Expression Levels Of P53 Research Articles

Structural simplification of Osimertinib to elaborate new indolyle-pyrimidine-5-carbonitrile derivatives with Anti-proliferative and Anti-SARS-CoV-2 activities assisted by molecular dynamic simulation

Based on the simplicity and modification of the medication osimertinib, two new series of indolyle-pyrimidine-5-carbonitrile scaffolds were created and synthesized with dual action as anti-SARS-Cov-2 and anticancer. The newly created heterocyclic compounds' chemical structures were effectively characterized. With IC50 values of 18.52, 20.89, and 19.85, respectively, compounds 9b, 10, and 15 had inhibitory actions against SARS-CoV-2 when compared to remdesivir and chloroquine, which served as pharmacological controls and had IC50 values of 3.38 μM and 24.9 μM, respectively.Furthermore, compounds 9a and 13 showed anti-proliferative activity against HepG2 cell lines with IC50s of 5.63 μM and 3.06 μM, respectively, in comparison to doxorubicin's IC50 of 7.4 μM. qRT-PCR revealed that HepG2 cells treated with compounds 9a and 13 showed increased p53 expression levels and decreased CDK1 and PI3K expression levels in comparison to doxorubicin. Molecular dynamics simulations of 20 ns duration were performed with PI3Kα, PI3Kγ, and CDK and active compound complexes. The results confirm that compound 13 has the potential to be a therapeutic candidate for additional preclinical and clinical research, as indicated by the molecular docking data.

Abstract A069: Development of an AAV-mediated GEM model to investigate the impact of age on pancreatic tumorigenesis

Abstract Much of our understanding of PDAC initiation comes from Genetically Engineered Mouse (GEM) models where a pancreas-specific Cre (such as Pdx- or p48-Cre) drives expression of specific mutations. While these GEM models generally recapitulate the key histopathological features of human PDAC, they have limitations when it comes to studying the impact of age on PDAC initiation since the Cre drivers induce recombination at the embryonic stage. This is an important consideration since in humans the disease manifests later in life, with the median age at diagnosis being 70 years old. Here, we describe the development of an intrapancreatic injection model using adeno-associated virus (AAV) to study how age influences PDAC initiation and the dynamics of tumor progression. This new viral transduction-based mouse model employs AAV-Cre to induce expression of an oncogenic KRasG12D allele together with a Trp53R172H tumor suppressor mutation within the pancreata of mice at different ages. Our approach utilizes mice with the genotype LSL-KrasG12D; LSL-Trp53R172H that are injected with different AAV-Cre directly into the parenchyma of the pancreas. In our initial studies, we used rAAV vectors expressing Cre recombinase and EGFP (AAV-Cre-EGFP) into young mice (10-14 weeks old) and older mice (32 weeks old) and analyzed EGFP expression using IVIS imaging and immunohistochemistry (IHC) at various timepoints post-injection. To evaluate tumor initiation, we assessed the extent of PanIN lesion formation using H&E staining. Our preliminary findings indicate that AAV-Cre-EGFP effectively transduces cells in vivo and triggers tumor development in the pancreas. To account for potential immunogenicity of the fluorescent protein, we then used rAAV vectors lacking EGFP expression. As controls, we administered identical rAAV vectors to wild-type cohorts matched for age and sex, along with mock injections. Transduction efficiency within the pancreas was determined by evaluating Cre and p53 expression levels, given that the R172H mutation stabilizes the protein. Notably, we found that EGFP is not necessary for either successful transduction or tumor initiation. We are currently evaluating PDAC initiation using a wide spectrum of ages, but our preliminary data using this model suggest that increased age may impact tumor progression, and that this effect may be sex-specific. Our novel AAV-Cre injection model holds promise for investigating age-related aspects of PDAC initiation, and in the future, such approaches could be leveraged to investigate how age impacts the effectiveness of different therapeutic modalities. Citation Format: Karen XD Duong-Polk, Madelaine Neff, Cosimo Commisso. Development of an AAV-mediated GEM model to investigate the impact of age on pancreatic tumorigenesis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A069.

Resveratrol inhibits pancreatic cancer proliferation and metastasis by depleting senescent tumor-associated fibroblasts.

