Nonstructural Protein 1 of SARS-CoV-2 Is a Potent Pathogenicity Factor Redirecting Host Protein Synthesis Machinery toward Viral RNA.

The causative virus of the COVID-19 pandemic, SARS-CoV-2, uses its nonstructural protein 1 (Nsp1) to suppress cellular, but not viral, protein synthesis through yet unknown mechanisms. We show here that among all viral proteins, Nsp1 has the largest impact on host viability in the cells of human lung origin. Differential expression analysis of mRNA-seq data revealed that Nsp1 broadly alters the cellular transcriptome. Our cryo-EM structure of the Nsp1-40S ribosome complex shows that Nsp1 inhibits translation by plugging the mRNA entry channel of the 40S. We also determined the structure of the 48S preinitiation complex formed by Nsp1, 40S, and the cricket paralysis virus internal ribosome entry site (IRES) RNA, which shows that it is nonfunctional because of the incorrect position of the mRNA 3' region. Our results elucidate the mechanism of host translation inhibition by SARS-CoV-2 and advance understanding of the impacts from a major pathogenicity factor of SARS-CoV-2.

Balance between the cell viability and death in 3D.

Cell death is a phenomenon, frequently perceived as an absolute event for cell, tissue and the organ. However, the rising popularity and complexity of such 3D multicellular 'tissue building blocks' as heterocellular spheroids, organoids, and 'assembloids' prompts to revise the definition and quantification of cell viability and death. It raises several questions on the overall viability of all the cells within 3D volume and on choosing the appropriate, continuous, and non-destructive viability assay enabling for a single-cell analysis. In this review, we look at cell viability and cell death modalities with attention to the intrinsic features of such 3D models as spheroids, organoids, and bioprints. Furthermore, we look at emerging and promising methodologies, which can help define and understand the balance between cell viability and death in dynamic and complex 3D environments. We conclude that the recent innovations in biofabrication, biosensor probe development, and fluorescence microscopy can help answer these questions.

The NF-κB/FXR/TonEBP pathway protects renal medullary interstitial cells against hypertonic stress.

Farnesoid X receptor (FXR), a ligand-activated transcription factor, plays an important role in maintaining water homeostasis by up-regulating aquaporin 2 (AQP2) expression in renal medullary collecting ducts; however, its role in the survival of renal medullary interstitial cells (RMICs) under hypertonic conditions remains unclear. We cultured primary mouse RMICs and found that the FXR was expressed constitutively in RMICs, and that its expression was significantly up-regulated at both mRNA and protein levels by hypertonic stress. Using luciferase and ChIP assays, we found a potential binding site of nuclear factor kappa-B (NF-κB) located in the FXR gene promoter which can be bound and activated by NF-κB. Moreover, hypertonic stress-induced cell death in RMICs was significantly attenuated by FXR activation but worsened by FXR inhibition. Furthermore, FXR increased the expression and nuclear translocation of hypertonicity-induced tonicity-responsive enhance-binding protein (TonEBP), the expressions of its downstream target gene sodium myo-inositol transporter (SMIT), and heat shock protein 70 (HSP70). The present study demonstrates that the NF-κB/FXR/TonEBP pathway protects RMICs against hypertonic stress.

m6A methylation of RNF43 inhibits the progression of endometriosis through regulating oxidative phosphorylation via NDUFS1.

