model was established to test the ABR thresholds before and after noise exposure. The proteomics technology was used to study the mechanism of differential protein expression in the cochlea by noise stimulation. The average hearing threshold of guinea pigs on the first day after noise exposure was 57.00 ± 6.78 dB Sound pressure level (SPL); the average hearing threshold on the seventh day after noise exposure was 45.83 ± 6.07 dB SPL. The proteomics technology identified 3122 different inner ear proteins, of which six proteins related to the hearing were down-regulation: Tenascin C, Collagen Type XI alpha two chains, Collagen Type II alpha one chain, Thrombospondin 2, Collagen Type XI alpha one chain and Ribosomal protein L38, and are enriched in protein absorption, focal adhesion, and extracellular matrix receptor pathways. Impulse noise can affect the expression of differential proteins through focal adhesion pathways. This data can provide an experimental basis for the research on the prevention and treatment of NIHL.

Angiogenesis, the formation of new vessels, is a critical step in the malignancy progression of solid tumors. Many investigations have demonstrated the usefulness of immunotoxins to halt angiogenesis in solid tumors. Pharmaceutically, Vascular Endothelial Growth Factor (VEGF) can deliver coupled toxins to the tumor vessels through VEGF Receptors. In the current study, we designed, expressed, and assessed the in vitro and in vivo toxicities of a novel immunotoxin consisting of mouse VEGF and heminecrolysin toxin (mVEGF-HNc). The fusion protein was expressed in E. coli and purified via Ni+2 affinity chromatography. The biological activity of immunotoxin was evaluated on NIH/3T3 cells and TC1-tumorized mouse model. The mVEGF-NHc showed significant cytotoxicity on the cells as VEGFR-expressing cells. Moreover, the size of the tumor in the mVEGF-HNc-treated group started to reduce after six injections, while it continued to grow in the PBS-received mice. Efficacious targeting of solid tumor cells via mVEGF-HNc suggests its prospective therapeutic potential for cancer therapy.

Fecal microbiota transplantation is an emerging disease-modifying therapy. The viability of the microbiome in feces and its successful transfer depends on the preparation of fecal microbiota suspension. However, currently, no standard operation procedure is proposed for fecal suspension preparation. This study aims to compare the effect of different preparation methods on the composition of fecal microbiota composition in the rat. Four methods were used to collect the fecal suspension from fresh rat fecal (Group A), including stirring with normal saline (Group B), stirring with normal saline and then standing (Group C), stirring with normal saline and filtered with gauze (Group D), and stirring with normal saline and centrifuged (Group E). 16S ribosomal RNA gene (16S rDNA) sequencing technology was used to analyze the microbiota diversity and composition of each group of samples. Compared with fresh feces, the bacterial richness of the fecal suspension obtained by the four methods was significantly decreased (P < 0.05). The structural similarity with fresh fecal microbiota from high to low is groups B, D, C, and E. All four methods changed the microbiota structure to varying degrees, thus may affect the effect of FMT. In conclusion, choosing different methods to prepare fecal suspensions may help to better optimize the application of FMT.

Dengue fever is a mosquito-borne disease that claims the lives of millions of people around the world. A number of factors like disease's non-specific symptoms, increased viral mutation, growing antiviral drug resistance due to reduced susceptibility, unavailability of an effective vaccine for dengue, weak immunity against the virus, and many more are involved. Dengue belongs to the Flaviviridae family of viruses. The two species of the vector transmitting dengue are Aedes aegypti and Aedes albopictus, with the former one being dominant. Serotypes 2 of dengue fever are spread to the human body and cause severe illness. Recently, dengue has imposed an aggressive effect synergistically with the COVID-19 pandemic. As a result, we concentrated our efforts on finding a potential therapeutic. For this, we chose natural compounds to fight dengue fever, which is currently regarded as successful among many drug therapies. Following this, we started the in silico experiment with 922 plant extracts as lead compounds to fight serotype 2. In this study, we used SwissADME for analyzing ligand drug-likeness, pkCSM for designing an ADMET profile, Autodock vina 4.2 and Swissdock tools for molecular docking, and finally Desmond for molecular dynamics simulation. Ultimately 45 were found effective against the 2'O methyltransferase protein of serotype 2. CHEMBL376820 was found as possible therapeutic candidates for inhibiting methyltransferase protein in this thorough analysis. Nevertheless, more in vitro and in vivo research are required to substantiate their potential therapeutic efficacy.

