Overexpression Of Protein Research Articles

Huanglian decoction prevents and treats radiation-induced intestinal injury in lung cancer by regulating endoplasmic reticulum stress.

Radiotherapy is a primary treatment method for lung cancer, and radiation-induced lung injury has been widely studied. However, the lung and intestines are closely related, and patients receiving lung radiotherapy often experience gastrointestinal reactions. Huanglian Decoction has proven effective in treating intestinal diseases, but its role in radiation-induced intestinal injury in lung cancer has not yet been demonstrated. The study investigated the potential protective mechanisms of Huanglian Decoction against radiation-induced intestinal injury in lung cancer. In vivo experiments were conducted to examine the morphological changes. Changes in endoplasmic reticulum stress-related proteins were assessed using electron microscopy, immunohistochemistry, immunofluorescence, and Western blotting. In vitro studies involved the overexpression of TRPA1 protein in NCM460 cells via lentiviral vectors. Tumor-bearing mice exhibited severe damage to both lung and colon tissues following radiotherapy, with elevated levels of IL-33, increased expression of ST2L and TRPA1 in colon tissues, higher levels of endoplasmic reticulum stress-related proteins, and the presence of apoptosis and inflammatory responses. Huanglian Decoction reduced radiation-induced intestinal injury by lowering IL-33 levels, which in turn reduced endoplasmic reticulum stress response in colon tissues. In TRPA1-overexpressing NCM460 cells, Huanglian Decoction decreased TRPA1 expression levels and significantly alleviated endoplasmic reticulum stress response. Conclusively, the results indicate that Huanglian Decoction alleviates radiation-induced intestinal endoplasmic reticulum stress by reducing IL-33 levels, subsequently inhibiting the expression of ST2L and TRPA1 in colon tissues. This demonstrates the protective effect of Huanglian Decoction on the intestines during lung cancer radiotherapy, highlighting its promising clinical application in radiation protection.

Electroacupuncture Mitigates TRPV1 Overexpression in the Central Nervous System Associated with Fibromyalgia in Mice

Background: Fibromyalgia (FM) is characterized by chronic pain, significantly affecting the quality of life and functional capabilities of patients. In addition to pain, patients may experience insomnia, chronic fatigue, depression, anxiety, and headaches, further complicating their overall well-being. The Transient Receptor Potential Vanilloid 1 (TRPV1) receptor responds to various noxious stimuli and plays a key role in regulating pain sensitivity and inflammation. Thus, targeting TRPV1 may provide analgesic and anti-inflammatory benefits. This study investigates the efficacy of electroacupuncture (EA) in alleviating chronic pain in FM through TRPV1 and its downstream molecules in the central nervous system (CNS). Methods: To model FM, we subjected mice to intermittent cold stress (ICS) for three days. The study comprised five rodent groups: Control (CON), ICS, ICS + EA, ICS + Sham EA, and ICS + KO (TRPV1 knockout mice). Results: Our findings revealed that ICS induced allodynia and hyperalgesia in mice by day four, persisting until day 21. EA at 2 Hz and TRPV1 KO significantly decreased both mechanical and thermal hypersensitivity (Withdrawal—Day 14: 2.43 ± 0.19 g; Day 21: 5.88 ± 0.47 g, n = 6, p < 0.05; Latency—Day 14: 2.77 ± 0.22 s; Day 21: 5.85 ± 0.41 s, n = 6, p < 0.05). In contrast, sham EA did not produce significant effects. Additionally, TRPV1 and several pain-related proteins were significantly elevated in the thalamus, somatosensory cortex (SSC), medial prefrontal cortex (mPFC), hippocampus, hypothalamus, cerebellum regions V (CB V), VI (CB VI) and VII (CB VII) after the ICS model. Both EA at the ST36 acupoint and TRPV1 KO mice showed diminished overexpression of pain-related proteins, with the sham EA group showing no significant changes compared to the ICS group. Conclusions: Chronic widespread pain was reduced by EA and TRPV1 KO, with the effects of EA on the TRPV1 pain pathway clearly evident in the CNS after 21 days.

