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  • Article
    Jiajing Zhou, Lanyue Zhang, Jifeng Yan, Aihua Hou, Wenchao Sui, Meiling Sun
    Discovery Medicine. 2023, 35(176): 251-263. https://doi.org/10.24976/Discov.Med.202335176.26

    Background: Cancer stem cells (CSCs) are characterized by an ability for unlimited proliferation and efficiency of self-renewal. The targeting of lung CSCs (LCSCs)-related signaling pathways represent a promising therapeutic strategy for treatment of lung cancer. Ferroptosis a potential strategy for LCSCs treatment, and curcumin cloud induce ferroptosis. In this study, we aimed to observe the effects of curcumin on LCSCs via ferroptosis-related pathways.

    Methods: In this study, A549 cluster of differentiation (CD)133+ and A549 CD133 cells were isolated using magnetic bead-based separation. Colony formation and sphere formation assays, as well as cells injection in non-obese diabetes/severe combined immune deficiency (NOD/SCID) mice, were used to analyze the tumorigenic ability of cells differentially expressing CD133. A549 CD133+ cells were treated with different doses of curcumin (0, 10, 20, 40, 80 μM). Cell viability, glutathione peroxidase 4 (GPX4) and ferroptosis suppressor protein 1 (FSP1) expressions were measured. The 50% inhibitory concentration (IC50) of curcumin, two ferroptosis inducers, inhibitor of GPX4 (RSL3) and inhibitor of FSP1 (iFSP1), and a ferroptosis inhibitor, ferrostatin-1 (Fer-1), were used to investigate the mechanism underlying the effect of curcumin on ferroptosis in A549 CD133+ cells.

    Results: A549 CD133+ cells had greater tumorigenic ability than A549 cells. Curcumin treatment suppressed the expressions of GPX4 (glutathione peroxidase 4) and FSP1 in A549 CD133+ cells, thereby inducing ferroptosis. RSL3 and iFSP1 respectively suppressed the GSH (glutathione)-GPX4 and FSP1 (ferroptosis suppressor protein 1)-CoQ10 (coenzyme Q10)-nicotinamide adenine dinucleotide (NADH) pathways in A549 CD133+ cells. However, the roles of curcumin were blocked by Fer-1 treatment.

    Conclusions: In this study, curcumin induced ferroptosis through inhibiting the GSH-GPX4 and FSP1-CoQ10-NADH pathways in A549 CD133+ cells, resulting in the inhibition of their self-renewal potential.

  • Article
    Yang Fang, Yan-jing Wang, Hong-li Zhao, Xin Huang, Yi-nan Fang, Wen-yi Chen, Ruo-zhen Han, Ai Zhao, Ji-min Gao
    Discovery Medicine. 2023, 35(176): 405-417. https://doi.org/10.24976/Discov.Med.202335176.41

    Objectives: Over the past two decades, great progress has been made in advancing the early detection and multimodal treatment of non-small cell lung cancer (NSCLC). However, overall cure rates and survival rates of NSCLC are still not satisfactory, and research into new therapies is needed. This study attempted to construct human Fibroblast Activation Protein-Chimeric Antigen Receptor Natural killer (NK)-92 cells (hFAP-CAR-NK-92 cells) and explore their potential therapeutic effects in NSCLC.

    Methods: Immunohistochemistry analysis was carried out to examine fibroblast activation protein (FAP) and Gasdermin E (GSDME) expression in clinical specimens of lung adenocarcinoma and squamous cell carcinoma tissue. Then the engineered hFAP-CAR-NK-92 cells efficiency was determined in vitro with lactate dehydrogenase (LDH) cytotoxicity assay and the cell morphology of A549, H226, and cancer-related fibroblast (CAF) was observed by electron microscopy. After the co-culture of target cells and effect cells, flow cytometry was employed for examining the CD107a expression in the effect cells, and western blotting was conducted for the cleavage levels of Caspase 3 and GSDME proteins in the target cells. The safety and efficacy of hFAP-CAR-NK-92 cells adoptive transfer immunotherapy in a tumor-bearing mouse were evaluated.