Oxidative phosphorylation is becoming increasingly important in the induction and development of endometriosis. Recently, it has been reported that ring finger protein 43 (RNF43) is involved in the process of oxidative phosphorylation, but the mechanism remains unclear. Our investigation is to delve into the roles of RNF43 in endometriosis and elucidate the related mechanisms. We found RNF43 was downregulated in ectopic endometrial tissue and primary ectopic endometrial stromal cells (ECESCs). Knockdown of RNF43 enhanced cell viability and migration by activating oxidative phosphorylation in eutopic endometrial stromal cells (EUESCs), while overexpression of RNF43 led to the opposite results. Moreover, RNF43 reinforced the ubiquitination and degradation of NADH dehydrogenase Fe-S protein 1 (NDUFS1) by interacting with it. Likewise to RNF43 overexpression, NDUFS1 silencing inhibited cell viability, migration, and oxidative phosphorylation in ECESCs. NDUFS1 was a downstream target of RNF43, mediating its biological role in endometriosis. Interestingly, the expression and stability of RNF43 mRNA were regulated by the Methyltransferase-like 3 (METTL3)/IGF2BP2 m6A modification axis. The results of rat experiments showed decreased RNF43 expression and increased NDUFS1 expression in endometriosis rats, which was enhanced by METTL3 inhibition. Those observations indicated that m6A methylation-mediated RNF43 negatively affects viability and migration of endometrial stromal cells through regulating oxidative phosphorylation via NDUFS1. The discovery of METTL3/RNF43/NDUFS1 axis suggested promising therapeutic targets for endometriosis.

Identification of the Candidate mGlu2 Allosteric Modulator THRX-195518 through In Silico Method and Evaluation of Its Neuroprotective Potential against Glutamate-Induced Neurotoxicity in SH-SY5Y Cell Line.

Glutamate (Glu) toxicity has been an important research topic in toxicology and neuroscience studies. In vitro and in vivo studies have shown that Group II metabotropic Glu2 (mGlu2) activators have cell viability effects. This study aims to determine a candidate ligand with high mGlu2 allosteric region activity among cytotoxicity-safe molecules using the in silico positioning method and to evaluate its cell viability effect in vitro. We investigated the candidate molecule's cell viability effect on the SH-SY5Y human neuroblastoma cell line by MTT analysis. In the study, LY 379268 (agonist) and JNJ-46281222 (positive allosteric modulator; PAM) were used as control reference molecules. Drug bank screening yielded THRX-195518 (docking score being -12.4 kcal/mol) as a potential novel drug candidate that has a high docking score and has not been mentioned in the literature so far. The orthosteric agonist LY 379268 exhibited a robust protective effect in our study. Additionally, our findings demonstrate that JNJ-46281222 and THRX-195518, identified as activating the mGlu2 allosteric region through in silico methods, preserve cell viability against Glu toxicity. Therefore, our study not only emphasizes the positive effects of this compound on cell viability against Glu toxicity but also sheds light on the potential of THRX-195518, acting as a mGlu2 PAM, based on in silico absorption, distribution, metabolism, excretion, and toxicity (ADMET) data, as a candidate drug molecule. These findings underscore the potential utility of THRX-195518 against both neurotoxicity and Central Nervous System (CNS) disorders, providing valuable insights.

Effects of free fatty acid receptor (FFAR) signaling on the modulation of cancer cell functions under hypoxic conditions.

In the tumor environment, hypoxia promotes tumor progression, such as cancer cell growth, migration and chemoresistance. This study aimed to evaluate the roles of free fatty acid receptors (FFARs) in the regulation of cancer cell functions under hypoxic conditions, using fibrosarcoma HT1080 cells. HT1080 cells expressed FFAR1, FFAR2 and FFAR3 genes, but not FFAR4 gene. FFAR1, FFAR2 and FFAR3 expression levels in HT1080 cells cultured at 1 % O2 were elevated, compared with 21 % O2. The cell growth activities of HT1080 cells cultured at 21 % O2 were inhibited by acetic acid (AA) and propanoic acid (PA), but not 1 % O2. HT1080 cell motility was markedly reduced by culturing at 1 % O2. The cell growth and motility of HT1080 cells were enhanced by FFAR2 knockdown. The cell viability to cisplatin (CDDP) of HT1080 cells cultured at 1 % O2 was increased, compared with 21 % O2. FFAR2 knockdown suppressed the cell viability to CDDP of HT1080 cells. On the other hand, the cell motility and viability to CDDP of HT1080 cells cultured at 21 % O2 were suppressed by TUG-770. When HT1080 cells were cultured at 1 % O2, the cell motility and viability to CDDP were decreased, correlating with FFAR1 expression level. Moreover, FFAR1 knockdown increased the cell viability to CDDP of HT1080 cells cultured at 1 % O2. These results suggest that FFAR-mediated signaling plays an important role in the modulation of cellular functions of HT1080 cells under hypoxic conditions.