Pancreatic cancer, a formidable gastrointestinal neoplasm, is characterized by its insidious onset, rapid progression, and resistance to treatment, which often lead to a grim prognosis. While the complex pathogenesis of pancreatic cancer is well recognized, recent attention has focused on the oncogenic roles of senescent tumor-associated fibroblasts. However, their precise role in pancreatic cancer remains unknown. Resveratrol is a natural polyphenol known for its multifaceted biological actions, including antioxidative and neuroprotective properties, as well as its potential to inhibit tumor proliferation and migration. Our current investigation builds on prior research and reveals the remarkable ability of resveratrol to inhibit pancreatic cancer proliferation and metastasis. To explore the potential of resveratrol in inhibiting pancreatic cancer by targeting senescent tumor-associated fibroblasts. Immunofluorescence staining of pancreatic cancer tissues revealed prominent coexpression of α-SMA and p16. HP-1 expression was determined using immunohistochemistry. Cells were treated with the senescence-inducing factors known as 3CKs. Long-term growth assays confirmed that 3CKs significantly decreased the CAF growth rate. Western blotting was conducted to assess the expression levels of p16 and p21. Immunofluorescence was performed to assess LaminB1 expression. Quantitative real-time polymerase chain reaction was used to measure the levels of several senescence-associated secretory phenotype factors, including IL-4, IL-6, IL-8, IL-13, MMP-2, MMP-9, CXCL1, and CXCL12. A scratch assay was used to assess the migratory capacity of the cells, whereas Transwell assays were used to evaluate their invasive potential. Specifically, we identified the presence of senescent tumor-associated fibroblasts within pancreatic cancer tissues, linking their abundance to cancer progression. Intriguingly, Resveratrol effectively eradicated these fibroblasts and hindered their senescence, which consequently impeded pancreatic cancer progression. This groundbreaking discovery reinforces Resveratrol's stature as a potential antitumor agent and positions senescent tumor-associated fibroblasts as pivotal contenders in future therapeutic strategies against pancreatic cancer.

Clinical, endoscopic, laboratory and immunomorphological parameters in predicting the occurrence of colorectal cancer in patients with diverticular disease of the colon

To define the role of clinical, endoscopic, laboratory and immunomorphological parameters in predicting the occurrence and course of colorectal cancer (CRC) in patients with diverticular disease of the colon (DDC). One hundred and seventy five people with DDC were examined, divided into 3 groups: group 1 - 85 patients with DDC; group 2 - 45 with DDC in combination with adenomatous polyps (AP); 3rd group - 45 with DDC with established CRC (I or II stage). The control group consisted of 30 practically healthy people. Patients and healthy people were examined according to a single program: clinical, laboratory, instrumental data and immunomorphological research methods [use of primary antibodies to p53 (mAb clone DO-7 product no. PA0057, Leica Biosystems, Leica Bond III) and Ki-67 (Ab16667, Abcam, UK)]. Among the main complaints in patients with DDC and CRC, constipation was more common than in patients with DDC and DDC with AP (p<0.05). In patients with DDC and colorectal neoplasia, a positive reaction to occult blood in the feces was more often verified, compared with the group with DDC (p<0.05). Higher levels of glucose and cholesterol in blood plasma, as well as body mass index were found in patients with DDC with AP and CRC, compared with the DDC group (p<0.05). A higher level of expression of Ki-67 and p53 was found in patients with DDC combined with AP and CRC, compared with patients with DDC without colorectal neoplasia (p<0.05). At the same time, in patients with DDC with CRC, the expression level of Ki-67 and p53 was higher than in patients with DDC with AP (p<0.05) Conclusion. In patients with DDC combined with AP and CRC, higher levels of glucose, plasma cholesterol, as well as body mass index were observed compared to the group of patients with DDC alone (p<0.05). Of note, the results of the determination of Ki-67 and p53 in the mucous membrane of the colon should be considered important prognostic markers for the development of CRC in patients with DDC.

Inhibitory Effects of Royal Jelly and Its Functional Components on the Proliferation of MKN-28 Gastric Cancer Cells.

Royal jelly (RJ) is a natural food product with nutritional value and anticancer activity. However, their effects on gastric cancer are unclear. Here, we show that treatment with 5-320 μg/mL of RJ, ethanol extract (RJEE), and protein hydrolyzate (RJPH) decreased the viability of MKN-28 gastric cancer cells, with a half-maximal inhibitory concentration of 123.22 μg/mL for RJEE. RJ, RJEE, and RJPH increase the lactate dehydrogenase release rate and change the morphology of the cells, resulting in cell shrinkage, nucleoplasm condensation, and the formation of apoptotic bodies. RJ and its functional components stagnated the cell cycle in the G0/G1 phase, accompanied by the accumulation of reactive oxygen species, decreased mitochondrial membrane potential, and increased expression levels of p53 and p21 proteins, caspase-3 activation, and apoptosis. Therefore, RJ, RJEE, and RJPH have potential inhibitory effects on the proliferation of gastric cancer cells.