    Results: Clinical studies have shown FAP positivity in patients with NSCLC. Compared with A549 or H226 cells alone, FAP expression was notably raised in A549+CAF cells or H226+CAF cells in nude mice, respectively (p < 0.05). The killing efficiency of K562 cells was not significantly different between hFAP-CAR-NK-92 and NK-92 cells (p > 0.05). The hFAP-CAR-NK-92 cells presented a higher killing efficiency against the hFAP-target (A549-hFAP, H226-hFAP and CAF-hFAP) cells than the NK-92 cells (p < 0.05). The degranulation of CD107a and cleavage levels of GSDME and Caspase 3 protein in the hFAP-CAR-NK-92 group were higher than those in the NK-92 group (p < 0.05). The 300 nM Granzyme B also induced pyroptosis in hFAP- or GSDME-positive cells (p < 0.05). In vivo experiments revealed that hFAP-CAR-NK-92 cells inhibited tumor progression of hFAP-positive NSCLC (p < 0.05).

    Conclusions: In this study, we successfully constructed hFAP-CAR-NK-92 cells and confirmed that hFAP-CAR-NK-92 cells could target hFAP-positive NSCLC to inhibit the progression of NSCLC by activating the Caspase-3/GSDME pyroptosis pathway.

  • Article
    Yingchao Gao, Yuanyuan Wang, Xin Wang, Jianwei Ma, Ming Wei, Na Li, Zengren Zhao
    Discovery Medicine. 2023, 35(176): 361-371. https://doi.org/10.24976/Discov.Med.202335176.37

    Background: Colorectal cancer is a common digestive tract malignancy. This study aimed to expound the functional role of fatty-acid-binding protein 4 (FABP4) and the potential underlying mechanisms in the development of colorectal cancer.

    Methods: Several techniques were utilized to investigate the role of FABP4 in colorectal cancer. FABP4 mRNA expression was quantified using Real time-quantitative PCR (RT-qPCR). Cell counting kit-8 (CCK-8), 5-ethynyl-2'-deoxyuridine (EdU), sphere formation assays and flow cytometry evaluated cell growth, stemness, and apoptosis in SW480 and HT29 cells. Glycolysis was assessed via extracellular acidification rate (ECAR) , lactate production, glucose uptake, adenosine triphosphate (ATP)/adenosine 5'-diphosphate (ADP) ratio, and Glut1 and Elevated lactate dehydrogenase A (LDHA) protein expression. Reactive oxygen species (ROS) levels were analyzed by flow cytometry. Western blot measured the protein expression of FABP4, Proliferating cell nuclear antigen (PCNA), Bax, Bcl-2, Glut1, LDHA, stemness makers (Sox2, Oct4, and ALDHA1), and extracellular regulated protein kinase (ERK)/mammalian target of rapamycin (mTOR) pathway proteins. In vivo experiments, BALB/c nude mice (n = 12) were inoculated with 200 μL HT29 cells (5 × 106 cells) transfected with sh-FABP4 or short hairpin (sh)-negative control (NC), forming two groups with 6 mice each. The in vivo mice tumor model allowed for evaluating FABP4's impact on tumor growth.

    Results: FABP4 was significantly upregulated in colorectal cancer tissues and cells (p < 0.05). FABP4 knockdown markedly inhibited cell proliferation, stemness, and glycolysis, while promoting apoptosis in these cells (p < 0.05). Additionally, FABP4 depletion led to a significant increase in ROS level (p < 0.05). However, N-acetyl-L-cysteine (NAC) (p < 0.05), a ROS scavenger, mitigates these effects. Furthermore, the effects of FABP4 depletion on cell growth, stemness, glycolysis, and apoptosis in colorectal cancer cells were also retarded by NAC (p < 0.05). Notably, FABP4 knockdown also suppressed the ERK/mTOR pathway, suggesting its regulation via ROS (p < 0.05). In vivo study results showed, FABP4 depletion significantly curbed tumor growth in colorectal cancer (p < 0.05).

    Conclusions: These results suggest that FABP4 depletion inhibits colorectal cancer progression by modulating cell growth, stemness, glycolysis and apoptosis. This regulation occurs through the ROS/ERK/mTOR pathway.

