Gene P53 Research Articles

Biological and Molecular Characterization of Moroccan Isolates of Spiroplasma citri

Spiroplasma citri, a bacterium from the class Mollicutes, is the causative agent of citrus stubborn disease, a serious threat to Moroccan citrus crops, with yield losses reaching 45%. Despite its long-standing presence since 1949 and regulations mandating disease-free citrus plants, data on S. citri in Morocco remain scarce. This study investigates the pathogenicity and symptom variability of Moroccan S. citri isolates using biological indexing and genetic mapping based on the Spiralin and P58 genes. Biological indexing through reverse inoculation revealed that seven out of ten isolates caused moderate to intense symptoms within 8 to 10 weeks, with symptom severity varying across citrus cultivars and regions. These findings suggest variations in pathogen titer. Molecular analysis showed that Moroccan isolates (27GH, 3GH, 8GH, 56MK, 16MK, 60MK, 2GLK, 13SS, and 30S1) exhibited complete (100%) sequence similarity with each other and the reference strain R2-A8. Furthermore, these isolates displayed a high degree of similarity (99.75%) to a Corsican isolate (U13995) and a 94% similarity to an Iranian isolate (KP666137). Analysis of the P58 gene confirmed a high level of homogeneity with Moroccan reference strain R8-A2, closely aligning (99.75%) with the American BR3-3X strain, and 98% similarity to isolates from Syria and Iran. This study lays a foundational insight into the molecular characterization of S. citri in Morocco and provides a groundwork for future research into managing citrus stubborn disease.

Quantitative Proteomics Analysis Reveals XDH Related with Ovarian Oxidative Stress Involved in Broodiness of Geese

Studies have demonstrated significant alterations in ovarian oxidative stress levels, ovarian degeneration, and follicular atresia during the broody period in geese. The results of this study showed that during the broody period, geese exhibited degraded ovarian tissues, disrupted follicular development, a thinner granulosa cell layer, and lower levels of ovarian hormones E2, P4, and AMH. Antioxidant activity (GSH, CAT, SOD, T-AOC, and the content of H2O2) and the mRNA expression levels of antioxidant genes (GPX, SOD-1, SOD-2, CAT, COX-2, and Hsp70) were significantly higher in pre-broody geese compared to laying geese, while the expression of apoptosis-related genes (p53, Caspase-3, and Caspase-9) increased and the anti-apoptotic gene Bcl-2 decreased. Additionally, proteomic analysis identified 703 differentially expressed proteins (DEPs), primarily concentrated in the GO categories of the biological process (biological regulation, response to stimulus, etc.) and enriched in the KEGG pathways (PI3K-Akt signaling pathway, etc.). Among them, XDH was central to the regulatory network. Furthermore, Western blotting revealed higher expression of XDH in the ovaries of pre-broody geese than those of laying geese. Pearson correlation analysis indicated a significant correlation between XDH expression and oxidative stress markers in the ovaries of geese (r > 0.75). Overall, these results demonstrated that geese experience ovarian atrophy and remarkably increased oxidative stress during the broody period, suggesting that XDH may be a key driver of broodiness in geese.

Inhibition of ASIC1a reduces ferroptosis in rheumatoid arthritis articular chondrocytes via the p53/NRF2/SLC7A11 pathway.

The activation of acid-sensing ion channel 1a (ASIC1a) in response to extracellular acidification leads to an increase in extracellular calcium influx, thereby exacerbating the degeneration of articular chondrocytes in rheumatoid arthritis (RA). It has been suggested that the inhibition of extracellular calcium influx could potentially impede chondrocyte ferroptosis. The cystine transporter, solute carrier family 7 member 11 (SLC7A11), is recognized as a key regulator of ferroptosis. Recent studies suggest that the tumor suppressor gene p53 facilitates the induction of ferroptosis by suppressing the upregulation of SLC7A11. This process is mediated by the nuclear factor erythroid 2-related factor 2 (NRF2), a key transcription factor integral to the maintenance of cellular redox homeostasis and the regulation of inflammatory responses. This study aims to investigate the role of ASIC1a in the ferroptosis of RA chondrocytes and to determine the involvement of the p53/NRF2/SLC7A11 pathway in its underlying mechanism. Invitro experiments revealed that acidosis induces ferroptosis and reduces the expression of NRF2 and SLC7A11 in chondrocytes. Moreover, acidification significantly increased p53 protein levels in chondrocytes. Pifithrin-α (PFN-α), a p53 inhibitor, mitigated acidosis-induced ferroptosis and restored the diminished expression of NRF2 and SLC7A11. Furthermore, PcTx-1, an ASIC1a inhibitor, inhibited acidification-induced ferroptosis, enhanced the protein levels of SLC7A11 and NRF2, and reduced p53 expression. Invivo experiments demonstrated that the ASIC1a-specific inhibitor PcTx-1 ameliorated histopathological characteristics of ankle joints in collagen-induced arthritis (CIA) mice, decreased p53 expression, and enhanced NRF2 and SLC7A11 expression in chondrocytes. These findings suggest that ASIC1a inhibition may mitigate acidification-induced ferroptosis in articular chondrocytes in RA, potentially via the p53/NRF2/SLC7A11 pathway.