Lycium barbarum oligosaccharide-derived carbon quantum dots inhibit glial scar formation while promoting neuronal differentiation of neural stem cells

Overexpression of glial fibrillary acidic protein (GFAP) in activated astrocytes following spinal cord injury is closely associated with glial scar formation, which harms axonal regrowth. In this study, we prepared ultrasmall cationic carbon quantum dots (CQDs) via one-step hydrothermal carbonization. Lycium barbarum oligosaccharides were used as the carbon source for the first time, and polyetherimide (PEI) and ethylenediamine (ED) were used as cationic reagents. Interestingly, the resultant CQDs show the bioactivity of specifically inhibiting GFAP protein expression, while promoting neuronal marker expression in neural stem cells (NSCs). Furthermore, CQDs together with NSCs can remarkably improve the motor activity of animals after implantation into the transection lesion of the rat spinal cord. Histological analysis confirmed that CQDs can enhance neuronal differentiation of NSCs while inhibiting glial scar formation in vivo. Altogether, this study represents the first report of producing CQDs from oligosaccharides and investigating their impact on NSCs differentiation, thus providing a paradigm for exploring the bioactivity of quantum dots.

Overexpression of enhanced yellow fluorescent protein fused with Channelrhodopsin-2 causes contractile dysfunction in skeletal muscle.

Skeletal muscle activation using optogenetics has emerged as a promising technique for inducing noninvasive muscle contraction and assessing muscle function both invivo and invitro. Transgenic mice overexpressing the optogenetic fusion protein, Channelrhodopsin 2-EYFP (ChR2-EYFP) in skeletal muscle are widely used; however, overexpression of fluorescent proteins can negatively impact the functionality of activable tissues. In this study, we characterized the contractile properties of ChR2-EYFP skeletal muscle and introduced the ChR2-only mouse model that expresses light-responsive ChR2 without the fluorescent EYFP in their skeletal muscles. We found a significant reduction in the contractile ability of ChR2-EYFP muscles compared with ChR2-only and WT mice, observed under both electrical and optogenetic stimulation paradigms. Bulk RNAseq identified the downregulation of genes associated with transmembrane transport and metabolism in ChR2-EYFP muscle, while the ChR2-only muscle did not demonstrate any notable deviations from WT muscle. The RNAseq results were further corroborated by a reduced protein-level expression of ion channel-related HCN2 in ChR2-EYFP muscles and gluconeogenesis-modulating FBP2 in both ChR2-EYFP and ChR2-only muscles. Overall, this study reveals an intrinsic skeletal dysfunction in the widely used ChR2-EYFP mice model and underscores the importance of considering alternative optogenetic models, such as the ChR2-only, for future research in skeletal muscle optogenetics.

Organic photoelectrochemical transistor biosensor based on BiVO4-ZnIn2S4 material for efficient and sensitive detection of MCF-7 cells

Recently, organic photoelectrochemical transistor (OPECT) has become a very interesting biological measurement method in photoelectrochemical (PEC) bioanalysis and future bio-related applications. OPECT is expected to be a powerful tool for disease detection and early warning. Circulating tumor cells (CTCs) are generally deemed to be the dominant factor of tumor metastasis, and 90 % of cancer patients die from this metastatic disease. Therefore, there is an imminent need to develop a highly sensitive CTCs detection sensing system to improve the survival rate of cancer patients. Here, we use a DNA tetrahedrons (DNA NTH) with an aptamer at the top to immobilize on the surface of the photoelectric material to capture cells (MCF-7). Specifically, the BiVO4-ZnIn2S4 hybrid was synthesized by a simple hydrothermal method, which can effectively modulated devices with high current gain. Au NPs were directly integrated on the electrode surface to construct an OPECT photoelectric sensing platform. Subsequently, the aptamer which is thiol-functionalized (SH-Apt) was immobilized on the electrode surface. Because of the overexpression of MUC1 protein on the cell membrane, it can specifically capture MCF-7 cells. The introduction of MCF-7 cells resulted in a significant decrease in the current signal. There is a relationship between the change of photocurrent and the logarithm of MCF-7 cell concentration, which is a good linear relationship ranging from 50 to 5 × 105 cell mL−1. The obtained detection limit is 43 cell mL−1. The biosensor has high selectivity and sensitivity, and achieves sensitive detection of MCF-7.