Electrofluidic control for textile-based cell culture: Identification of appropriate conditions required to integrate cell culture with electrofluidics.

Electric field-driven microfluidics, known as electrofluidics, is a novel attractive analytical tool when it is integrated with low-cost textile substrate. Textile-based electrofluidics, primarily explored on yarn substrates, is in its early stages, with few studies on 3D structures. Further, textile structures have rarely been used in cellular analysis as a low-cost alternative. Herein, we investigated novel 3D textile structures and develop optimal electrophoretic designs and conditions that are favourable for direct 3D cell culture integration, developing an integrated cell culture textile-based electrofluidic platform that was optimised to balance electrokinetic performance and cell viability requirements. Significantly, there were contrasting electrolyte compositional conditions that were required to satisfy cell viability and electrophoretic mobility requiring the development of and electrolyte that satisfied the minimum requirements of both these components within the one platform. Human dermal fibroblast cell cultures were successfully integrated with gelatine methacryloyl (GelMA) hydrogel-coated electrofluidic platform and studied under different electric fields using 5 mM TRIS/HEPES/300 mM glucose. Higher analyte mobility was observed on 2.5% GelMA-coated textile which also facilitated excellent cell attachment, viability and proliferation. Cell viability also increased by decreasing the magnitude and time duration of applied electric field with good cell viability at field of up to 20 V cm-1.

Adaptive resistance of tumor cells to anti-vascular endothelial growth factor therapy: A reversible phenomenon.

Vascular endothelial growth factor (VEGF) inhibition is an essential targeted strategy for malignant tumors, but its efficacy is severely constrained by drug resistance. The traditional view holds that the target of VEGF inhibition is endothelial cells, and thus compensatory angiogenesis is considered the main mechanism of drug resistance. In this study, we found that tumor cells themselves could develop acquired resistance to VEGF therapy, indicating an independent resistance mechanism apart from angiogenesis. Notably, this acquired resistance was temporary, disappearing completely four days after discontinuing exposure to the drug in vitro. Our findings suggest that tumor cells may also be targets of VEGF inhibition, and their response to treatment should not be overlooked in contributing to drug resistance.

Associations of motility and auto-aggregation with biofilm-formation capacity levels in Clostridioidesdifficile.

Clostridioides difficile (C. difficile) is responsible for one of the most common nosocomial infections worldwide. This work assessed associations between biofilm-formation capacity levels of C. difficile and cell viability, motility, flagella, motility and auto-aggregation in 118 clinical isolates. Biofilm production was assessed by the crystal violet method. Cell viability was determined by BacTiter-Glo™ Microbial Cell Viability Assay and live-imaging microscopy. Expression levels of LuxS, Cwp84, Spo0A, PilA, and FliC were measured by real-time PCR. Motility was visually assessed in agar tubes. Auto-aggregation levels were determined by OD600 measurements. Out of 118 isolates, 66 (56 %) were biofilm producers, with most being strong or moderate producers. Cell viability, motility and auto-aggregation positively correlated with biofilm-production capacity (p = 0.0001, p = 0.036 and p < 0.0001, respectively). Positive associations were found between pilA, fliC and luxS expression levels and biofilm-production capacity (p = 0.04, p = 0.01, p = 0.036, respectively). This is the first report of associations between biofilm-formation capacity and cell viability, pilA, fliC, and luxS gene expression, auto-aggregation and motility. These correlations should be further explored to expand knowledge on the regulation of C. difficile biofilm formation, and pathogenesis, which will have notable implications on treatment options.