Unveiling the potential anticancer activity of Spirulina maxima extract-nanoemulsion through in vitro and in vivo studies.

Being the second leading cause of death globally, cancer has been a long-standing and rapidly evolving focus of biomedical research and practice in the world. Recently, there has been growing interest in cyanobacteria. This focus is particularly evident in developing innovative anticancer treatments to reduce reliance on traditional chemotherapy. This study investigates the anticancer potential of the Spirulina maxima extract nanoemulsion (SMNE) technique to improve the delivery, stability, and solubility of the S. maxima extract (SME). SMNE, prepared in three concentrations (SMNEC1, SMNEC2, SMNEC3), was characterized and confirmed to successfully load SME into silica-coated nanoparticles. Cytotoxicity tests on HepG2 and MCF-7 cell lines revealed a significant reduction in cell viability after 48-hour SMNE treatment, with IC50 values of 1488µg/mL and 1721.936µg/mL, respectively. SMNE also demonstrated efficacy in inhibiting tumor growth in mice with Ehrlich ascites carcinoma, normalizing alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, and reducing oxidative stress markers such as catalase (CAT) and malondialdehyde (MDA). Histopathological examination showed that SMNEC3-treated groups had almost normal liver architecture. Additionally, SMNE downregulated oncogenic miR-221-3p and miR-222-3p, activating cancer suppression genes p27 and PTEN. The study concludes that SMNE, with its anti-inflammatory and antioxidant properties and ability to modulate key miRNAs, enhances SME delivery and shows promise as an effective cancer treatment.

GH inhibits ALV-J replication and restricts cell cycle by activating PI3K/Akt signaling pathway

Growth hormone (GH) plays a crucial role in growth, sexual maturity, and immunity in chickens. Avian leukosis virus subgroup J (ALV-J) is an exogenous tumorigenic retrovirus that primarily induces immunosuppression, growth retardation, decreased egg production, tumors formation, and even death in chickens. Previous studies have suggested that GH is involved in the regulation of innate immunity and inflammation. However, the specific role of GH in response to ALV-J remains unclear. In this study, we observed a significant upregulation of GH protein expression in the plasma of ALV infected chickens, and a marked increase in GH mRNA in ALV-J infected cells. We found that lower gp85 expression correlated with higher GH expression in immune tissues, suggesting that GH may inhibit gp85 expression. Additionally, GH overexpression enhanced the expression of interferons (IFN-α, IFN-β), interferon-stimulating genes (Mx1, ASCL1, CH25H), and pro-inflammatory factors (Mx1, ASCL1, CH25H) in DF-1 cells infected with ALV-J. GH also affected the cell cycle by regulating the expression of cell proliferation-related genes (p21, PCNA, Cyclin B2, Cyclin D1, Cyclin D2) and cell apoptosis-related genes (p53, Fas, Cyct, Caspase-1, Caspase-3, Caspase-8). More importantly, we found that GH restricted cell proliferation and apoptosis, and inhibited the replication of ALV-J by activating the PI3K/Akt signaling pathway in DF-1 cells. In conclusion, these results indicate GH plays a role in the antiviral response against the replication of ALV-J, providing evidence of an interaction between GH and the innate immunity in chickens.

Effective Targeting of Glutamine Synthetase with Amino Acid Analogs as a Novel Therapeutic Approach in Breast Cancer

Cancer cells undergo metabolic rewiring to support rapid proliferation and survival in challenging environments. Glutamine is a preferred resource for cancer metabolism, as it provides both carbon and nitrogen for cellular biogenesis. Recent studies suggest the potential anticancer activity of amino acid analogs. Some of these analogs disrupt cellular nucleotide synthesis, thereby inhibiting the formation of DNA and RNA in cancer cells. In the present study, we investigated the anticancer properties of Acivicin and Azaserine in the breast cancer MCF-7 cell line, comparing their effects to those on the non-tumorigenic MCF-10 epithelial cell line in vitro. Interestingly, at lower concentrations, both Acivicin and Azaserine showed potent inhibition of MCF-7 cell proliferation, as assessed by the MTT assay, without detectable toxicity to normal cells. In contrast, Sorafenib (Nexavar), a commonly used drug for solid tumors, showed harmful effects on normal cells, as indicated by increased lactate dehydrogenase (LDH) production in treated cells. Furthermore, unlike Sorafenib, treatment with Acivicin and Azaserine significantly affected apoptotic signaling in treated cells, indicating the role of both amino acid analogs in activating programmed cell death (PCD), as assessed by the Annexin-V assay, DAPI staining, and the relative expression of tumor suppressor genes PTEN and P53. ELISA analysis of MCF-7 cells revealed that both Acivicin and Azaserine treatments promoted the production of anti-inflammatory cytokines, including IL-4 and IL-10, while significantly reducing the production of tumor necrosis factor alpha (TNF-α). Mechanistically, both Acivicin and Azaserine treatment led to a significant reduction in the expression of glutamine synthetase (GS) at both the RNA and protein levels, resulting in a decrease in intracellular glutamine concentrations over time. Additionally, both treatments showed comparable effects on Raf-1 gene expression and protein phosphorylation when compared with Sorafenib, a Raf-1 inhibitor. Moreover, docking studies confirmed the strong binding affinity between Acivicin, Azaserine, and glutamine synthetase, as evidenced by their docking scores and binding interactions with the enzyme crystal. Collectively, these findings provide evidence for the anticancer activity of the two amino acid analogs Acivicin and Azaserine as antagonists of glutamine synthetase, offering novel insights into potential therapeutic strategies for breast cancer.