The study on 4D culture system of squamous cell carcinoma of tongue

Traditional cell culture methods often fail to accurately replicate the intricate microenvironments crucial for studying specific cell growth patterns. In our study, we developed a 4D cell culture model-a precision instrument comprising an electromagnet, a force transducer, and a cantilever bracket. The experimental setup involves placing a Petri dish above the electromagnet, where gel beads encapsulating magnetic nanoparticles and tongue cancer cells are positioned. In this model, a magnetic force is generated on the magnetic nanoparticles in the culture medium to drive the gel to move and deform when the magnet is energized, thereby exerting an external force on the cells. This setup can mimic the microenvironment of tongue squamous cell carcinoma CAL-27 cells under mechanical stress induced by tongue movements. Electron microscopy and rheological analysis were performed on the hydrogels to confirm the porosity of alginate and its favorable viscoelastic properties. Additionally, Calcein-AM/PI staining was conducted to verify the biosafety of the hydrogel culture system. It mimics the microenvironment where tongue squamous cell carcinoma CAL-27 cells are stimulated by mechanical stress during tongue movement. Electron microscopy and rheological analysis experiments were conducted on hydrogels to assess the porosity of alginate and its viscoelastic properties. Calcein-AM/PI staining was performed to evaluate the biosafety of the hydrogel culture system. We confirmed that the proliferation of CAL-27 tongue squamous cells significantly increased with increased matrix stiffness after 5 d as assessed by MTT. After 15 d of incubation, the tumor spheroid diameter of the 1%-4D group was larger than that of the hydrogel-only culture. The Transwell assay demonstrated that mechanical stress stimulation and increased matrix stiffness could enhance cell aggressiveness. Flow cytometry experiments revealed a decrease in the number of cells in the resting or growth phase (G0/G1 phase), coupled with an increase in the proportion of cells in the preparation-for-division phase (G2/M phase). RT-PCR confirmed decreased expression levels of P53 and integrinβ3 RNA in the 1%-4D group after 21 d of 4D culture, alongside significant increases in the expression levels of Kindlin-2 and integrinαv. Immunofluorescence assays confirmed that 4D culture enhances tissue oxygenation and diminishes nuclear aggregation of HIF-1α. This device mimics the microenvironment of tongue cancer cells under mechanical force and increased matrix hardness during tongue movement, faithfully reproducing cell growthin vivo, and offering a solid foundation for further research on the pathogenic matrix of tongue cancer and drug treatments.

Green synthesis of copper oxide nanoparticles using walnut shell and their size dependent anticancer effects on breast and colorectal cancer cell lines

Metal oxide nanoparticles(NPs) contain unique properties which have made them attractive agents in cancer treatment. The CuO nanoparticles were green synthesized using walnut shell powder in different calcination temperatures (400°, 500°, 700°, and 900 °C). The CuO nanoparticles are characterized by FTIR, XRD, BET, SEM and DLS analyses. SEM and DLS analyses showed that by increasing the required calcination temperature for synthesizing the NPs, their size was increased. DPPH analysis displayed no significant anti-oxidative properties of the CuO NPs. The MTT analysis showed that all synthesized CuO NPs exhibited cytotoxic effects on MCF-7, HCT-116, and HEK-293 cell lines. Among the CuO NPs, the CuO-900 NPs showed the least cytotoxic effect on the HEK-293 cell line (IC50 = 330.8 µg/ml). Hoechst staining and real-time analysis suggested that the CuO-900 NPs induced apoptosis by elevation of p53 and Bax genes expression levels. Also, the CuO-900 NPs increased the Nrf-2 gene expression level in MCF-7 cells, despite the HCT-116 cells. As can be concluded from the results, the CuO-900 NPs exerted promising cytotoxic effects on breast and colon cancer cells.

Dietary Zn proteinate with moderate chelation strength alleviates heat stress-induced intestinal barrier function damage by promoting expression of tight junction proteins via the A20/NF-κB p65/MMP-2 pathway in the jejunum of broilers