Astragaloside IV inhibits cell viability and glycolysis of hepatocellular carcinoma by regulating KAT2A-mediated succinylation of PGAM1.

BACKGROUND: Astragaloside IV (AS-IV) is one of the basic components of Astragali radix, that has been shown to have preventive effects against various diseases, including cancers. This study aimed to explore the role of AS-IV in hepatocellular carcinoma (HCC) and its underlying mechanism. METHODS: The cell viability, glucose consumption, lactate production, and extracellular acidification rate (ECAR) in SNU-182 and Huh7 cell lines were detected by specific commercial kits. Western blot was performed to analyze the succinylation level in SNU-182 and Huh7 cell lines. The interaction between lysine acetyltransferase (KAT) 2 A and phosphoglycerate mutase 1 (PGAM1) was evaluated by co-immunoprecipitation and immunofluorescence assays. The role of KAT2A in vivo was explored using a xenografted tumor model. RESULTS: The results indicated that AS-IV treatment downregulated the protein levels of succinylation and KAT2A in SNU-182 and Huh7 cell lines. The cell viability, glucose consumption, lactate production, ECAR, and succinylation levels were decreased in AS-IV-treated SNU-182 and Huh7 cell lines, and the results were reversed after KAT2A overexpression. KAT2A interacted with PGAM1 to promote the succinylation of PGAM1 at K161 site. KAT2A overexpression promoted the viability and glycolysis of SNU-182 and Huh7 cell lines, which were partly blocked following PGAM1 inhibition. In tumor-bearing mice, AS-IV suppressed tumor growth though inhibiting KAT2A-mediated succinylation of PGAM1. CONCLUSION: AS-IV inhibited cell viability and glycolysis in HCC by regulating KAT2A-mediated succinylation of PGAM1, suggesting that AS-IV might be a potential and suitable therapeutic agent for treating HCC.

Evidence of the protective role of Carvacrol in a retinal degeneration animal model.

Neurodegenerative pathologies affecting the posterior segment of the eye, are characterized by being devastating and responsible for the majority of visual dysfunctions worldwide. These diseases are primarily degenerative, progressing chronically, and can inflict gradual harm to the optic nerve, retinal ganglion cells (RGC), photoreceptors, and other retinal cells. This retinal damage leads to a progressive loss of vision, marking these conditions as a significant health concern worldwide. The intravitreal administration of the phytochemical Carvacrol (CAR) is expected to demonstrate a neuroprotective and antiapoptotic effect on retinal cells, with a specific focus on RGC. This effect will be observed in a retinal degeneration model (RDM) in rabbits induced by cytotoxic and oxidative agents, namely glutamate (GLUT) and L-buthionine-S, R-sulfoximine (BSO). An in vivo study was conducted using New Zealand rabbits in which retinal damage was created to evaluate the effectiveness of CAR. The effectiveness of CAR on the functionality of retinal neuronal cells in RDM was evaluated using pupillary light reflection (PLR). Furthermore, the phytotherapeutic's influence on cell viability was determined through flow cytometry analysis. Finally, the neuroprotective and antiapoptotic capabilities of CAR were specifically scrutinized in RGC through histological studies, quantifying cell survival, and employing immunohistochemical assays to detect the apoptotic index (%) using the TUNEL technique. Our results demonstrated that CAR promoted the recovery of the pupillary contraction profile over time, maintaining the functionality of retinal cells as healthy controls. Additionally, it showed increased cell viability under oxidative and cytotoxic conditions given by GLUT-BSO agents. Finally, we found that CAR protects the survival of RGC and decreases the percentage of apoptotic cells when compared to RDM. CAR demonstrated to have positive effects on the functionality of photoreceptive nerve cells by restoring pupillary contraction. Likewise, it was shown to have neuroprotective and antiapoptotic effects when evaluated in a general and specific way on retinal nerve cells.

Histone lactylation regulates autophagy of hyperplastic scar fibroblasts by inhibiting the transcriptional activity of phosphatase and tensin homologue.