Modeling and measuring how codon usage modulates the relationship between burden and yield during protein overexpression in bacteria.

Excess utilization of translational resources is a critical source of burden on cells engineered to over-express exogenous proteins. To improve protein yields and genetic stability, researchers often use codon optimization strategies that improve translational efficiency by matching an exogenous gene's codon usage with that of the host organism's highly expressed genes. Despite empirical data that shows the benefits of codon optimization, little is known quantitatively about the relationship between codon usage bias and the burden imposed by protein overexpression. Here, we develop and experimentally evaluate a stochastic gene expression model that considers the impact of codon usage bias on the availability of ribosomes and different tRNAs in a cell. In agreement with other studies, our model shows that increasing exogenous protein expression decreases production of native cellular proteins in a linear fashion. We also find that the slope of this relationship is modulated by how well the codon usage bias of the exogenous gene and the host's genes match. Strikingly, we predict that an overoptimization domain exists where further increasing usage of optimal codons worsens yield and burden. We test our model by expressing sfGFP and mCherry2 from constructs that have a wide range of codon optimization levels in Escherichia coli . The results agree with our model, including for an mCherry2 gene sequence that appears to lose expression and genetic stability from codon overoptimization. Our findings can be leveraged by researchers to predict and design more optimal cellular systems through the use of more nuanced codon optimization strategies.

Hepatovirus translation requires PDGFA-associated protein 1, an eIF4E-binding protein regulating endoplasmic reticulum stress responses.

The overexpression and misfolding of viral proteins in the endoplasmic reticulum (ER) may cause cellular stress, thereby inducing a cytoprotective, proteostatic host response involving phosphorylation of eukaryotic translation initiation factor 2 subunit alpha (eIF2α). Here, we show that hepatitis A virus, a positive-strand RNA virus responsible for infectious hepatitis, adopts a stress-resistant, eIF2α-independent mechanism of translation to ensure the synthesis of viral proteins within the infected liver. Cap-independent translation directed by the hepatovirus internal ribosome entry site and productive hepatovirus infection of mice both require platelet-derived growth factor subunit A (PDGFA)-associated protein 1 (PDAP1), a small phosphoprotein of unknown function with eIF4E-binding activity. PDAP1 also interacts with eIF1A and is essential for translating stress-resistant host messenger RNAs that evade the proteostatic response to ER stress and that encode proteins promoting the survival of stressed cells.

Regulation of the HMGA2-SNAI2/CXCR4 axis in atherosclerosis and retinal neovascularization: new therapeutic insights.

Atherosclerosis (AS) is a vascular disease associated with endothelial damage, plaque formation, and retinal neovascularization (RNV), leading to visual impairment. Research indicates that vascular endothelial dysfunction, lipid deposition, and inflammatory responses contribute to the formation of plaques and atherosclerotic lesions. Among the common complications, studies have shown that RNV and the molecular mechanisms of AS hold significant clinical importance. In this study, we identified the overexpression of the gene heat shock protein 90 (HSP90) through transcriptome sequencing. Subsequent protein expression analysis and inhibition experiments in corresponding animal models confirmed the crucial role of HSP90 in the modulation of this disease. Research findings revealed an increase in the expression of HSP90, HMGA2, Snail family transcriptional repressor 2 gene (SNAI2), CXC chemokine receptor 4 (CXCR4), and vascular endothelial growth factor (VEGF) in atherosclerotic mouse tissues. Inhibition of HSP90 expression reduced vascular neovascularization and downregulated the expression of HMGA2 and VEGF. Given that HSP90 can promote HMGA2 expression, which, in turn, facilitates angiogenesis, we conducted lentiviral infection experiments on primary retinal endothelial cells obtained from atherosclerotic mice, confirming the regulatory role of HSP90 in modulating HMGA2 expression through the SNAI2/CXCR4 signaling pathway and its involvement in retinal endothelial neovascularization. In conclusion, our study highlights the significant regulatory role of HSP90 in AS-induced RNV, providing a new target for disease treatment. Furthermore, this research extensively explores the mechanism of HSP90 in regulating RNV and associated signaling pathways, offering novel insights and laying a solid foundation for future studies in this disease domain.