  • Review
    Siresha Bathina, Undurti N Das
    Discovery Medicine. 2023, 35(178): 653-663. https://doi.org/10.24976/Discov.Med.202335178.64

    Mitochondria-associated membranes (MAMs) play a significant role in multiple cellular processes including lipid metabolism and neuronal survival. Fatty acids constitute 80% of the dry mass of the brain and are vital for life. Apart from mitochondrial β-oxidation, fatty acids are metabolized in part by peroxisomes to regulate the generation of acyl Coenzyme A and adenosine triphosphate (ATP). Ablation of mitochondria and its associated genes tether endoplasmic reticulum (ER)-Mitochondria contact and results in loss of function leading to aberrant lipid metabolism. Additionally, an increase in reactive oxygen species (ROS) levels along with free radicals' generation may lead to alteration in the integrity of membrane phospholipids, proteins, and DNA. Hence, it is critical to understand the effect of structural and functional aspects of mitochondria on lipid homeostasis. This review explains the role of mitochondrial dysfunction in lipid metabolism and its impact on various neurodegenerative diseases and metabolic disorders.

  • Article
    Jun Chen, Cheng Lei Xia, Rui Dong, Xian Guo Liu, Jing Xia
    Discovery Medicine. 2023, 35(178): 777-786. https://doi.org/10.24976/Discov.Med.202335178.72

    Background: Doxorubicin (Dox) is a clinical first-line broad-spectrum anticancer agent. A dose-dependent cardiotoxic and myelosuppressive response limits the clinical use of Dox. Recent research indicates that Dox-induced cardiotoxicity is associated with senescent cell accumulation and that antiaging therapy can alleviate aging-related disorders. Cepharanthine (Cep) is commonly used to treat various acute and chronic illnesses, including leukopenia, snakebites, dry mouth, and hair loss. Whether Cep alleviates Dox-induced senescence is unknown.

    Methods: The expression of genes and proteins associated with aging was examined using NIH3T3 cell lines. The experiments were divided into a control group, a Dox group, and a Cep group on different days. NIH3T3 senescent cells were detected by senescence-β-galactosidase (SA-β-Gal) staining, and Western blotting was used to detect the protein levels of p16, p53, AMP-activated protein kinase (AMPK), mammalian target of the rapamycin (mTOR), p62, and Light Chain 3 (LC3). Fluorescence was used to detect the expression of monomeric red fluorescence protein-green fluorescence protein-Light Chain 3 (mRFP-GFP-LC3) and LC3 puncta in NIH3T3 cells. Real-time quantitative reverse transcription polymerase chain reaction (RT‒qPCR) was used to test the expression of senescence-associated secretory phenotypes (SASP: Interleukin 6 (IL-6), Interleukin 1 beta (IL-1β), and Interleukin 8 (IL-8)). Cell Counting Kit-8 (CCK-8) was used to assess NIH3T3 cell viability.

    Results: Here, we reported that Cep reversed the Dox-induced increase in the proportion of SA-β-Gal-positive cells and the high expression of aging-related proteins (p53, p < 0.05; p16, p < 0.05) and aging-related genes (IL-6, p < 0.05; IL-1β, p < 0.05; IL-8, p < 0.05) on the 3rd day. Mechanistically, Cep reduced the increase in the levels of phospho-mTOR (p < 0.05) on Days 1 and 3 and p62 protein (p < 0.05) caused by Dox on Day 1 and reversed the decline in LC3II/LC3I levels (p < 0.05) caused by Dox on Day 3, which is associated with the regulation of senescence. Additionally, the viability of NIH3T3 cells was significantly increased in the concentration range of 0.5–5 μM Cep (p < 0.05).

    Conclusions: We first found that Cep could suppress SA-β-Gal activity (p < 0.05) and the development of SASP. Additionally, in Cep-treated cells, Cep could restore autophagy dysfunction and suppress the mTOR signaling pathway. This research provides a new view on the mechanics of aging and autophagy and aids in developing novel antiaging drugs.

  • Article
    Qing Tang, Anli Xu, Ying Yang, Yunmei Zhang, Jianan Sun
    Discovery Medicine. 2023, 35(176): 208-220. https://doi.org/10.24976/Discov.Med.202335176.22

    Background: The emergence of chemotherapy resistance usually causes therapeutic failure in advanced cervical cancer. Forkhead box protein M1 (FOXM1) and threonine tyrosine kinase (TTK) are closely associated with cancer drug sensitivity, but the mechanism of FOXM1 on TTK involvement in chemo-treated cervical cancer remains unclear. Here, we aimed to observe the effects of FOXM1 on TTK and on chemotherapy sensitivity in cervical cancer.