Pleiotrophin Prevents H2O2-Induced Senescence of Dental Pulp Stem Cells.

Dental pulp stem cells (DPSCs) are widely used in research on dental tissue regeneration and systemic disease treatment. However, the oxidative microenvironment often causes cellular senescence, leading to decreased function. Our previous study demonstrated that pleiotrophin (PTN), a secreted extracellular matrix-associated protein, could rescue the proliferative capacity and osteogenic differentiation of replicative senescent DPSCs. This study aimed to explore the influence and mechanism of PTN on dental pulp stem cells under H2O2-induced oxidative microenvironment. DPSCs isolated from human third molars were treated with 100 μm H2O2 for 4 h, mimicking the oxidative microenvironment. To investigate the influence of PTN on DPSC under H2O2-induced oxidative microenvironment, 50 pg/mL PTN was added in the culture medium for 48 h. RT-qPCR, western blotting, SA-β-gal staining, intracellular ROS production and immunofluorescence staining assays were used to analyse the cellular senescence, osteogenic differentiation capacity, oxidative stress conditions and possible mechanism. H2O2 treatment increased the ratio of SA-β-gal-positive DPSCs and upregulated the senescence-related gene expression, including P53, P21 and P16. PTN pretreatment downregulated the ratio of SA-β-gal-positive DPSCs and the expression of these genes. Besides, PTN pretreatment partially reversed the H2O2-induced decreased osteogenic differentiation potential of DPSCs, total antioxidant capacity and Nrf2 and HO-1 mRNA expression in DPSCs. Western blotting and immunofluorescent staining results indicated that PTN pretreatment enhanced the Nrf2 nuclear translocation under oxidative stress conditions and observable higher fluorescence signals in the nucleus denoted PTN and Nrf2 colocalisation. Western blotting results showed that PTN reversed the decreased expression of p-AKT in the H2O2-induced oxidative environment. However, the PI3K inhibitor LY294002 blocked the upregulated levels of total Nrf2. Immunofluorescence staining displayed that LY294002 also inhibited the nuclear translocation of Nrf2 which was enhanced under PTN pretreatment. This study demonstrated that PTN could prevent senescent damage induced by H2O2 on DPSCs, mainly by combining with Nrf2 and enhancing its nuclear translocation.

Effect of WW-domain transcription regulator 1 on aging regulation of human dental pulp stem cells

Objective: Investigating the changes of phenotype and moleculars associated with aging with the increase of passage times of human dental pulp stem cells (hDPSC), to explore the role of WW-containing transcriptional regulator 1 (WWTR1) in the aging mechanism. Methods: hDPSCs were cultured by tissue block method, and were divided into 4 groups according to the age, algebra, cell knockdown and overexpression of WWTR1 in hDPSCs. Group Ⅰ: hDPSCs from human teeth were further divided into youth group (15-25 years old) and group middle-aged group (40-50 years old) according to different ages. Group Ⅱ: according to different passage, hDPSCs were divided into young cells group (hDPSCs were transmitted to P3 generation), and old cells group (hDPSCs were transmitted to P10 generation). Group Ⅲ: hDPSCs were knocked down of WWTR1, which were further divided into knockdown group and knockdown carrier group. Group Ⅳ: hDPSCs were overexpressed of WWTR1, which were further divided into overexpression group and overexpression carrier group. Real-time fluorescence quantitative PCR (RT-qPCR) was used to detect the changes of WWTR1 expression in groups Ⅰ and Ⅱ, and cell counting kit-8 (CCK-8) was used for groups Ⅱ, Ⅲ, and Ⅳ. Cell proliferation capacity was detected by CCK-8 assay. The ability of osteogenic differentiation was detected by alizarin red staining. Cell senescence positive rate was detected by age-related β-galactosidase staining. The expression levels of age-related genes p53 and p21 were detected by RT-qPCR. Results: The proportion of senescent cells increased gradually with continuous culture. The proliferation and osteogenic differentiation of hDPSCs in the old group were significantly lower than those in the young group (P<0.001). The expression levels of senescence related genes p53 (2.09±0.24) and p21 (4.91±0.54) in old cell group were higher than those in young cell group respectively [p53: (1.08±0.09) and p21: (1.09±0.08)] (P<0.01, P<0.001). The WWTR1 expression levels of hDPSCs in middle-aged group and old cells group were both decreased compared with those in young group and young cells group (P<0.01). The proportion of senescent cells in knockdown group (44.50±2.42) was higher than that in knockdown carrier group (22.27±0.56) (P<0.001). After knocking down WWTR1 in hDPSCs, the expression levels of age-related genes p53 and p21 were up-regulated (P<0.001), and the abilities of proliferation and osteogenic differentiation in the knockdown group were lower than those in the knockdown carrier group (P<0.001). The proportion of senescent cells in overexpression empty carrier group (20.40±0.79) was higher than that in overexpression group (10.07±0.61) (P<0.001). After WWTR1 overexpression ins hDPSCs, the expression levels of age-related genes p53 and p21 were down-regulated, and the proliferation and osteogenic differentiation ability in overexpression group were higher than those in overexpression carrier group (P<0.001). Conclusions: WWTR1 can inhibit the expression levels of age-related genes p53 and p21, thus delaying the aging process as well as promoting the proliferation and osteogenic differentiation of hDPSCs.