Characterization of the exopolysaccharides produced by the industrial yeast Komagataella phaffii.

The yeast Komagataella phaffii has become a popular host strain among biotechnology startup companies for producing recombinant proteins for food and adult nutrition applications. K. phaffii is a host of choice due to its long history of safe use, open access to protocols and strains, a secretome free of host proteins and proteases, and contract manufacturing organizations with deep knowledge in bioprocess scale-up. However, a recent publication highlighted the abundance of an unknown polysaccharide that accumulates in the supernatant during fermentation. This poses a significant challenge in using K. phaffii as a production host. This polysaccharide leads to difficulties in achieving high purity products and requires specialized and costly downstream processing steps for removal. In this study, we describe the use of the common K. phaffii host strain YB-4290 for production of the bioactive milk protein lactoferrin. Upon purification of lactoferrin using membrane-based separation methods, significant amounts of carbohydrate were co-purified with the protein. It was determined that the carbohydrate is mostly composed of mannose residues with minor amounts of glucose and glucosamine. The polysaccharide fraction has an average molecular weight of 50 kDa and consists mainly of mannan, galactomannan and amylose. In addition, a large fraction of the carbohydrate has an unknown structure likely composed of oligosaccharides. Additional strains were tested in fermentation to further understand the source of the carbohydrates. The commonly used industrial hosts, BG10 and YB-4290, produce a basal level of exopolysaccharide; YB-4290 producing slightly more than BG10. Overexpression of recombinant protein stimulates exopolysaccharide production well above levels produced by the host strains alone. Overall, this study aims to provide a foundation for developing methods to improve the economics of recombinant protein production using K. phaffii as a production host.

Emergence of carbapenem resistance in persistent Shewanella algae bacteremia: the role of pdsS G547W mutation in adaptive subpopulation dynamics

This study elucidates the in vivo genetic mechanisms contributing to the emerging resistance to carbapenem in Shewanella algae through a lens of adaptive microbial evolution. Leveraging PacBio amplification-free sequencing, we tracked the evolution of β-lactam resistance in clinical isolates from a persistent S. algae bacteremia case amidst antimicrobial therapy. Our investigation spotlighted a recurrent G547W mutation in the sensor histidine kinase (pdsS), which was associated with the overexpression of an OmpA-like protein (pdsO) within a proteobacteria-specific sortase system. Intriguingly, we observed a recurrent switch between wild-type and G547W alleles, revealing an adaptive expansion and contraction of underlying cell subpopulations in response to β-lactam exposure. Comparative transcriptome analyses further demonstrated the overexpression of genes pivotal for membrane integrity, biofilm formation, immune evasion, and β-lactamase activation in resistant samples. This underscores the pre-existence of resistant cells at minuscule frequencies even without antibiotic pressure, potentially explaining the within-host emergence of resistance during antibiotic treatments. Our findings provide pivotal insights into the dynamic genetic adaptations of S. algae under therapeutic pressures, unmasking intricate resistance mechanisms and highlighting the critical role of subpopulation dynamics in treatment outcomes.

De novo strategy of organic semiconducting polymer brushes for NIR-II light-triggered carbon monoxide release to boost deep-tissue cancer phototheranostics

The integration of photoacoustic imaging (PAI) and photothermal therapy (PTT) within the second near-infrared (NIR-II) window, offering a combination of high-resolution imaging and precise non-invasive thermal ablation, presents an attractive opportunity for cancer treatment. Despite the significant promise, the development of this noninvasive phototheranostic nanomedicines encounters challenges that stem from tumor thermotolerance and limited therapeutic efficacy. In this contribution, we designed an amphiphilic semiconducting polymer brush (SPB) featuring a thermosensitive carbon monoxide (CO) donor (TDF-CO) for NIR-II PAI-assisted gas-augmented deep-tissue tumor PTT. TDF-CO nanoparticles (NPs) exhibited a powerful photothermal conversion efficiency (43.1%) and the capacity to trigger CO release after NIR-II photoirradiation. Notably, the liberated CO not only acts on mitochondria, leading to mitochondrial dysfunction and promoting cellular apoptosis but also hinders the overexpression of heat shock proteins (HSPs), enhancing the tumor’s thermosensitivity to PTT. This dual action accelerates cellular thermal ablation, achieving a gas-augmented synergistic therapeutic effect in cancer treatment. Intravenous administration of TDF-CO NPs in 4T1 tumor-bearing mice demonstrated bright PAI signals and remarkable tumor ablation under 1064 nm laser irradiation, underscoring the potential of CO-mediated photothermal/gas synergistic therapy. We envision this tailor-made multifunctional NIR-II light-triggered SPB provides a feasible approach to amplify the performance of PTT for advancing future cancer phototheranostics.Graphical abstract