    Methods: The expressions of FOXM1 and TTK in cervical cancer tissues and para-cancerous tissues were analyzed by immunohistochemistry. SiHa and Hela cells were transfected with human lentivirus-FOXM1, small interfering RNA (siRNA) or pcDNA3.1/FOXM1 to analyze the changes in TTK protein expression. Furthermore, the cells were treated with paclitaxel (8 μM) or cisplatin (10 μM) to analyze the effects of FOXM1 on chemotherapy sensitivity. SiHa cells were used to construct a xenograft model to study the effects of FOXM1 expression in response to paclitaxel treatment. The tumor size and weight were observed. The expressions of Ki-67, FOXM1, and TTK protein in tumor tissues were measured by immunohistochemistry.

    Results: High expression of FOXM1 and TTK were found in the cervical cancer tissues (p < 0.05). The TTK protein expressions were decreased by FOMX1-siRNA transfection in SiHa and Hela cells (p < 0.01). The cell viability and cell cycle were also suppressed by FOMX1-siRNA transfection (p < 0.01) but enhanced by pcDNA3.1/FOXM1 transfection (p < 0.01). For paclitaxel or cisplatin treatment, the cell viability and cell DNA damage were improved due to the FOXM1 overexpression (p < 0.01). TTK inhibitor significantly suppressed the effects of FOXM1 overexpression (p < 0.01).

    Conclusions: FOXM1 regulated TTK and affected the therapeutic efficacy of cisplatin and paclitaxel in cervical cancer.

  • Review
    Can Zhang
    Discovery Medicine. 2023, 35(178): 757-776. https://doi.org/10.24976/Discov.Med.202335178.71

    Alzheimer's disease (AD) is a progressive neurodegenerative disorder and the primary cause of dementia in the elderly. The research on AD has markedly evolved since discovering the pathological hallmarks in the brain over the past years. However, the etiology of AD has not been completely elucidated, and presently there is no cure for AD. Furthermore, despite the disease hallmarks commonly manifested in all AD cases, considerable evidence suggests complex heterogeneity of AD pathogenesis. Specifically, studies have identified several disease genes that cause or associate with AD and discovered multiple modifiable risk factors of AD, which may further aggregate the complex etiology of AD. Understanding molecular mechanisms of AD genes and modifying risk factors of AD are making it possible to develop effective interventions to prevent or cure the disease, which may in turn help understand the disease etiology. Recently, there has been keen interest in carrying out multidisciplinary investigations on AD, e.g., the studies on the molecular basis in association with neural network abnormalities in AD. Furthermore, the molecular findings on the pathogenetics of AD have provided translational proof-of-concept to intervene in AD, which has inspired clinical trials by reducing disease neuropathology related to AD genes or risk factors of AD. This article's main aim is to highlight recent progress on the mechanisms and approaches with an attempt to gain an integrated perspective of AD etiology. We envision that this article and its referenced reports may provide a valuable point of reflection that may not only acknowledge past discoveries, but also stimulate future studies, advancing the understanding of the complex etiology of AD and the development of effective therapeutics for AD.

  • Article
    Jin Li, Yanyun Ruan, Chenhui Zheng, Yue Pan, Bangyi Lin, Qi Chen, Zhibao Zheng
    Discovery Medicine. 2023, 35(174): 45-56. https://doi.org/10.24976/Discov.Med.202335174.6

    Background: The aberrant expression of adipocyte enhancer binding protein 1 (AEBP1) has been observed in many cancers and it seems to be involved in the tumorigenesis, progression, and metastasis in numerous tumor types. However, the contribution of AEBP1 in breast cancer (BCa) remains inexplicable.

    Methods: Information related to the diagnostic significance and expression of AEBP1 in BCa was obtained from the public dataset Kaplan–Meier Plotter (http://kmplot.com/analysis/) and the dataset UALCAN (https://ualcan.path.uab.edu/index.html). The MTT (methyl thiazolyl tetrazolium) assay, colony formation assay, Transwell® assay, and FACS (fluorescence-activated cell sorting) assay were used to detect the proliferation, invasive and apoptotic ability of cells before and after treatment. In addition, we constructed an AEBP1 overexpression vector and silenced AEBP1, combined with Real-Time Quantitative Reverse Transcription PCR (qRT-PCR), western blot, immunohistochemistry and TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling) assay to investigate the prognostic significance, biological functions and potential mechanisms of AEBP1 in BCa.