Hyperplastic scar (HS) is an overreaction of tissue to skin injury caused by local fibroblast proliferation and excessive collagen production. Histone posttranslational modification patterns are important epigenetic processes that control various biological activities. This study was designed to investigate the effects of histone lactylation on HS and the underlying mechanism. Western blot was used to analyse the lactylation level in HS patients and fibroblasts (HSFs). In vitro experiments, western blot, cell counting kit-8, and immunofluorescence staining were performed to detect the collagen level, cell viability, and autophagy, respectively. The relationship between snai2 (SLUG) and phosphatase and tensin homologue (PTEN) was assessed by RNA immunoprecipitation and dual-luciferase reporter assays. The results showed that the histone lactylation level was upregulated in HS tissues and HSFs. HSFs showed increased collagen production and cell viability, and decreased autophagy. Silencing of lactate dehydrogenase A (LDHA) promoted the transcription of PTEN by inhibiting SLUG, thus promoting autophagy. Knockdown of LDHA inhibited collagen deposition and cell viability, and increased autophagy in HSFs, and the results were reversed after PTEN inhibition. In summary, histone lactylation inhibited the transcription activity of PTEN by promoting SLUG, thereby suppressing autophagy and promoting collagen deposition and cell viability of HSFs, which might provide effective therapeutic strategies in HS.

Synthesis of alga-coated copper oxide nanoparticles with potential applications in shrimp farming.

Copper (Cu) is a crucial element that plays a vital role in facilitating proper biological activities in living organisms. In this study, copper oxide nanoparticles (CuO NPs) were synthesized using a straightforward precipitation chemical method from a copper nitrate precursor at a temperature of 85 °C. Subsequently, these NPs were coated with the aqueous extract of Sargassum angustifolium algae. The size, morphology, and coating of the NPs were analyzed through various methods, revealing dimensions of approximately 50 nm, a multidimensional shaped structure, and successful algae coating. The antibacterial activity of both coated and uncoated CuO NPs against Vibrio harveyi, a significant pathogen in Litopenaeus vannamei, was investigated. Results indicated that the minimum inhibitory concentration (MIC) for uncoated CuO NPs was 1000 µg/mL, whereas for coated CuO NPs, it was 500 µg/mL. Moreover, the antioxidant activity of the synthesized NPs was assessed. Interestingly, uncoated CuO NPs exhibited superior antioxidant activity (IC50 ≥ 16 µg/mL). The study also explored the cytotoxicity of different concentrations (10-100 µg/mL) of both coated and uncoated CuO NPs. Following 48 h of incubation, cell viability assays on shrimp hemocytes and human lymphocytes were conducted. The findings indicated that CuO NPs coated with alga extract at a concentration of 10 µg/mL increased shrimp hemocyte viability. In contrast, uncoated CuO NPs at a concentration of 25 µg/mL and higher, as well as CuO NPs at a concentration of 50 µg/mL and higher, led to a decrease in shrimp hemocyte survival. Notably, this study represents the first quantitative assessment of the toxicity of CuO NPs on shrimp cells, allowing for a comparative analysis with human cells.

Hyaluronic acid enhances cell migration, viability, and mineralized tissue-specific genes in cementoblasts.