Spatially Controlled MicroRNA Imaging in Mitochondria via Enzymatic Activation of Hybridization Chain Reaction.

Live-cell imaging of RNA in specific subcellular compartments is essential for elucidating the rich repertoire of cellular functions, but it has been limited by a lack of simple, precisely controlled methods. Here such an approach is presented via the combination of hybridization chain reaction and spatially restricted enzymatic activation with organelle-targeted delivery. The system can localize engineered DNA hairpins in the mitochondria, where target RNA-initiated chain reaction of hybridization events is selectively activated by a specific enzyme, enabling amplified RNA imaging with high precision. It is demonstrated that the approach is compatible with live cell visualization and enables the regulatable imaging of microRNA in mitochondria. Since in situ activation of the signal amplification with enzyme eliminates the need for genetically encoded protein overexpression, it is envisioned that this simple platform will be broadly applicable for precise RNA imaging with subcellular resolution in a variety of biological processes.

Overexpression of outer membrane protein A (OmpA) increases aminoglycoside sensitivity in mycobacteria

BackgroundTuberculosis (TB), caused by Mycobacterium tuberculosis (Mtb) complex infection, is a leading cause of death worldwide from a single infectious agent. The emergence of drug resistance Mtb clinical strains makes the situation more serious. The role of Mtb outer membrane protein A (OmpA) in antimicrobial resistance remains unclear. This study aimed to evaluate the effect of OmpA expression on mycobacterial drug resistance. In this study, a Mycobacterium smegmatis (Ms) strain overexpressing OmpA (Ms-OmpA) and a Mycobacterium bovis (Mb) strain overexpressing OmpA (Mb-OmpA) were constructed, and their susceptibility to anti-TB drugs was determined by performing the minimal inhibitory concentrations (MICs), the plate assay and the macrophage infection assays.ResultsThe streptomycin MIC of the overexpressing strain was 2-fold lower than those of the wide-type (Ms) and empty plasmid strains (pMV-261) as well as amikacin and gentamicin. Moreover, both the plate and the macrophage infection assays indicate that overexpression of OmpA increases streptomycin sensitivity in Mycobacteria. The other aminoglycosides like amikacin and gentamicin have the same phenotypes as streptomycin on the plates for the virulent strain Mb-OmpA. The porin inhibitor spermidine can increase streptomycin tolerance in the overexpressing strain, and overexpressing OmpA can increase the intracellular accumulation of hydrophilic ethidium bromide, which indicates that porin protein OmpA contributes to aminoglycosides sensitivity in Mycobacteria.ConclusionsIn this study, we have characterized the contribution of OmpA in the antimicrobial resistance phenotype of Mycobacteria, which may provide valuable insights for understanding antibiotic resistance and designing new strategies for TB treatment.