    Results: Higher expression of AEBP1 mRNA (message RNA) was observed in BCa patients with later-stage, who obtained poorer overall survival. Meanwhile, compared with adjacent noncancerous tissues, AEBP1 protein expression was dramatically upregulated in the BCa ones. Furthermore, overexpressed AEBP1 enhanced cell proliferation, migration, invasion, and blocked cell apoptosis in BCa cells. Moreover, the research certificated that AEBP1 upregulated the expression of MMP (matrix metalloproteinase)-2, 9, vimentin, N-cadherin (neural-cadherin), phosphorylation of ERK (extracellular signal-regulated kinase), Smad2/3 (Abbreviated from Sma for nematode and Mad for Drosophila) and AKT (V-akt murine thymoma viral oncogene homolog), while down-regulated the expression of E-cadherin (epithelial cadherin) and PTEN (phosphatase and tensin homolog deleted on chromosome 10). To inhibit cell apoptosis, enforced expression of AEBP1 effectively blocked the cleavage of caspase 9 and p53 (protein 53) and promoted the expression of anti-apoptotic protein Bcl-2 (B-cell lymphoma-2). Finally, AEBP1 accelerated subcutaneously transplanted tumor growth in nude mice by increasing the expression of the cell proliferation biomarker ki67, the phosphorylation of AKT, and blocked apoptosis in vivo.

    Conclusions: In summary, these data suggested the important role of AEBP1 in the BCa progression, which could be used as a potential biomarker for prognostic hallmark and a novel therapeutic strategy.

  • Article
    Xiaoyuan Ning, Dongping Luo, Yong Chen, Yeqing Shao, Jiayun Xu
    Discovery Medicine. 2023, 35(176): 372-382. https://doi.org/10.24976/Discov.Med.202335176.38

    Objective: Mesangial proliferative glomerulonephritis (MPGN) is a prevalent form of primary glomerulonephritis, distinguished by the proliferation of mesangial cells and the accompanying inflammatory response. Baicalin, the active ingredient in the Scutellaria baicalensis Georgi plant, has been observed to have a protective effect on the kidneys. However, its specific impact on MPGN has yet to be studied widely. Hence, this study aimed to investigate the effect on MPGN and the underlying mechanisms of Baicalin.

    Methods: Thirty-six Sprague-Dawley (SD) rats, aged 6 to 8 weeks, were randomly allocated into different subgroups: control, model, benazepril, and three baicalin subgroups (low, medium, and high dose), each consisting of six rats. The concentrations of 24-hour urinary protein, blood urea nitrogen (BUN), serum creatinine (SCr), triglycerides (TG), total cholesterol (TC), interleukins (IL-1α, IL-2, IL-10), and interferon-γ (IFN-γ) were measured with biochemistry. The pathological alterations in the renal tissue were examined using Hematoxylin and Eosin (HE) along with Periodic Acid-Schiff (PAS) staining. Concurrently, the extent of apoptosis was evaluated using TdT-mediated dUTP nick end labeling (TUNEL) staining. In vitro, mesangial cells were exposed to 30 μg/mL lipopolysaccharide for 24 h, with or without varying concentrations of baicalin (10, 20, 40 μM). MTT assay was applied to estimate cell activity, flow cytometry to evaluate the cell cycle, and 5-ethynyl-2-deoxyuridine (EdU) detection to measure cell proliferation. IL-1α, IL-2, IL-10, and IFN-γ concentrations in the cell supernatant were assayed with biochemistry. Furthermore, the expression of apoptosis-related proteins, concluding BCL2-Associated X (Bax), Bcl-2, NOD-like receptor thermal protein domain associated protein 3 (NLRP3), and caspase-1, NF-E2-related factor 2/antioxidant response element (Nrf2/ARE) pathway-related proteins (Nrf2 and HO-1), and phosphatidylinositol 3 kinase/protein kinase B (PI3K/AKT) pathway-related proteins (p-PI3K, PI3K, p-AKT, and AKT) in both the renal tissue and cell supernatant were measured.