Atorvastatin calcium alleviates UVB-induced HaCat cell senescence and skin photoaging

Excessive exposure to ultraviolet radiation B (UVB) has been shown to contribute to the aging of human skin cells. Previous research has demonstrated that atorvastatin calcium (Ato) can mitigate the aging effects caused by chemotherapy drugs. However, it remains unclear whether Ato can alleviate skin aging induced by ultraviolet radiation. In this study, through in vitro experiments with Hacat cells, we found that Ato can significantly reduce the UVB-induced increased expression of age-related protein p16 and age-related gene p21, and also reduce the up-regulation of inflammatory factors such as IL-1 and IL-6. Besides, it can reduce the expression of metallomatrix protein (MMP1 and MMP9), and inhibit cell senescence and inflammatory damage. Similarly, we found that Ato can enhance skin collagen fiber reduction and collagen volume decrease, repair skin photoaging and damage induced by UVB rays, and speed up the rate at which the wounded location heals in vivo using Balb/c mice. In the mechanism, Ato markedly decreased the expression of p-p38, p-p65, p-mTOR in vivo and in vitro, suggesting that it may act on Mitogen-activated protein kinase (MAPK), Nuclear factor κB (NF- κB) and Mammalian target of rapamycin (mTOR) signaling pathways to produce above marked effects. In conclusion, Ato obviously relieved UVB-induced photoaging and damage, thus providing evidence for its potential in mitigating skin aging caused by ultraviolet radiation.

Exercise alleviates hematopoietic stem cell injury following radiation via the carnosine/Slc15a2-p53 axis

Ionizing radiation (IR) can cause severe dysfunction of hematopoietic stem cells (HSCs), leading to acute or prolonged myelosuppression. In recent years, physical exercise has been recognized as a healthy lifestyle as it can fight a variety of diseases. However, whether it provides protection against IR is not fully understood. In this study, we revealed that long-term moderate exercise mitigated IR-induced hematopoietic injury by generating carnosine from skeletal muscles. We found that exercised mice displayed reduced loss of HSC number and function after IR, accompanied by alleviated bone marrow damage. Interestingly, these effects were largely abrogated by specific deletion of carnosine synthase Carns1 in skeletal muscles. In contrast, carnosine treatment protected HSCs against IR-induced injury. Mechanistically, we demonstrated that exercise-generated carnosine was specifically transported to HSCs via Slc15a2 and then inhibited p53 transcriptional activity by directly interacting with its core DNA-binding domain, which led to downregulation of the p53 target genes p21 and Puma, thus promoting the proliferation and survival and inhibiting the senescence of irradiated HSCs. More importantly, a similar role of the carnosine/Slc15a2-p53 axis was observed in human cord blood-derived HSCs. Collectively, our data reveal that moderate exercise or carnosine supplementation may be potential antiradiation strategies.