There is an increasing demand for natural food preservatives due to consumers' concern on the negative effects of chemical preservatives in food products. Nisin (bacteriocin) is an effective food biopreservative that has been approved globally. However, its low yield proves to be a limiting factor and must be addressed to meet the increasingly high demand from the food industry. The present work thus investigated the effects of individual and combined fermentation factors on Lactococcus lactis ATCC 11454 growth and nisin activity using the one-factor-at-a-time (OFAT) method. The level of each factor that gave the highest nisin production was then selected and combined to further improve its activity. The best combined conditions for highest cell growth and nisin activity were 30 °C, pH 6.0, and mild agitation with the addition of 1.0% w/v glucose, 1.0% w/v skim milk, and 0.5% v/v Tween 20. This increased nisin production by 22.7% as compared to control (basic condition). The present work provided critical information on the relationship between fermentation conditions, growth, and nisin activity of L. lactis ATCC 11454 that could be explored to understand the potential and limitation of the strain. This fermentation strategy can also serve as a benchmark to further enhance the production of bacteriocin or other biopreservative compounds.

N6-methyladenosine (m6A) methylation regulates pathological processes of cerebral stroke, which can lead to disability and death. Herein, we explored the role of a m6A "reader" YTHDF1 in stroke. MCAO (middle cerebral artery occlusion) rat model and hypoxia/reoxygenation (H/R)-induced neurocytes cell model were established. TTC staining assay assessed the infarction area and TUNEL assay analyzed apoptosis. Neurological score was analyzed to evaluate the brain function. Cell counting kit-8, LDH release, and flow cytometry assessed cellular proliferation, cell death, and cell apoptosis in vitro. The expression of YTHDF1, PTEN, and the factors in the PI3K/AKT/mTOR pathway was measured using western blot. The interaction between YTHDF1 and PTEN was confirmed luciferase assay and RNA immunoprecipitation assay. The results indicated that YTHDF1 was upregulated in the brain tissues of MCAO mice and H/R-treated cells. Knockdown of YTHDF1 inhibited the infarct area, neuron damage, and apoptosis. Additionally, YTHDF1 depletion promoted viability and inhibited apoptosis of H/R-treated cells. Moreover, YTHDF1 inactivated the PI3K/AKT/mTOR pathway. Mechanistically, YTHDF1 binds to PTEN to increase PTEN mRNA stability. Overexpressing PTEN rescued the effects of YTHDF1 depletion on cell viability and apoptosis. In conclusion, silencing of YTHDF1 decelerated the progression of cerebral stroke through promoting PTEN degradation and activating the PTEN/AKT/mTOR pathway, suggesting that YTHDF1 has the potential to be a therapeutic target for stroke.

Prime Editing is a CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) based genome editing technique having promising potential in terms of reducing off target activity. It introduces fragments of DNA sequences into the target site using a guide RNA (gRNA) molecule, composed of both the sequence that is to be inserted into the target site along with an inactive Cas9 nickase and a reverse transcriptase. Prime Editing can cause insertions, deletions, and various point mutations for reverting the phenetic characteristics of a disease specially tested in human adult stem cells and cancer cell lines. The main aim of our review is to explore how Prime Editing and its various forms are being utilized as an emerging tool to cure deleterious diseases like cancer, also as a delivery strategy of the tool into cells. There are almost five generations of Prime Editors (PE) with increasing levels of efficiency from one level to another that have huge clinical potential in correcting mutations; however, the necessity for a pegRNA design is extremely significant. But besides having such advantages, the limitations of this technology particularly include generation of double nicks while optimizing the efficiency of PE3. So, it is important to consider all such consequences and customize PE as per requirements.