    Results: Baicalin treatment significantly reduced the 24-hour urinary protein, serum levels of BUN, SCr, TG, TC, IL-1α, IL-2, IL-10, and IFN-γ in vivo experiments. Baicalin treatment also improved the pathological condition of renal tissue and decreased the occurrence of apoptosis. In vitro, findings confirmed that baicalin inhabits the proliferation of mesangial cells triggered by Lipopolysaccharide (LPS), induces a G1 phase cell cycle arrest, and reduces the concentrations of IL-1α, IL-2, IL-10, and IFN-γ. Baicalin also decreased the ratios of p-PI3K/PI3K and p-AKT/AKT while enhancing the levels of Nrf2 and HO-1 in both renal tissue and cell supernatant.

    Conclusions: Baicalin can mitigate MPGN by impeding the proliferation and inflammation of mesangial cells by activating Nrf2/ARE and PI3K/AKT pathways.

  • Article
    Shan Tan, Rui Chao
    Discovery Medicine. 2023, 35(176): 300-311. https://doi.org/10.24976/Discov.Med.202335176.31

    Background: The high rate of the recurrence and metastasis of osteosarcoma (OS) is the major cause of its poor prognosis. There is a strong correlation between tumor-associated neutrophils (TANs) and tumor progression, progression, and metastasis. This study aimed to identify potential markers that could predict OS metastasis based on analysis of TANs in the tissues of OS patients.

    Methods: A single-cell sequencing dataset (GSE152048), containing seven primary OS lesions, two recurrent OS lesions, and two lung metastatic OS lesions was used for TANs subset identification using the R software (version 4.1.0, R Project for Statistical Computing, Vienna, Austria; https://www.r-project.org/). Immune cell infiltration and immune score were analyzed using CIBERSORT algorithm and ESTIMATE database, respectively. The differentially expressed genes (DEGs) of TANs were used for weighted gene co-expression network analysis (WGCNA) to screen key genes associated with OS metastasis. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) functional enrichment analysis were used to analyze the functions and signaling pathways involved in key genes. The mRNA levels of protein phosphatase 2 regulatory subunit B'gamma (PPP2R5C) were validated in human osteosarcoma cell lines U2-OS and MG63, human normal cervical endometrial cell line HUCEC, and human foreskin fibroblast (HFF-1) cell line by real-time qPCR (RT-qPCR). PPP2R5C-siRNA991 was transfected into U2-OS and MG63 for 48 h, then the expression levels of PPP2R5C, AKT serine/threonine kinase (AKT), and phospho-AKT (p-AKT) were determined by RT-qPCR and Western blotting. Cell proliferation, migration, and apoptosis were measured by cell counting kit-8 (CCK-8), Transwell, and flow cytometry, respectively.

    Results: We identified TANs subsets in primary, metastatic, and recurrent OS. Immune infiltration analysis showed that TANs were expressed in OS. Compared with non-metastatic OS, metastatic OS had lower stromal score, immune score, ESTIMATE score, and higher tumor purity. WGCNA classified DEGs into five clusters, according to their function and identified PPP2R5C, protein phosphatase 2 regulatory subunit B'epsilon (PPP2R5E), tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein gamma (YWHAG) and CREB binding protein (CREBBP), as potential markers that may affect TANs-induced OS metastasis via hypoxia inducible factor 1 (HIF-1), phosphatidylinositol 3-kinases (PI3K)-AKT and Janus kinase (JAK)-signal transducers and activators of transcription (STAT) signaling pathways. In vitro experiments demonstrated that the mRNA and protein expressions of PPP2R5C, PPP2R5E, YWHAG, and CREBBP were highly expressed in U2-OS and MG63 cells (p < 0.01). Furthermore, PPP2R5C reduced proliferation and migration (p < 0.01) and increased apoptosis and p-AKT protein levels in U2-OS and MG6 cells (p < 0.01).

    Conclusions: PPP2R5C affects OS metastasis via PI3K/AKT pathway, which may be a potential marker for OS metastasis and recurrence.

  • Article
    Jing Xia, Yu Zhou, Siyue He, Manoj Kumar Vashisth, Huijie Jia, Qianlong Dai, Yufen He, Xiaobo Wang
    Discovery Medicine. 2023, 35(176): 264-274. https://doi.org/10.24976/Discov.Med.202335176.27

    Background: Amonafide (Amo), due to hematotoxicity and digestive tract symptoms, the clinical application of which is limited. Several studies have reported that chemotherapy side effects are closely related to cellular senescence accumulation. Our study aims to examine whether amonafide causes senescence in human umbilical vein endothelial cell (HUVEC) lines and investigate its mechanisms associated with senescence.