EXTH-07. UNRAVELING THE ROLE OF SHH SIGNALING PATHWAY IN RETINOBLASTOMA

Abstract Retinoblastoma (RB) is a pediatric cancer affecting the retina, often requiring surgical removal of one or both eyes due to the limited effectiveness of current treatments. There is an urgent need for advanced therapeutic strategies capable of effectively eliminating tumors, thereby obviating the need for surgical removal of eyes in affected children. Achieving this goal requires a deeper understanding of the cellular and molecular mechanisms governing RB initiation and progression. Using a transgenic mouse model, we identified the crucial role of the Shh signaling pathway in Rb1 mutation-induced tumorigenesis, highlighting its potential as a novel therapeutic target for RB treatment. We have observed frequent overexpression of Shh signaling proteins (SHH, GLI1, and GLI2) in human RB, correlating with aggressive clinicopathological features. GLI2 overexpression is necessary for Rb1 deletion-induced RB tumor initiation in mouse retinal progenitor cells (RPCs). These mouse intraocular tumors accurately replicate human RB at both cellular and molecular levels. Additionally, Shh pathway activation extends to normal retinal cells adjacent to human RB tumors, suggesting the involvement of Shh signaling in RB-microenvironment interactions. Importantly, our studies have identified a novel small molecular inhibitor targeting the Shh pathway, effectively inhibiting RB growth in vitro and in vivo by targeting both tumor cells and the tumor microenvironment (TME). The proposed research aims to provide unprecedented insights into the role of SHH signaling pathway in RB initiation and progression and the supporting communication network between RB tumors and TME. These outcomes have the potential to treat RB effectively while improving the well-being of young individuals with RB and those at risk of RB.

Characteristics of stem sings of EpCAM-negative and EpCAM-positive tumor cells in primary tumor and 2D and 3D cultures in breast cancer

One of the predictors of adverse prognosis in breast cancer is the overexpression of the EpCAM protein. However, a signifcant part of tumor cells have low or no EpCAM expression. The molecular features and the ability to grow in 2D and 3D cultures, indirectly refecting metastatic potential of tumor cells without EpCAM expression, have not been sufficiently studied.The aim of the study was to compare phenotypic variants of stem cells and EMT depending on the EpCAM expression in the primary tumor of breast cancer and 2D and 3D cultures.Material and Methods. The study included 7 patients with invasive breast carcinoma of no special type. Luminal A subtype was found in 2/7 patients (29 %), and luminal B HER2-negative molecular subtype was found in 5/7 patients (71 %). The patients didn’t have neoadjuvant chemotherapy. Cells from the primary tumor were cultured in 2D and 3D. The primary tumor cells, 2D and 3D cultures were phenotyped using fow cytometry with antibodies targeting: CD45, EpCAM (CD326), CD44, CD24, N-cadherin (CD325), and CD133, CK7/8, EpCAM (CD326).Results. No significant differences were found in the frequency of detection and the number of cells exhibiting stem cell features among EpCAM- and EpCAM+ tumor cells, 2D and 3D cultures. All phenotypic variants of co-expression of stemness markers CD44, CD24, CD133, and ALDH were present both in the primary tumor and 2D/3D cultures. Stem cells in mammospheres had features of both epithelial and hybrid EMT phenotypes. In primary tumors and 2D/3D cultures, the smallest proportion consisted of stem cells with the CD44+CD24- phenotype and cells co-expressing CD44+CD24- with other stemness markers, while the largest proportion comprised stem cells with ALDH+ and ALDH+CD133+ phenotypes. Marked intra- and inter-tumor heterogeneity in the phenotypic composition of primary tumors, 2D, and 3D cultures was noted.Conclusion. The results indicate that EpCAM-negative and EpCAM-positive tumor cells of luminal molecular subtype of breast cancer are capable of exhibiting various phenotypic variants of stemness and EMT in the primary tumor and 2D and 3D cultures. In most cases, individual tumor cells show co-expression of several stemness markers. The phenotypic composition of primary tumors, 2D and 3D cultures is characterized by pronounced intra- and inter-tumor heterogeneity.

Comparative Proteomic Analysis of Cell Wall Proteins of Aminoglycosides Resistant and Sensitive Mycobacterium tuberculosis Clinical Isolates.