BACKGROUND/OBJECTIVES: It has been repeatedly demonstrated that cementum formation is a crucial step in periodontal regeneration. Hyaluronic acid (HA) is an important component of the extracellular matrix which regulates cells functions and cell-cell communication. Hyaluronic acid/derivatives have been used in regenerative periodontal therapy, but the cellular effects of HA are still unknown. To investigate the effects of HA on cementoblast functions, cell viability, migration, mineralization, differentiation, and mineralized tissue-associated genes and cementoblast-specific markers of the cementoblasts were tested. MATERIALS AND METHODS: Cementoblasts (OCCM-30) were treated with various dilutions (0, 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, 1:128) of HA and examined for cell viability, migration, mineralization, and gene expressions. The mRNA expressions of osteocalcin (OCN), runt-related transcription factor 2 (Runx2), bone sialoprotein (BSP), collagen type I (COL-I), alkaline phosphatase (ALP), cementum protein-1 (CEMP-1), cementum attachment protein (CAP), and small mothers against decapentaplegic (Smad) -1, 2, 3, 6, 7, ß-catenin (Ctnnb1) were performed with real-time polymerase chain reaction (RT-PCR). Total RNA was isolated on days 3 and 8, and cell viability was determined using MTT assay on days 1 and 3. The cell mineralization was evaluated by von Kossa staining on day 8. Cell migration was assessed 2, 4, 6, and 24 hours following exposure to HA dilutions using an in vitro wound healing assay (0, 1:2, 1:4, 1:8). RESULTS: At dilution of 1:2 to 1:128, HA importantly increased cell viability (p < .01). HA at a dilution of 1/2 increased wound healing rates after 4 h compared to the other dilutions and the untreated control group. Increased numbers of mineralized nodules were determined at dilutions of 1:2, 1:4, and 1:8 compared with control group. mRNA expressions of mineralized tissue marker including COL-I, BSP, RunX2, ALP, and OCN significantly improved by HA treatments compared with control group both on 3 days and on 8 days (p < .01). Smad 2, Smad 3, Smad 7, and ß-catenin (Ctnnb1) mRNAs were up-regulated, while Smad1 and Smad 6 were not affected by HA administration. Additionally, HA at dilutions of 1:2, 1:4, and 1:8 remarkably enhanced CEMP-1 and CAP expressions in a dilution- and time-dependent manner (p < .01). CONCLUSIONS: The present results have demonstrated that HA affected the expression of both mineralized tissue markers and cementoblast-specific genes. Positive effects of HA on the cementoblast functions demonstrated that HA application may play a key role in cementum regeneration.

CDKN2A promoter methylation enhances self-renewal of glioblastoma stem cells and confers resistance to carmustine.

BACKGROUND: Glioblastoma, a highly aggressive form of brain cancer, poses significant challenges due to its resistance to therapy and high recurrence rates. This study aimed to investigate the expression and functional implications of CDKN2A, a key tumor suppressor gene, in glioblastoma cells, building upon the existing background of knowledge in this field. METHOD: Quantitative reverse transcription PCR (qRT-PCR) analysis was performed to evaluate CDKN2A expression in U87 glioblastoma cells compared to normal human astrocytes (NHA). CDKN2A expression levels were manipulated using small interfering RNA (siRNA) and CDKN2A overexpression vector. Cell viability assays and carmustine sensitivity tests were conducted to assess the impact of CDKN2A modulation on glioblastoma cell viability and drug response. Sphere formation assays and western blot analysis were performed to investigate the role of CDKN2A in glioblastoma stem cell (GSC) self-renewal and pluripotency marker expression. Additionally, methylation-specific PCR (MSP) assays and demethylation treatment were employed to elucidate the mechanism of CDKN2A downregulation in U87 cells. RESULT: CDKN2A expression was significantly reduced in glioblastoma cells compared to NHA. CDKN2A overexpression resulted in decreased cell viability and enhanced sensitivity to carmustine treatment. CDKN2A inhibition promoted self-renewal capacity and increased pluripotency marker expression in U87 cells. CDKN2A upregulation led to elevated protein levels of p16INK4a, p14ARF, P53, and P21, which are involved in cell cycle regulation. CDKN2A downregulation in U87 cells was associated with high promoter methylation, which was reversed by treatment with a demethylating agent. CONCLUSION: Our findings demonstrate that CDKN2A downregulation in glioblastoma cells is associated with decreased cell viability, enhanced drug resistance, increased self-renewal capacity, and altered expression of pluripotency markers. The observed CDKN2A expression changes are mediated by promoter methylation. These results highlight the potential role of CDKN2A as a therapeutic target and prognostic marker in glioblastoma.