BackgroundThe aim of this study was to determine whether and how Zn proteinate with moderate chelation strength (Zn-Prot M) can alleviate heat stress (HS)-induced intestinal barrier function damage of broilers. A completely randomized design was used for comparatively testing the effects of Zn proteinate on HS and non-HS broilers. Under high temperature (HT), a 1 (Control, HT-CON) + 2 (Zn source) × 2 (added Zn level) factorial arrangement of treatments was used. The 2 added Zn sources were Zn-Prot M and Zn sulfate (ZnS), and the 2 added Zn levels were 30 and 60 mg/kg. Under normal temperature (NT), a CON group (NT-CON) and pair-fed group (NT-PF) were included.ResultsThe results showed that HS significantly reduced mRNA and protein expression levels of claudin-1, occludin, junctional adhesion molecule-A (JAMA), zonula occludens-1 (ZO-1) and zinc finger protein A20 (A20) in the jejunum, and HS also remarkably increased serum fluorescein isothiocyanate dextran (FITC-D), endotoxin and interleukin (IL)-1β contents, serum diamine oxidase (DAO) and matrix metalloproteinase (MMP)-2 activities, nuclear factor kappa-B (NF-κB) p65 mRNA expression level, and protein expression levels of NF-κB p65 and MMP-2 in the jejunum. However, dietary supplementation with Zn, especially organic Zn as Zn-Prot M at 60 mg/kg, significantly decreased serum FITC-D, endotoxin and IL-1β contents, serum DAO and MMP-2 activities, NF-κB p65 mRNA expression level, and protein expression levels of NF-κB p65 and MMP-2 in the jejunum of HS broilers, and notably promoted mRNA and protein expression levels of claudin-1, ZO-1 and A20.ConclusionsOur results suggest that dietary Zn, especially 60 mg Zn/kg as Zn-Prot M, can alleviate HS-induced intestinal barrier function damage by promoting the expression of TJ proteins possibly via induction of A20-mediated suppression of the NF-κB p65/MMP-2 pathway in the jejunum of HS broilers.

Lovastatin Combination Therapy Increases the Survival and Proliferation of Rat Bone Marrow-Derived Mesenchymal Stem Cells Against the Inflammatory Activity of Lipopolysaccharide.

Oxidative stress hurts the survival of transplanted mesenchymal stem cells (MSCs). Lipopolysaccharide (LPS) preconditioning inhibits apoptotic death in MSCs. Also, Lovastatin's protective effect was reported on MSCs. Here, we investigated the potential of LPS and Lovastatin combination therapy on the survival and proliferation of MSCs. MSCs harvested from adult rats (240-260 g) femur and tibia bone marrow. Third passage MSCs were divided into 6 groups control group, LPS, LPS + Lovastatin (10 and 15 µM), and Lovastatin (10 and 15 µM). Cell survival and proliferation were assessed using an MTT assay 24 h after LPS, Lovastatin, or LPS + Lovastatin treatment. Also, Malondialdehyde (MDA) as a lipid peroxidation marker and antioxidant enzymes such as Glutathione peroxidase (GPX) and Superoxide dismutase (SOD) activity levels evaluated. Finally, the expression level of tumor protein P53 (P53) and octamer-binding transcription factor 4 (OCT4) genes were measured by qRT-PCR test. Lovastatin 10 μM potentiated proliferation and survival of MSCs. It can increase the activity of GPX and SOD. 10 µM Lovastatin could not affect MDA amounts but decreased the expression levels of P53 and Oct4 significantly. Nevertheless, treatment with LPS reduced the survival and proliferation of MSCs, along with a significant reduction in GPX activity. LPS + Lovastatin could increase SOD activity, however, GPX enzyme activity and MSCs proliferation did not change so, and it was not effective. We propose Lovastatin at the dose of 10 µM as a suitable combination agent to increase the survival and proliferation of MSCs in oxidative stress conditions.

Hypoxia-induced autophagy inhibits myometrial proliferation in pregnant mice via the mTOR pathway in vitro.

Hypoxia is closely associated with physiological and pathological conditions in the human body, and the myometrium is affected by hypoxic stress during pregnancy and delivery. Autophagy is a catabolic pathway involved in the regulation of apoptosis, proliferation and migration of a variety of cells, which can be activated under hypoxia. However, the mechanism and function of autophagy in uterine smooth muscle cells remained unclear. The aim of this study was to investigate the changes of autophagy in pregnant uterine smooth muscle cells (pUSMCs) under hypoxia and the effect of autophagy on myometrial cells proliferation during pregnancy. In this study, primary uterine smooth muscle cells were isolated from mice in late pregnancy and cultured under normoxic and hypoxic conditions respectively. Western blotting and immunofluorescence were used to detect the expression levels of autophagy-related proteins LC3B, P62, mTOR and p-mTOR under different culture conditions. Cell proliferation was assessed by CCK-8 assay. In addition, 3-Methyladenine (3-MA) was used to inhibit autophagy in hypoxia-treated pUSMCs and MHY1485 was used to activate mTOR. Studies have confirmed that under hypoxic conditions, autophagy is enhanced and cell proliferative viability is reduced in pUSMCs. Autophagy inhibitor 3-MA restored cell proliferation inhibited by hypoxia. Furthermore, hypoxia in pUSMCs led to a downregulation of p-mTOR/mTOR levels. The mTOR activator MHY1485 inhibited autophagy by preventing the binding of autophagosomes to lysosomes and reversed the hypoxia-induced inhibition of cell proliferation. Collectively, our results indicate that hypoxia upregulates autophagy through the mTOR pathway in pUSMCs, thereby inhibiting cell proliferation during pregnancy.