Lumbar degenerative disc disease (LDDD) is frequently misdiagnosed as other spine conditions, and the accurate diagnosis is challenging. This study was conducted to explore the role of circRNA GRB10 in the accurate diagnosis of LDDD. This study included 60 cases of LDDD, 60 cases of patients with sacroiliac joint pain (SJP), 60 cases of lumbar disc herniation (LDH), 60 cases of piriformis syndrome (PS), 60 cases of entrapment neuropathy (EN) and 60 cases of healthy controls (HCs). Plasma was obtained from each patient before and after treatment. Expression of GRB10 was studied with RT-qPCR. The role of plasma GRB10 in the accurate diagnosis of LDDD was analyzed by ROC curve analysis. Compared to HCs, decreased accumulation of GRB10 RNA was only observed in LDDD group, but not in SJP, LDH, PS and EN groups. With plasma expression level of GRB10 measured before treatment as a biomarker, LDDD patients were separated from SJP, LDH, PS, EN and HC groups. After treatment, increased expression levels of GRB10 were only observed in LDDD group, but not in other groups. Therefore, GRB10 was downregulated in LDDD and may serve as a biomarker for the accurate diagnosis of LDDD.

Epidermal growth factor receptor (EGFR) is a transmembrane glycoprotein receptor with intracellular tyrosine kinase activity. Mutations in the EGFR gene, including deletions in exon 19 and the mutation L858R, induce responsiveness of non-small cell lung cancer (NSCLC) to a group of drugs known as tyrosine kinase inhibitors. Here, we report the development of the CRISPR-based fluorescent reporter (CBFR) assay including a two-step strategy combining PCR amplification and Cas12a-driven cleavage to detect the delE746_A750 subtype of EGFR exon 19 deletions. Sensitivity and specificity of the CBFR assay were analyzed with different concentrations of fluorescence reporter and different amounts of PCR product. The results demonstrated that increasing the fluorescent reporter to 4 μM and the PCR product to 5 μl enhanced sensitivity. The CBFR assay could detect EGFR exon 19 deletion even with a frequency of 1% in samples. In clinical NSCLC samples, optimized CBFR assay enabled visual detection of the delE746_A750 subtype in less than 1 h. The CBFR assay provides a sensitive, specific, and simple strategy designed based on a straightforward and inexpensive process. We suggest that the CBFR assay could serve as a diagnostic approach to detect mutations, deletions, and pathogens in underequipped laboratories and promote personalized therapeutic approaches.

Lung adenocarcinoma (LUAD), a prevalent form of non-small cell lung cancer (NSCLC), has a high incidence and mortality rate. However, its molecular regulatory mechanisms have yet to be fully understood. The purpose of this study was to look into how NADPH quinone oxidoreductase-1 (NQO1) and it miR-485-5p and affected LUAD cells. The levels of miR-485-5p and NQO1 expression in LUAD cells and tissues were determined by means of quantitative reverse transcription polymerase chain reaction. The viability, proliferation, migration, and apoptosis of LUAD cells were assessed using cell counting Kit-8, 5-bromo-2'-deoxyuridine, transwell, and caspase-3 assays, respectively. Western blot experiments were used to examine the relative protein expression of matrix metallopeptidase 2 and matrix metallopeptidase 9, as well as the phosphorylation of phosphoinositide 3-kinase (PI3K) and protein kinase B (Akt) in LUAD cells. Luciferase and RNA pull-down experiments were also conducted for the verification of miR-485-5p's underlying relationship with NQO1. In our study, we found that LUAD cells and tissues had miR-485-5p downregulation and NQO1 upregulation. The experimental outcomes indicated that miR-485-5p overexpression in LUAD cells reduced their malignant behaviors, suppressed PI3K and Akt phosphorylation, and facilitated apoptosis. The results also revealed that NQO1 was a direct miR-485-5p target, and that NQO1 could reverse miR-485-5p's inhibitory effect on the malignant phenotype of LUAD cells. Furthermore, it was also observed that through targeting NQO1, miR-485-5p could suppress LUAD cell migration and proliferation, further blocking the phosphorylation of PI3K and Akt and inducing apoptosis among LUAD cells. In conclusion, the miR-485-5/NQO1 axis regulates LUAD progression through the PI3K/Akt pathway.