The rising prevalence of Mycobacterium tuberculosis (M.tb) strains resistant to aminoglycosides (amikacin and kanamycin) challenges effective TB control and treatment. Understanding the mechanisms behind this resistance is crucial since aminoglycosides are a mainstay of TB therapy. The study aimed to analyze the cell wall proteins overexpressed in aminoglycoside-resistant isolates of Mycobacterium tuberculosis using proteomics approaches. We used two-dimensional electrophoresis and mass spectrometry to compare the cell wall proteomes of aminoglycosides-resistant and susceptible clinical isolates. The overexpressed protein spots were excised and identified using liquid chromatography-mass spectrometry (LC/MS). The identified proteins were subsequently analyzed for molecular docking, pupylation site identification, and STRING analysis. We found a total of nine significantly upregulated proteins in aminoglycosides-resistant isolates. Three of these proteins were the same (isoform), resulting in the identification of seven unique proteins. Specifically, Rv3841 and Rv1308 belonged to intermediary metabolism and respiration; Rv2115c to the cell wall and cell processes; Rv2501c, Rv2247 and Rv0295c to lipid metabolism; and Rv2416c to virulence, detoxification/adaptation. Notably, variations in these proteins support cell wall integrity, aiding mycobacteria's establishment and proliferation. Molecular docking study revealed that both drugs bind strongly to the proteins' active site regions. Additionally, the GPS-PUP algorithm successfully identified possible pupylation sites within these proteins, except Rv0295c. Based on interactome analysis using the STRING 12.0 database, we have identified potential interactive partners suggesting their role in aminoglycosides resistance. Overexpressed proteins not only act to counteract or regulate drug effects but also have a role in protein dynamics that allow for resistance. Some of these identified proteins may serve as innovative drug targets and biomarkers for the early detection of drug-specific resistance in M.tb. Further research is needed to elucidate the mechanisms by which these potential protein targets contribute to resistance in AK and KM M.tb isolates.

Multi-Omics Approach Reveals Genes and Pathways Affected in Miller-Dieker Syndrome.

Miller-Dieker syndrome (MDS) is a rare neurogenetic disorder resulting from a heterozygous deletion of 26 genes in the MDS locus on human chromosome 17. MDS patients often die in utero and only 10% of those who are born reach 10years of age. Current treatments mostly prevent complications and control seizures. A detailed understanding of the pathogenesis of MDS through gene expression studies would be useful in developing precise medical approaches toward MDS. To better understand MDS at the molecular level, we performed RNA sequencing on RNA and mass spectrometry on total protein isolated from BJ (non-MDS) cells and GM06097 (MDS) cells, which were derived from a healthy individual and an MDS patient, respectively. Differentially expressed genes (DEGs) at the RNA and protein levels involved genes associated with phenotypic features reported in MDS patients (CACNG4, ADD2, SPTAN1, SHANK2), signaling pathways (GABBR2, CAMK2B, TRAM-1), and nervous system development (CAMK2B, BEX1, ARSA). Functional assays validated enhanced calcium signaling, downregulated protein translation, and cell migration defects in MDS. Interestingly, overexpression of methyltransferase-like protein 16 (METTL16), a protein encoded in the MDS locus, restored defects in protein translation, phosphorstates of mTOR (mammalian target of rapamycin) pathway regulators, and cell migration in MDS cells. Although DNA- and RNA-modifying enzymes were among the DEGs and the intracellular SAM/SAH ratio was eightfold lower in MDS cells, global nucleoside modifications remained unchanged. Thus, this study identified specific genes and pathways responsible for the gene expression changes, which could lead to better therapeutics for MDS patients.

Venetoclax in combination with a pediatric-inspired regimen for the treatment of newly diagnosed adults with Philadelphia chromosome-negative acute lymphoblastic leukemia.

BCL-2 protein overexpression, common in B-cell acute lymphoblastic leukemia (B-ALL), including the Philadelphia (Ph)-like subtype, mediates leukemic cell survival. We treated 24 patients with 14 days of BCL-2 inhibitor, venetoclax, 400 mg daily (dose level 1) during induction and consolidation cycles combined with the CALGB 10403 regimen in newly diagnosed adults with Ph-negative B-ALL. Median age was 31 (range: 18-53) years, 92% were Hispanic, and 12 (50%) patients had Ph-like ALL. No dose limiting toxicity occurred in the phase 1 part. Median times to neutrophil and platelet count recovery were 20 and 21 days from start of induction, respectively. The most common grade ≥3 treatment-related adverse events were leukopenia (96%), neutropenia (83%), anemia (83%), thrombocytopenia (79%), lymphopenia (71%), hyperbilirubinemia (38%), and elevated ALT (33%). One patient with non-Ph-like ALL died from asparaginase-associated pancreatitis, and 23 (96%) patients achieved complete remission (CR) or CR with incomplete count recovery (CRi) following induction with or without extended induction phase. Of 22 patients who started consolidation, 20 (91%) achieved negative minimal residual disease status (MRD-) (.