The anti-inflammatory effect of arsenic trioxide effectively mitigates the pathogenic process in local chickens with avian leukosis

Arsenic trioxide (ATO) is a classic first-line treatment for acute promyelocytic leukemia (APL). An increasing number of studies regarding the use of ATO in tumor treatment have shown consistently remarkable results. In this study, subgroup J avian leukosis virus (ALV-J) was used as a model virus, and different doses of ATO were used to treat ALV-J-positive chickens. Sexually mature green-shelled laying hens from the same ALV-J-positive offspring were grouped and treated with one of 3 different doses of ATO. The anti-inflammatory effects of different doses of ATO in ALV-J-positive chickens and their mechanisms were investigated by analyzing levels of inflammatory cytokines, antioxidant parameters and apoptosis-related genes. The results showed that ATO administration mitigated ALV-induced lymphoid leukosis in the liver. ATO inhibited the activation of the TLR4/MyD88/NF-κB signaling pathway and downregulated the expression levels of the inflammatory cytokines IL-1β, IL-6 and TNF-α. The SOD and GSH-Px activities were also increased, and the MDA content was decreased in the serum of ALV-J-positive chickens treated with different doses of ATO, so the antioxidant capacity of ALV-J-positive chickens was improved. The mRNA expression levels of p53, p21 and Bcl-2 in the livers of ALV-J-positive chickens treated with different doses of ATO were significantly downregulated, which induced the apoptosis of tumor cells and slowed the inflammatory response. The combined analysis revealed that the therapeutic effect of 2 mg/kg/dose ATO was superior to that of the other 2 treatments (0.5 and 1 mg/kg/dose ATO). In conclusion, the anti-inflammatory effect of ATO can effectively alleviate the ALV-J pathogenic process. ALV-J serves as a model virus for antiviral tumor research, while ATO provides references for the treatment of such tumors.

Oxaliplatin activates P53/miR-34a/survivin axis in inhibiting the progression of gastric cancer cells.

The purpose of this research was to determine how the P53/microRNA-34a (miR-34a)/survivin pathway contributes to oxaliplatin-induced (L-OHP) cell inhibition in gastric cancer. The BGC-823 gastric cancer cells were selected, and we examined their viability following treatment with L-OHP at different concentrations and time periods. The expression levels of miR-34a, P53, and survivin in the cells were determined. In the 12- and 24-h groups, drug concentration of 15 μg/cm² (p < .005 in both) significantly lowered cell viability. In comparison to the control group, miR-34a mRNA expression, P53 mRNA expression, and protein expression were all significantly greater in the 24-h group (p = .0324, p = .0069, p = .0260, respectively), but survivin mRNA and protein expressions were significantly lower than those in the control group (p = .0338, p = .0032, respectively). There was a significant decrease in gastric cancer cells in the miR-34a overexpression group (p = .0020), a significant increase in P53 mRNA and protein expression compared to the control group (p = .0080, p = .0121, respectively), and a significant decrease in survivin mRNA and protein expression compared to the control group. (p = .0213, p = .0069, respectively). Oxaliplatin inhibits tumor growth, invasion, and metastasis by upregulating miR-34a, activating the expression of the upstream P53 gene, and driving the downregulation of survivin (P53/miR-34a/survivin axis) in BGC-823 gastric cancer cells.

Inhibition of HDAC8 mitigates AKI by reducing DNA damage and promoting homologous recombination repair.