Elucidating the potential of Annona muricata L. grown in Sri Lanka to be used in developing an anticancer drug against colorectal and breast cancers

BackgroundSince ancient times many traditional medicine systems around the world have been using different parts of Annona muricata L. (AM), to treat cancer. Indeed, numerous in vitro and in vivo studies also have shown anticancer properties of different solvent extracts of different parts of AM. Even the same part of the plant has shown different levels of anticancer properties based on geographical variations. Therefore, in the present study, the anticancer potential of the leaves, fruit pulp and the fruit peel of the AM that is grown in Sri Lanka was comparatively analyzed with the intention of identifying the most suitable part to be developed into a nutraceutical with anticancer effects.MethodsFreeze-dried aqueous extracts of immature leaves (ILAM), mature leaves (MLAM), pulp (PAM) and peel (PLAM) of AM were analyzed for their antioxidant activity using 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging and 2,2-azinobis-3-ethylbenzothiozoline-6-sulfonic acid (ABTS) cation decolorization assays. Their cytotoxicity on breast cancer (MCF-7) cells, colorectal adenocarcinoma (DLD-1) cells and normal human gingival fibroblasts (HGF-1) were determined by the 3-(4,5- dimethylthiazole-2-yl)-2,5-diphenyl-tetrazoliumbromide (MTT) assay. Their effect on mRNA expression of proapoptotic (Bax and caspase-7) and cell cycle arresting (p21) genes was analyzed by RT- qPCR in the same cell lines.ResultsILAM demonstrated the highest antioxidant activity in both DPPH and ABTS assays followed by MLAM, PLAM and PAM. In the MTT assay, both ILAM and MLAM demonstrated strong cytotoxic activity against MCF-7 and DLD-1 cell lines while there were no cytotoxic effects on the normal human cell line HGF-1. Both ILAM and MLAM demonstrated concentration-dependent upregulation of mRNA expression of cell cycle arresting gene p21 and apoptosis inducing genes Bax and caspase-7 in MCF-7 and DLD-1 cells.ConclusionThe AEAM leaves grown in Sri Lanka has significantly higher antioxidant activity as well as selective cytotoxic effects on MCF-7 and DLD-1 cancer cells compared to its PL and P counterparts. Further, the AEAM leaves induced mRNA expression of the anticancer genes p21, Bax and caspase-7, indicating its potential to be developed into an anticancer drug against breast and colorectal cancer.

Cytotoxic activity of silver nanoparticles prepared by eco-friendly synthesis using Lythrum salicaria extract on breast cancer cells.

Metal nanoparticles (NPs) have widely been investigated due to their several applications in therapeutic activities. The current investigation highlights the cytotoxic effects of the eco-friendly phytosynthesis route for silver nanoparticles using Lythrum salicaria (L. salicaria) extract (AgNPs-LS). The change in color from colorless to brown confirmed the reduction of silver ions to AgNPs. x-ray diffraction (XRD) analysis demonstrated high crystallinity. The surface morphology of AgNPs-LS was spherical, and their average sizes were 50nm. energy-dispersive x-ray analysis (EDAX) confirmed that silver was the predominant component, indicating the involvement of L. salicaria plant extract in the green synthesis process. In vitro dimethyl thiazolyl tetrazolium bromide (MTT) assay showed significant cytotoxicity of AgNPs-LS against MCF7 cells, with an IC50 of 113µg mL- 1. In contrast, AgNPs-LS showed minimal cytotoxicity to HEK293 cells (IC50: 254µg mL- 1), demonstrating a higher sensitivity of cancer cells to AgNPs-LS. Moreover, AgNPs-LS resulted in MCF7 cells producing reactive oxygen species (ROS) and undergoing cell cycle arrest at the G2/M phase, serving as barriers to the proliferation of cancer cells. Annexin V fluorescein isothiocyanate (FITC) assays and fluorescence microscopy confirmed the induction of apoptosis in MCF7 cells by AgNPs-LS. Gene expression analysis revealed upregulated pro-apoptotic genes (Bax, p53, caspase-3, and caspase-9) and downregulated an anti-apoptotic gene (Bcl2) in michigan cancer foundation7 (MCF7) cells treated with AgNPs-LS. These results indicate that AgNPs-LS induced apoptosis via the intrinsic pathway (mitochondrial-mediated mechanism) and involved p53-dependent regulation. The current study results implied that AgNPs-LS fabricated by a bio-green approach could be helpful to the future of nanomedicine.

Dissecting the Impact of Genetic Background on Oncogenic Response to Radiation Exposure in the Ptch1+/- Mouse Model.

Medulloblastoma (MB) is a common primary brain cancer in children. The sonic hedgehog (SHH) pathway is indispensable for the normal development of the cerebellum, and MB is often caused by persistent SHH activation owing to mutations in pathway components. Patched1 (PTCH1) is the primary receptor for the SHH ligand and a negative regulator of the SHH signal transduction pathway. Mice heterozygous for the Ptch1 gene (Ptch1+/-) are predisposed to MB development. Irradiation of newborn Ptch1+/- mice dramatically increases MB occurrence. A genetic background carrying the Ptch1 mutation significantly influences the risk of developing MB. This study aims to investigate the genetic background-related mechanisms that regulate radiation-induced cellular response and oncogenesis in the cerebellum. We employed multiple approaches, including: (a) analysis of cellular radiosensitivity in granule cell precursors (GCPs), the MB cells of origin, derived from Ptch1 mice with a genetic background that is sensitive (CD1) or resistant (C57Bl/6) to the induction of radiogenic MB; (b) identification of genes differentially expressed in spontaneous and radiation-induced MBs from these two mouse strains; (c) bioinformatic analysis to correlate the expression of radiation-induced genes with survival in MB patients; and (d) examining the expression of these genes in ex vivo MBs induced by single or repeated radiation doses. We have identified a potential gene expression signature-Trp53bp1, Bax, Cyclin D1, p21, and Nanog-that influences tumor response. In ex vivo cultured spontaneous MBs, the expression levels of these genes increase after irradiation in CD1 mice, but not in mice with a C57Bl/6 genetic background, suggesting that this signature could predict tumor response to radiation therapy and help develop strategies for targeting DNA damage repair in tumors. A detailed understanding of the mechanisms behind genetic background-related susceptibility to radiation-induced oncogenic responses is crucial for translational research.