In recent years, within tissue engineering, cell growth on patterned surfaces have gained significant attention. Growing cells in patterns is important to manufacture polymeric tissues that can be used within the medical field. For this reason, the main focus of this study was to prepare patterned scaffolds using Titanium (Ti) and polyvinyl chloride (PVC) covered on microscope lamellas and examine their liability for cell growth. A polydimethylsiloxane stamp was initially prepared which was then used to transfer a predefined pattern onto PVC- and Ti-covered surfaces. Cell growth experiments were performed on the prepared materials by seeding L929 mouse fibroblasts. The growth of cells seeded on the surface of the scaffolds were spectroscopically followed using Neutral Red uptake assay. The results showed cell proliferation on both patterned surfaces, however, it was higher on Ti-covered samples. In addition, three different alkanethiols were tested for cell adhesion on patterned surfaces. A higher number of cell proliferation was observed with undecanethiol, which has a shorter alkane group among them. The morphological properties of the samples before and after cell-seeding were analyzed via scanning electron microscope and optical microscopy. Significant amount of cell proliferation was observed on all of the prepared samples.

Erratum in Mol Biotechnol. 2022 Oct 31;:

An ongoing pandemic of coronavirus disease 2019 (COVID-19) is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). So far, there have been various approaches for SARS-CoV-2 detection, each having its pros and cons. The current gold-standard method for SARS-CoV-2 detection, which offers acceptable specificity and sensitivity, is the quantitative reverse transcription-PCR (qRT-PCR). However, this method requires considerable cost and time to transport samples to specialized laboratories and extract, amplify, and detect the viral genome. On the other hand, antigen and antibody testing approaches that bring rapidity and affordability into play have lower sensitivity and specificity during the early stages of COVID-19. Moreover, the immune response is variable depending on the individual. Methods based on clustered regularly interspaced short palindromic repeats (CRISPR) can be used as an alternative approach to controlling the spread of disease by a high-sensitive, specific, and low-cost molecular diagnostic system. CRISPR-based detection systems (CRISPR-Dx) target the desired sequences by specific CRISPR-RNA (crRNA)-pairing on a pre-amplified sample and a subsequent collateral cleavage. In the present article, we have reviewed different CRISPR-Dx methods and presented their benefits and drawbacks for point-of-care testing (POCT) of suspected SARS-CoV-2 infections at home or in small clinics.

Hepatocellular carcinoma (HCC) is one of the most commonly diagnosed malignancy globally with a pessimistic prognosis. Previous studies have demonstrated that abnormal expression of genes in the lysine-specific histone demethylase 3 (KDM3) family with epigenetic changes and dysregulation of enzymes promotes cancer progression. In this study, multiomics analyses were utilized to analyze differential expression, prognostic value, genetic alteration, protein-protein interaction, associated biological pathways and immune cell infiltration of KDM3s in patients with HCC. KDM3A-C were significantly upregulated to different extents based on pathologic and tumor grades in patients with HCC compared to normal tissue. Of note, higher KDM3A expression was associated with poor survival in HCC patients, whereas KDM3B and KDM3C were not associated with survival. Furthermore, KDM3A-B genetic alterations had significant effects on survival in patients with HCC. Analyses of the KEGG pathway and miRNAs targets of KDM3A and KDM3B in HCC may provide potential value in tumor behaviors and treatment. The differential expression of the KDM3 family has a strongly significant correlation with the infiltration of the abundance of immune cells, including B cells, CD8+ T cells, CD4+ T cells, macrophages, neutrophils, and dendritic cells in HCC. This study indicates that KDM3A may have the potential to be a promising molecular target in terms of prognostic biomarkers or therapeutic targets for HCC treatment.