Nephrotoxicity is a major side effect of platinum-based antineoplastic drugs, and there is currently no available therapeutic intervention. Our study suggests that targeting histone deacetylase 8 could be a potential treatment for cisplatin-induced acute kidney injury (AKI). In a murine model of AKI induced by cisplatin, the administration of PCI-34051, a selective inhibitor of HDAC8, resulted in significant improvement in renal function and reduction in renal tubular damage and apoptosis. Pharmacological inhibition of HDAC8 also decreased caspase-3 and PARP1 cleavage, attenuated Bax expression and preserved Bcl-2 levels in the injured kidney. In cultured murine renal epithelial cells (mRTECs) exposed to cisplatin, treatment with PCI-34051 or transfection with HDAC8 siRNA reduced apoptotic cell numbers and diminished expression of cleaved caspase-3 and PARP1; conversely, overexpression of HDAC8 intensified these changes. Additionally, PCI-34051 reduced p53 expression levels along with those for p21, p-CDK2 and γ-H2AX while preserving MRE11 expression in the injured kidney. Similarly, pharmacological and genetic inhibition of HDAC8 reduced γ-H2AX and enhanced MRE11 expression; conversely, HDAC8 overexpression exacerbated these changes in mRTECs exposed to cisplatin. These results support that HDAC8 inhibition attenuates cisplatin-induced AKI through a mechanism associated with reducing DNA damage and promoting its repair.

Effects of Life-Long Exercise on Age-Related Inflammation, Apoptosis, Oxidative Stress, Ferroptosis Markers, and NRF2/KAEP 1/Klotho Pathway in Rat Kidneys

To investigate the effects of life-long exercise (LLE) on age-related inflammatory cytokines, apoptosis, oxidative stress, ferroptosis markers, and the NRF2/KAEP 1/Klotho pathway in rats. Eight-month-old female Sprague-Dawley rats were divided into four groups: 1) LLE: 18-month LLE training starting at 8 months of age, 2) Old moderate-intensity continuous training (OMICT): 8 months of moderate-intensity continuous training starting at 18 months of age, 3) Adult sedentary (ASED): 8 month-old adult sedentary control group, and 4) Old sedentary (OSED): a 26-month-old sedentary control group. Hematoxylin eosin staining was performed to observe the pathological changes of kidney tissue injury in rats; Masson’s staining to observe the deposition of collagen fibers in rat kidney tissues; and western blotting to detect the expression levels of IL-6, IL 1β, p53, p21, TNF-α, GPX4, KAEP 1, NRF2, SLC7A11, and other proteins in kidney tissues. Results: Compared with the ASED group, the OSED group showed significant morphological changes in renal tubules and glomeruli, which were swollen and deformed, with a small number of inflammatory cells infiltrated in the tubules. Compared with the OSED group, the expression levels of inflammation-related proteins such as IL-1β, IL-6, TNF α, and MMP3 were significantly lower in the LLE group. Quantitative immunofluorescence analysis and western blotting revealed that compared with the ASED group, KAEP 1 protein fluorescence intensity and protein expression levels were significantly enhanced, while Klotho and NRF2 protein fluorescence intensity and protein expression levels were reduced in the OSED group. Compared with the OSED group, KAEP 1 protein fluorescence intensity and protein expression levels were reduced in the LLE and OMICT groups. Klotho and KAEP 1 protein expression levels and immunofluorescence intensity were higher in the LLE group than in the OSED group. The expression levels of GPX4 and SLC7A11, two negative marker proteins associated with ferroptosis, were significantly higher in the LLE group than in the OSED group, while the expression of p53 a cellular senescence-associated protein that negatively regulates SLC7A11, and the downstream protein p21 were significantly decreased. LLE may ameliorated aging-induced oxidative stress, inflammatory response, apoptosis, and ferroptosis by regulating Klotho and synergistically activating the NRF2/KAEP 1 pathway.

Structural characterization and antioxidant activities of polysaccharides extracted from Polygonati rhizoma pomace

The present study aimed to prepare polysaccharides from the pomace of Polygonatum rhizome and characterize their structural features and biological activities. After hot water extraction and DEAE-52 cellulose fractionation, a neutral polysaccharide (PKP) was obtained with 91.85% sugars and 0.45% proteins. Structural characterization indicated that PKP contained a main fraction with a molecular weight of 4.634 × 103 Da and was composed of →1)-β-D-Fruf-(2→ and →6)-β-D-Fruf-(2→ residues. PKP was a semi-crystalline polymer, and the Congo red assay suggested the presence of triple-helix structure in PKP. PKP exhibited moderate radical scavenging activity (including 15.55% inhibition of DPPH, 21.48% inhibition of ABTS, and 22.52% inhibition of ·OH) and could effectively protect MRC-5 cells from H2O2-induced oxidative damage at 0.01 mg/mL through inhibiting apoptosis, decreasing SA-β-galactosidase activity, and downregulating the expression levels of p16 and p53. Therefore, PKP could be used in functional foods and pharmaceuticals as an antioxidant. This study provides an attractive method for utilizing polysaccharides from waste materials.

Assessment of the Circulating PD-1 and PD-L1 Levels and P53 Expression as a Predictor of Relapse in Pediatric Patients with Wilms Tumor and Hypernephroma.