Regulation mechanism of oxidative status, immunity and apoptosis induced by hypoxia and heat exposure via PI3K/Akt signaling pathway in Megalobrama amblycephala

Megalobrama amblycephala, a main herbivorous fish with notable economic benefits in China, often faces serious challenges to its survival and growth due to hypoxia and heat caused by factors such as global warming and intensive aquaculture. To evaluate the combined effects of these stressors, we performed a two-factor crossover test to assess the impacts of simultaneous exposure to hypoxia (2 mg/L) and heat (35 °C) on oxidative stress, immunity and apoptosis in M. amblycephala. These results showed that hypoxia and heat exposure significantly enhanced the expression of oxygen-sensing and heat shock protein (HSP) genes, hypoxia inducible factor 1α (Hif-1α), HIF-prolyl hydroxylase-2 (phd2) and factor inhibiting Hif-1 (fih-1), as well as hsp70 and hsp90α. Furthermore, M. amblycephala suffering from hypoxia and heat exposure exhibited several changes in liver tissues, with the most severe lesions and up-regulation of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) observed in those subjected to simultaneous exposure. Moreover, the combined hypoxia and heat exposure initially triggered an increase in the activities of total antioxidant capacity (T-AOC), superoxide dismutase (SOD) and catalase (CAT), and glutathione (GSH) contents, followed by a reduction, and the accumulation of malondialdehyde (MDA), which induced oxidative stress. This was accompanied by an increase and subsequent reduction in the contents of alkaline phosphatase (AKP), acid phosphatase (ACP), complement component 3 (C3) and C4, immunoglobulin M (IgM), and interferon-γ (IFN-γ) leading to immunosuppression. Additionally, hypoxia and heat exposure up-regulated the expression of antioxidant enzyme genes (nrf2, cu/zn-sod, mn-sod cat, ho-1, pi3k and gpx-1a), inflammatory genes (interleukin il-1β, il-8 and tnf-α), immunity effectors (igm and lyz), as well as apoptosis genes (casp3, casp8, casp9 and p53) and activated p-Akt/Akt, suggesting apoptosis may be linked with oxidative stress and inflammation and mediated through the PI3K/Akt signaling pathway. In short, the combined hypoxia and heat exposure disrupted homoeostasis in M. amblycephala, with a more pronounced detrimental effect than exposure to either stressor alone These results will contribute to understanding the mechanism of combined exposure to hypoxia and heat in fish and provide a fundamental base for fisheries management.

In vitro anti-prostate cancer efficacy and phytochemical composition of the dichloromethane and ethyl acetate leaf extracts of Vitex doniana (sweet).

Prostate cancer is a significant global health concern, particularly among ageing male populations, with a disproportionately higher burden in sub-Saharan Africa. Conventional treatments, though effective, are costly and cause devastating side effects which limit their clinical benefits. Hence, this study evaluated the in vitro antiprostate cancer properties and secondary metabolites of dichloromethane and ethyl acetate lead extracts of Vitex doniana to explore safer and efficacious natural alternatives based on ethnomedicinal claims. Phytochemical profiling was conducted using gas chromatography-mass spectrometry (GC-MS) analysis to identify secondary metabolites in the extracts. The cytotoxic effects of the extracts were determined through the MTT assay using Vero CCL-81 cells and DU-145 cells. The expression profile of the selected genes (ar, bcl2, caspase-3, cdk1, and p53) in DU-145 cells treated with the study extracts was investigated using RT-qPCR. GC-MS analysis revealed 10 secondary metabolites in the dichloromethane extract and 27 secondary metabolites in the ethyl acetate extract of V. doniana leaves, with the majority being sesquiterpenes, diterpenoids, and phytosterols. The dichloromethane and ethyl acetate leaf extracts of V. doniana exhibited low cytotoxicity against normal mammalian epithelial cells (Vero CCL-81), with CC50 values of 1,238.85μg/mL and 964.81μg/mL, respectively. Besides, the ethyl acetate leaf extract of the studied plant demonstrated potent anti-prostate cancer activity against DU-145 cells, with an IC50 of 35.68μg/mL and a high selectivity index (SI) of 27.04. Likewise, the dichloromethane leaf extract of this plant displayed cytotoxic effects (IC50: 287.01μg/mL) and a selectivity index of 4.32. The reference drug (Doxorubicin) showed a higher toxicity against Vero CCL-81(IC50: 0.41μg/mL) and DU-145 (IC50: 0.28μg/mL) cells and a lower selectivity index of 1.46. The DU-145 cells treated with the studied plant extracts exhibited notable upregulation of ar and bcl2, and normalization of caspase 3, cdk1 and p53 expression. The studied plant extracts possess in vitro anti-prostate cancer properties and could be promising candidates for further preclinical studies aimed at developing novel botanical-based therapies for the management of prostate cancer.