Suppression of CDK1/Drp1-Mediated Mitochondrial Fission Attenuates Dexamethasone-Induced Extracellular Matrix Deposition in the Trabecular Meshwork.

Aims: Deposition of extracellular matrix (ECM) in the trabecular meshwork (TM), as induced by dexamethasone (Dex), is believed to play an important role in the onset of glucocorticoid-induced glaucoma (GIG). Abnormal ECM deposition is a consequence of mitochondrial dysfunction. We aimed to clarify how mitochondrial dysfunction leads to ECM deposition within the TM and to support the development of novel therapeutic strategies. Results: In primary human TM cells (pHTMCs) and a Dex acetate-induced murine model of GIG, glucocorticoid administration stimulated both mitochondrial fission and ECM deposition. Excessive mitochondrial fission leads to dysfunction and the overexpression of ECM proteins in pHTMCs. Notably, when pHTMCs were treated with the dynamin-related protein 1 (Drp1) inhibitor Mdivi-1 or with Drp1 siRNA, we observed a marked reduction in Dex-induced mitochondrial damage and ECM proteins in vitro. Furthermore, in C57BL/6J mice, treatment with Mdivi-1 mitigated mitochondrial damage and blocked ECM deposition within the TM. We then used Ro3306 to inhibit the cyclin-dependent kinase (CDK)1-mediated phosphorylation of Drp1 at Ser 616, which restored mitochondrial function and diminished Dex-induced ECM protein expression in pHTMCs. Innovation: This study illuminates the pathogenic mechanism linking mitochondrial dysfunction to ECM deposition in GIG. Our innovative approach revealed that Dex stimulates mitochondrial fission via CDK1-mediated p-Drp1s616 overexpression, which drives ECM accumulation. It offered a novel therapeutic strategy for reducing ECM protein expression by inhibiting excessive mitochondrial fission and restoring mitochondrial function. Conclusion: By targeting the CDK1/Drp1-driven mitochondrial fission process, we can counteract Dex-induced ECM deposition in the TM both invivo and in vitro.

Role of vitamin D3 in mitigating sodium arsenite-induced neurotoxicity in male rats.

Arsenic is associated with various neurological disorders, notably affecting memory and cognitive functions. The current study examined the protective effects of vitamin D3 (Vit. D3) in countering oxidative stress, neuroinflammation and apoptosis induced by sodium arsenite (SA) in the cerebral cortex of rats. Male Wistar rats were subjected to a daily oral administration of sodium arsenite (NaAsO2, SA) at a dosage of 5mg/kg, along with 500IU/kg of Vit. D3, and a combination of both substances for four weeks. The results indicated that Vit. D3 effectively mitigated the SA-induced increase in oxidative stress markers, thiobarbituric acid reactive substances (TBARS) and nitric oxide (NO), the decrease in antioxidants (reduced glutathione; GSH, superoxide dismutase; SOD, catalase; CAT, and glutathione peroxidase; GPx), as well as the increase in pro-inflammatory markers including, tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and amyloid-beta (Aβ)1-42. Furthermore, Vit. D3 reversed the alterations in the neurochemicals acetylcholinesterase (AchE), monoamine oxidase (MAO), dopamine (DA), and acetylcholine (Ach) and ameliorated the histopathological changes in the cerebral cortex. Moreover, immunohistochemical analyses revealed that Vit. D3 reduced the SA-induced overexpression of cerebral cysteine aspartate-specific protease-3 (caspase-3) and glial fibrillary acidic protein (GFAP) in the cerebral cortex of male rats. Consequently, the co-administration of Vit. D3 can protect the cerebral cortex against SA-induced neurotoxicity, primarily through its antioxidant, anti-inflammatory, anti-apoptotic, and anti-astrogliosis effects.