Background/Objectives: Wilms tumor (WT) is the most common form of pediatric renal tumor, accounting for over 90% of cases followed by hypernephroma. Some pediatric patients with WT (10%) experience relapse or metastasis and have poor survival rates. PD-L1 assists cancer cells in escaping damage from the immune system. P53 mutations are found in relapsed WT tumor samples. We hypothesized that testing circulating PD-1 and PD-L1 and P53 expression levels could offer a simple method to predict patient relapse and explore novel treatments for pediatric WTs and hypernephroma. Methods: Flow cytometric detection of cPD-1, cPD-L1, and P53 expression in relapsed and in-remission WT and hypernephroma before and after one year of chemotherapy was performed. Results: Our data shows increased levels of cPD-L1 in relapsed pediatric patients with WT or hypernephroma before and after chemotherapy. There were also slight and significant increases in cPD-1 levels in relapsed groups before chemotherapy. Additionally, we observed significant decreases in P53 expression after one year of chemotherapy in relapsed pediatric patients. Conclusions: Our study found that circulating PD-L1 can be used as a predictor marker for WT and hypernephroma relapse. In conclusion, these circulating markers can assist in monitoring relapse in WT and hypernephroma patients without the need for several biopsies.

Bioinformatics analysis reveals that CBX2 promotes enzalutamide resistance in prostate cancer

Enzalutamide (Enz) is commonly utilized as the initial treatment strategy for advanced prostate cancer (PCa). However, a notable subset of patients may experience resistance to Enz, resulting in reduced effectiveness. Utilizing Gene Expression Omnibus (GEO) databases, we identified CBX2 as a crucial factor in mediating resistance to Enz, primarily due to its inhibitory effect on the P53 signaling pathway. Silencing of CBX2 using small interfering RNA (siRNA) led to elevated levels of P53 expression in LNCaP cells. This indicates that CBX2 may have a critical effect on PCa Enz resistance and could serve as a promising therapeutic target for individuals with Enz resistance.

Role of NF-κB signaling pathway in H2O2-induced oxidative stress of hiPSCs.

The balance between oxidation and antioxidation is crucial for the development of embryo. It is harmful to the early embryonic development if embryonic stem cells (ESCs) encounter the serious oxidative stress in vivo. Induced pluripotent stem cells (iPSCs) are very similar to ESCs and are the important cell source to replace ESCs for research and therapy. Studies show that iPSCs have better resistant ability to oxidative stress, but the involved mechanism remains unclear. In this study, we predicted that the NF-κB pathway might be involved in H2O2-induced developmental damage by network toxicology analysis. Then, the oxidative stress model was established with different concentrations of H2O2 to investigate the mechanism of NF-κB pathway in oxidative stress of human induced pluripotent stem cells (hiPSCs). The results showed as follows: With the increase of H2O2 concentration, the ROS level gradually went up leading to an increasing damage degree of hiPSCs; however, the MDA content was obviously high only in the 400μM H2O2 group; the activities of some antioxidant indexes such as SOD2 and T-AOC were significantly upregulated in the 100μM group, while most of antioxidant indexes showed downregulated tendency to different degrees with the increase of H2O2 concentration. The expression levels of P65, P50, IκB, SOD2, and FHC mRNA were upregulated in most H2O2-treated groups, showing a dose-dependent relationship. In subsequent experiments, the inhibitor of IκB-α phosphorylation, Bay11-7082, reversed the upregulation of P65, IκB, and FHC mRNA expression induced by 400μM H2O2. The protein levels of P65, p-P65, P50, p-P50, IκB, p-IκB, SOD2, and FHC were upregulated in most H2O2-treated groups. However, the upregulation induced by 400μM H2O2 could be reversed by BAY 11-7082, except for IκB and SOD2. In conclusion, H2O2 could promote the expressions and phosphorylations of NF-κB that could upregulate the expressions of its downstream antioxidant genes to minimize the damage of hiPSCs caused by oxidative stress. These results contribute to a fundamental understanding of the antioxidant mechanism of iPSCs and will further facilitate the application of iPSCs, as well as provide a reference for controlling the oxidative stress encountered in the early development stage of embryo.

Based on SIRT1/NF-κB/NLRP3 Signal Pathway to Explore the Effect of Yiqi Huatan Huoxue Recipe on Inflammatory Injury in AIT Mice by Pyroptosis.