Design, synthesis, and cytotoxicity screening of novel pyrazolopyrimidines over renal cell carcinoma (UO-31 cells) as p38α inhibitors, and apoptotic cells inducing activities

A series of novel molecules with pyrazolopyrimidine-4-amine core were designed and synthesized as potential cytotoxic agents over Renal Cell Carcinoma cells (UO-31). Results of cytotoxic activity against UO-31 cells showed that pyrazolopyrimidines 19 and 31 were found to be more cytotoxic than sorafenib (SOR). The cytotoxic activity of these compounds appeared to correlate with their ability to inhibit p38α MAPK which are 2.53- and 2.27- folds more potent than SOR. Moreover, results of the cell cycle analysis as well as the results of annexin-V on the (UO-31) cells showed that pyrazolopyrimidines 19 and 31 had a pro-apoptotic activity higher than SOR by 1.42- and 1.20- folds, respectively. Furthermore, compounds 19 and 31 were found to be effective in arresting the cell cycle throughout the accumulation of the cells at G2/M phase. Finally, the tested compounds decreased the TNF concentration as well as increased the expression of tumor suppressor gene p53, Bax/BCL-2 ratio and caspase 3/7.

Autologous transplantation of P63+ lung progenitor cells in patients with bronchiectasis: A randomized, single-blind, controlled trial

Non-cystic fibrosis bronchiectasis is a progressive respiratory disease with limited treatment options, prompting the exploration of regenerative therapies. This study investigates the safety and efficacy of autologous P63+ progenitor cell transplantation in a randomized, single-blind, controlled, phase 1/2 trial. Thirty-seven patients receive bronchoscopic airway clearance (B-ACT) (n= 19) or B-ACT plus P63+ progenitor cells (n= 18). Results show that compared to the control group, the change in DLCO levels from baseline to 24weeks post therapy is significantly higher in the cell treatment group (p value= 0.039). Furthermore, the patients in the cell treatment group demonstrate significantly reduced lung damaged area, improved SGRQ score, and ameliorated BSI and FACED scores within 4-12weeks post therapy. Transcriptomic analysis reveals that progenitor cells with higher expression of P63 gene have better therapeutic efficacy. These findings suggest that P63+ progenitor cells may offer a promising therapeutic approach for bronchiectasis. This study was registered at ClinicalTrials.gov(NCT03655808).

Biological effects of radiation exposure in patients treated with X-ray guided endovascular aortic repair

Abstract Background X-ray guided cardiovascular interventions such as endovascular aortic repair (EVAR) expose patients and operators to significant amounts of repeated ionising radiation over a long period of time. The biological effect of such protracted radiation exposure is poorly understood. We have previously demonstrated a higher incidence of dicentric chromosome (DC) aberrations in high-volume endovascular interventionalists and differences in acute DNA damage in lymphocytes between individuals following in vitro irradiation. Studies suggested a higher incidence of cancer in patients who underwent EVAR compared with those who have had open aneurysm repair, but the evidence is inconclusive. Further work is required to evaluate the biological radiation risk of X-ray guided cardiovascular interventions. Purposes We aimed to examine radiation-induced genome instability from patients after complex EVAR. We also aimed to investigate the intrinsic response of each patient to radiation using biomarkers of acute DNA damage following in vitro irradiation of their blood and to correlate this response to the expression of radiation-responsive genes. Methods Lymphocytes were isolated from patients after complex (branched/fenestrated) EVAR and non-irradiated controls. DC, a type of chromosomal aberration caused by radiation exposure were enumerated. γ-H2AX, a marker of acute DNA damage/repair, was measured by immunofluorescence after in vitro irradiation at 0.2 Gy and 1 Gy, along with the expression of the radiation-responsive genes FDXR, CCNG1, P21 and PHPT1 using qPCR. Results 17 patients (82% male, median age 73[59 – 85years]) and 16 controls (56% male, median age 68[53 – 83years]) were recruited. The mean incidence of DC was 3.782 (95% CI 3.13 - 4.43) and 0.898 (95% CI 0.48 -1.32) per 1000 cells for patients and controls, respectively (P&amp;lt;0.0001). Patients had higher background of γ-H2AX foci than unexposed controls, (0.7056, 95% CI 0.316 - 1.10) vs (0.241 95% CI 0.077 - 0.405) per cell, P&amp;lt;0.05. FDXR expression was most increased following irradiation (P&amp;lt;0.0001), reaching a mean 13.1-fold and 11.4-fold increase at 4-hour and 24-hour time-point, respectively, but this upregulation was not significantly different between the two groups. Conclusion We have shown an increased frequency of chromosomal aberrations in patients after complex EVAR, a biological finding that may support a propensity to developing malignancies. Patients also have higher basal DNA damage/repair (γ-H2AX) activity. Expression of ferredoxin reductase, FDXR gene could be a potential radio-responsive biomarker to provide personalised biological dosimetry information for clinical monitoring. Our study further underlines the long-term radiation risk of X-ray guided cardiovascular interventions and the potential of biological assays that may guide personalised care.