Comparative analysis of sorafenib and lenvatinib on HepG2 cells and human umbilical vein endothelial cells: Involvement of transforming growth factor-ß signaling in their molecular effects.

AIM: This study aimed to compare the effects of the molecular targeted drugs, sorafenib and lenvatinib, on the survival, invasion, and angiogenesis of hepatocellular carcinoma cells. Additionally, we investigated the involvement of transforming growth factor beta (TGF-ß) signaling in their molecular mechanisms. METHODS: To investigate the effects of sorafenib and lenvatinib, we conducted cell viability, invasion, and angiogenesis assays, as well as western blotting analyses. RESULTS: In human hepatocellular carcinoma cells (HepG2), sorafenib demonstrated potent inhibitory effects on cell proliferation, but induced cell invasion similar to TGF-ß. In contrast, lenvatinib showed weaker cytotoxicity compared with sorafenib, but suppressed cell invasion induced by TGF-ß. The actions of these two molecular targeted drugs were suggested to involve the regulation of the TGFßR2/ERK pathway. Moreover, in human umbilical vein endothelial cells, Sorafenib showed weaker cytotoxicity and enhanced the effects of TGF-ß on angiogenesis. Conversely, lenvatinib showed potent cytotoxic abilities and suppressed angiogenesis induced by TGF-ß. The actions of these two molecular targeted drugs were suggested to involve the regulation of the crosstalk between TGF-ß signaling and vascular endothelial growth factor signaling. CONCLUSIONS: Our findings indicate that both sorafenib and lenvatinib possess anticancer abilities by inducing the cytotoxicity of hepatocellular carcinoma cells. Furthermore, they show opposing effects on TGF-ß-induced cell invasion and angiogenesis, thereby enhancing the understanding of the multifaceted functions of molecular targeted drugs in treating hepatocellular carcinoma.

FZD3 regulates the viability, apoptosis, and extracellular matrix degradation of vaginal wall fibroblasts in pelvic organ prolapse via the Wnt signaling pathway.

The occurrence of pelvic organ prolapse (POP) seriously affects women's quality of life. However, the pathogenesis of POP remains unclear. We aimed to clarify the role of Frizzled class receptor 3 (FZD3) in POP. FZD3 expression in the vaginal wall tissues was detected using immunohistochemistry, real-time polymerase chain reaction, and western blot analysis. Then, vaginal wall fibroblasts (VWFs) were isolated from patients with POP and non-POP, and were identified. Cell viability and apoptosis were evaluated using Cell Counting Kit-8 and flow cytometry, respectively. Extracellular matrix (ECM) degradation was assessed by western blot analysis. The results illustrated that FZD3 was downregulated in POP. VWFs from POP had lower cell viability, ECM degradation, and higher apoptosis. Knockdown of FZD3 inhibited cell viability, ECM degradation, and promoted apoptosis of VWFs, whereas overexpression of FZD3 had opposite results. Moreover, IWP-4 (Wingless-type [Wnt] pathway inhibitor) reversed the role of FZD3 overexpression on biological behaviors. Taken together, FZD3 facilitates VWFs viability, ECM degradation, and inhibits apoptosis via the Wnt pathway in POP. The findings provide a potential target for the treatment of POP.

A comprehensive review on anticancer evaluation techniques.

The development of effective anticancer strategies and the improvement of our understanding of cancer need analytical tools. Utilizing a variety of analytical approaches while investigating anti-cancer medicines gives us a thorough understanding of the traits and mechanisms concerned to cancer cells, which enables us to develop potent treatments to combat them. The importance of anticancer research may be attributed to various analytical techniques that contributes to the identification of therapeutic targets and the assessment of medication efficacy, which are crucial things in expanding our understanding of cancer biology. The study looks at methods that are often used in cancer research, including cell viability assays, clonogenic assay, flow cytometry, 2D electrophoresis, microarray, immunofluorescence, western blot caspase activation assay, bioinformatics, etc. The fundamentals, applications, and how each technique analytical advances our understanding of cancer are briefly reviewed.