Autoimmune thyroiditis (AIT) is the most common autoimmune thyroid disease. In recent decades, its incidence and prevalence have sharply increased. Yiqi Huatan Huoxue recipe is a traditional Chinese medicine formula we use to treat AIT. Its clinical efficacy is clear, but the specific mechanism remains unclear. This study aims to explore whether pyroptosis mediated by the SIRT1/NF-κB/NLRP3 signaling pathway is one of the therapeutic mechanisms of Yiqi Huatan Huoxue recipe. Forty 8-week-old female NOD.H-2h4 mice were randomly divided into four groups: the normal group (NG), model group (MG), Yiqi Huatan Huoxue recipe group (YG), and western medicine group (selenium yeast tablet, SeG). The normal group was gavaged with distilled water, while the remaining groups were gavaged with 0.05% sodium iodide (NaI) solution for 8 weeks. After the AIT animal model formed naturally, the mice were euthanized by gavage after 8 weeks. Hematoxylin-eosin staining was used to observe thyroid tissue changes, and enzymelinked immunosorbent assay (ELISA) was used to detect serum anti-thyroglobulin antibodies (TGAb) and mouse anti-thyroid peroxidase antibodies (TPOAb). Real-time quantitative PCR (qRT-PCR), Western blot, and immunohistochemistry were used to detect the expression of sirtuin 1 (SIRT1), nuclear factor κB p65 (NF-κB p65), nod-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC), caspase- 1, gasdermin D (GSDMD), and interleukin (IL)-1β in thyroid tissue. Compared with the NG group, the thyroid structure of rats in the MG group was severely damaged, with significant lymphocyte infiltration, significantly increased serum TGAb and TPOAb levels, and significantly increased expression levels of SRIT1, NF-κB p65, NLRP3, ASC, Caspase-1, GSDMD, IL-1β mRNA, and protein. Compared with the MG group, the thyroid structure damage and lymphocyte infiltration in rats of each treatment group were improved, and the serum TGAb, TPOAb, SRIT1, NF-κB p65, NLRP3, ASC, Caspase-1, GSDMD, IL-1β mRNA, and protein expression levels were significantly reduced. Yiqi Huatan Huoxue recipe can alleviate thyroid structural damage in AIT mice, and its mechanism may be related to the upregulation of SIRT1, NF-κB deacetylation, and inhibition of NLRP3-mediated pyroptosis.

Effect of impaired autophagic flux on breast carcinogenesis through the enhanced LC3-p62-NRF2 feedback loop.

6 Background: Breast cancer affects more women worldwide than any other type of cancer. Although some genes have been identified as potential causes, the exact molecular mechanism of the disease is still unknown. In our study, we aimed to investigate the role of NRF2 and the autophagy markers p62 and LC3 in breast cancer cell lines that have different Tp53 status using a new compound called 6,6'dihydroxythiobinupharidine (DTBN). In our previous research, we have shown that DTBN modulates key cellular signal transduction pathways that are relevant to disease biology, including cancer. Furthermore, we analyzed online databases to determine the correlation between NRF2, p62, LC3, and Tp53 in breast cancer patients. Methods: To evaluate the effect of DTBN on NRF2, p62 and LC3 in different types of breast cancer cell lines with varying TP53 status, we conducted molecular studies by inhibiting NRF2 activity. We synthesized the correlation between NRF2, p62, LC3 and TP53 expression and clinical parameters by analyzing TCGA and GEO datasets. Results: Our findings suggest that DTBN induces cell death and autophagy in all breast cancer cell lines, regardless of their Tp53 status. However, they also led us to hypothesize that NRF2 may hinder the sensitivity of cancer cells to DTBN-induced cell death and autophagy. To test this hypothesis, we inhibited NRF2 activity using an NRF2 inhibitor called brusatol. The results of the study indicate that the inhibition of NRF2 significantly increased cell death and autophagy in DTBN-treated cells. We observed a decrease in the expression of the autophagy marker p62, while an increase in the expression of another autophagy marker LC3. These findings suggest that the interaction between NRF2, LC3 and p62, which is activated in response to DTBN treatment, may serve as a potential anticancer drug target. We have validated the positive correlation between NRF2 and the autophagic gene p62 and LC3, in different Tp53 status breast cancer cell lines by analyzing TCGA and GEO data. Conclusions: Our study shows that high levels of Nrf2, LC3 and p62 expression are significantly associated with increased breast cancer cell proliferation and migration. This implies that patients with breast cancer have lower overall survival and a higher rate of recurrence. Our findings highlight the role of DTBN-induced NRF2 and LC3 expression and reduced p62 expression in breast cancer cells with varying Tp53 status. These results may represent a paradigm for better understanding the cancer cell response to therapies and designing more efficient combined anticancer therapies targeting NRF2, LC3 p62, and Tp53.