Neurotoxic and developmental effects of scented incense stick smoke: Network toxicology and zebrafish model study

Burning incense sticks is a traditional practice in many cultures, especially in Southeast Asia. While it is often regarded as sacred and beneficial, modern incense sticks contain various chemicals that can pose health risks. A GCMS analysis of the ICS revealed potential compounds. Network toxicology revealed that ICS contains compounds violating Lipinski's rule of five, leading to potential neurotoxic effects. Key pathways affected include neuroactive ligand-receptor interaction and calcium signaling, associated with neurodegenerative diseases like Parkinson's and Alzheimer's. Significant genes involved are STAT3, BCL2, and MTOR, emphasizing the chemical hazards of ICS exposure. We investigated the toxicity of ICS using zebrafish (Danio rerio) embryos as a mode. ICS exposure resulted in a dose-dependent increase in toxicity. High concentrations (7 and 14 µg/ml) led to immediate mortality, while lower concentrations (0.1, 0.3, 0.5, and 1 µg/ml) caused developmental defects such as yolk sac edema, skeletal malformations, and pericardial edema. Mortality rates increased with higher concentrations, confirming dose-dependent ICS exposure caused hypoactive locomotion, with reduced distance traveled and velocity toxicity. Higher concentrations of ICS led to increased ROS levels and cellular damage, as evidenced by enhanced staining levels. A dose-dependent increase in lipid peroxidation (DPPP assay) and lipid accumulation (Nile red assay) was observed. Higher ICS concentrations led to significant oxidative damage to lipids and increased lipid deposition. Enzymatic assays showed that ICS exposure significantly decreased the activities of antioxidant enzymes SOD and CAT, indicating impaired antioxidant defense, while increasing LDH activity, signaling tissue damage and cytotoxicity. Gene expression analysis revealed downregulation of SOD1 and CAT genes, upregulation of inflammatory genes TNF-α and IL-1β, and increased expression of the apoptotic gene p53 with decreased expression of Bcl-2 and BDNF. These findings highlight ICS's potential to cause oxidative stress, inflammation, apoptosis, and neurodevelopmental impairments.

Investigating the Mechanisms of Anti-tumoral Effect of Combination Therapy of Calcitriol and Sodium Pentaborate Pentahydrate on HepG2 Hepatocellular Carcinoma Cells.

Hepatocellular carcinoma (HCC) is one of the most common primary liver cancers worldwide and is often associated with poor prognosis due to drug resistance. Combination therapies demonstrate superior efficacy at lower drug dosages on cancer cells compared to single treatments, resulting in less drug resistance in the cells. This study investigates the synergistic anti-tumoral effects of calcitriol, the biologically active form of vitamin D, and sodium pentaborate pentahydrate (NaB) on HepG2 cells. We examined the cell viability of NaB, calcitriol, or the combination of NaB and calcitriol on HepG2 cells and healthy human hepatic stellate cells (HHSC) using MTS. Our findings showed that combination therapy with 3.3mM NaB and 1µM calcitriol has a synergistic effect and a more cytotoxic effect on HepG2 cells. This combination significantly increased apoptosis and ROS levels compared to treatment alone with NaB or calcitriol. Gene expression and proteomics analysis revealed inhibition of DNA replication and the cell cycle process, further confirming the potent anti-proliferative effects of the combination therapy. When HepG2 cells were treated with a combination of 3.3mM NaB and 1µM calcitriol, mRNA levels of apoptosis-related genes AKT1 and MDM2 were downregulated, while p53 was upregulated. Additionally, cell cycle-related genes CDKN1A, GADD45A, and p27 were upregulated, whereas MCM2, MCM5, and MCM7 were downregulated. Furthermore, genes associated with the vitamin D receptor (VDR), including VDR and CYP24A1, were upregulated, while CYP27B1 was downregulated. Our proteomic analysis revealed decreased MCM2 and MCM5 protein expressions which was confirmed by western blotting. In conclusion, this study highlights the potential of NaB and calcitriol as a promising therapeutic strategy for HCC.