Cefixime loaded bare and functionalized halloysite nanocarriers and their biomedical applications.

Effective drug delivery for is the foremost requirement for the complete recovery of the disease. Nanomedicine and nanoengineering has provided so many spaces and ideas for the drug delivery design, whether controlled, targeted, or sustained. Different types of nanocarriers or nanoparticles are aggressively designed for the drug delivery applications. Clay minerals are identified as a one of the potential nanocarrier for the drug delivery. Owing to their biocompatibility and very low cytotoxicity, clay minerals showing effective therapeutic applications. In the present investigation, clay mineral, i.e., Halloysite nano tubes are utilized as a nanocarrier for the delivery of antibiotic cefixime (CFX), a third-generation cephalosporin. The HNT was first functionalized with the sulfuric acid and then further treated with the 3-(aminopropyl)triethoxysilane (APTES). The drug is loaded on three different classifications of HNTs, i.e., Bare-CFX-HNT, Acid-CFX-HNT, and APTES-CFX-HNT and their comparative analysis is established. Different characterization techniques such as X-ray diffractometry (XRD), Fourier transform infra-red (FT-IR), Transmission electron microscopy TEM), Brunauer-Emmett-Teller (BET), adsorption studies, and Thermogravimetric analysis (TGA) were performed to evaluate their chemical, structural, morphological, and thermal properties. TGA confirmed the encapsulation efficiency of Bare-CFX-HNT, Acid-CFX-HNT, and APTES-CFX-HNT as 42.65, 52.19, and 53.43%, respectively. Disk diffusion and MTT assay confirmed that the drug loaded HNTs have potential antibacterial activities and less cytotoxicity. The adsorption capacity of CFX with different HNTs are evaluated and Different adsorption and kinetic models have been discussed. Drug release studies shows that APTES-CFX-HNT showing sustained release of cefixime as compared to Bare-CFX-HNT and Acid-CFX-HNT.

TAPI-1 Exhibits Anti-tumor Efficacy in Human Esophageal Squamous Cell Carcinoma Cells via Suppression of NF-κB Signaling Pathway.

BACKGROUND: TNF-α processing inhibitor-1 (TAPI-1) is a known metalloproteinase inhibitor with potential anti-inflammatory effects. However, its anti-cancer effects on esophageal squamous cell carcinoma (ESCC) have not been uncovered. AIM: In the present study, the effects of TAPI-1 on ESCC cell viability, migration, invasion, and cisplatin resistance and the underlying molecular mechanisms were investigated in TE-1 and Eca109 cells. METHODS: To this end, TE-1 and Eca109 cells were exposed to TAPI-1 for indicated time intervals. Cell viability was assessed using cell counting kit-8 assay and apoptosis was evaluated using flow cytometry assay. Migration and invasion were assessed using Transwell assays. Gene expressions were analyzed using quantitative reverse transcription polymerase chain reaction. The activation of NF-κB signaling pathway was elucidated via Western blot and chromatin immunoprecipitation assay. RESULTS: We observed that higher doses (10, 20 µM) of TAPI-1 inhibited ESCC cell viability, while a lower dose (5 µM) of TAPI-1 inhibited ESCC cell migration and invasion and enhanced the chemosensitivity of ESCC cells to cisplatin. Moreover, TAPI-1 suppressed the activation of NF-κB signaling and the target genes expression in the stage of transcription initiation. Furthermore, blocking NF-κB signaling in advance could abolish all the effects of TAPI-1 on ESCC cells. CONCLUSION: Overall, these results indicated that TAPI-1 impairs ESCC cell viability, migration, and invasion and facilitates cisplatin-induced apoptosis via suppression of NF-κB signaling pathway. TAPI-1 may serve as a potential adjuvant agent with cisplatin for ESCC therapy.