    Methods: The experiments of expression of genes and proteins associated with aging were carried out with HUVEC cell lines. The experiments were divided into a control group and an amonafide group with different days. The HUVEC senescence cells were detected by SA-β-Gal staining, Western blotting detected the protein levels of p16, p53, AMPK (Adenosine 5'-Monophosphate (AMP)-Activated Protein Kinase), mTOR (mechanistic Target of Rapamycin), p62, and LC3 (microtubule-associated protein1 light chain 3, MAP1LC3). Fluorescence detected the expression of mRFP (monomeric Red Fluorescent Protein)-GFP (Green Fluorescent Protein)-LC3 and LC3 puncta of HUVEC cells. RT-qPCR (Real-Time Quantitative Polymerase Chain Reaction) tested the expressions of p53, p21, IL (Interleukin)-1β, IL-6 (Interleukin-6), IL-8 (Interleukin-8), and MCP-1 (Monocyte Chemoattractant Protein-1). CCK-8 (Cell Counting Kit-8) assessed the HUVEC cell viability.

    Results: Here, we reported that amonafide resulted in an increased proportion of SA-β-Gal positive cells, high expression of aging-related proteins (p53 p < 0.05; p16 p < 0.05), and aging-related genes (p53 p < 0.05; p21 p < 0.05; IL-1β p < 0.05; IL-6 p < 0.05; IL-8 p < 0.05; MCP-1 p < 0.05) on the 3rd day. Mechanistically, amonafide could cause an increase in the levels of the mTOR (p < 0.05) on days 1 and 3, and p62 protein (p < 0.05) on day 1, and a decline in LC3II (microtubule-associated protein1 light chain 3Ⅱ)/LC3I levels (p < 0.05) on day 3, which is associated with the regulation of senescence. Additionally, the viability of HUVECs (human umbilical vein endothelial cells) was significantly inhibited by amonafide starting with a concentration of 0.8 μm (p < 0.05).

    Conclusions: We first discovered that amonafide caused normal cellular senescence in our experiments. Amonafide-induced cellular aging by inhibiting autophagy and activating the mTOR pathway. The findings may offer new strategies for managing adverse reactions to amonafide.

  • Article
    Wang Luo, Yanbin He, Jianhui Xu, Shuhua Zhang, Chunxi Li, Jiangfeng Lv, Youfeng Shen, Zhao Ou, Hangming Dong
    Discovery Medicine. 2023, 35(176): 332-342. https://doi.org/10.24976/Discov.Med.202335176.34

    Background: It is common to obtain a low detection rate and unsatisfactory detection results in complex infection or rare pathogen detection. This retrospective study aimed to illustrate the application value and prospect of the third-generation sequencing technology in lower respiratory tract infection disease.

    Methods: This study recruited 70 patients with lower respiratory tract infection (LRTI). Pathogen detection of bronchoalveolar lavage fluid (BALF) from all patients was performed using nanopore metagenomic sequencing technology and traditional culture. BALF culture combined with quantitiative PCR (qPCR) was used as a reference standard to analyze the sensitivity and specificity of nanopore sequencing technology. The current study also collected the examination results of enrolled samples using technical methods sputum culture, tuberculosis DNA (TB-DNA), and Xpert MTB/RIF and analyzed the detection efficiency of nanopore sequencing for Mycobacterium tuberculosis.

    Results: The positive rates of pathogens in 70 BALF samples detected by conventional culture and nanopore sequencing were 25.71% and 84.29%, respectively. Among the 59 positive BALF cases using nanopore sequencing, a total of 31 pathogens were identified, of which the proportions of bacteria, fungi, viruses, and other pathogens were 50%, 17%, 32%, and 1%, respectively. Using the results combined with culture and qPCR detection methods as the standard, the pathogen detection of BALF using nanopore sequencing had a sensitivity of 70% and a specificity of 91.7%. Additionally, the positive rate of the detection of M. tuberculosis using nanopore sequencing was 33.3% (6/18). The clinical medication plans of 74.3% (52/70) of the patients were referred to the nanopore sequencing results, of which 31 cases changed their treatment strategy, 21 supported the previous treatment plans, and 90% (47/52) of the patients finally had clinical improvement.

    Conclusions: BALF detection using nanopore sequencing technology improves the process of detecting pathogens in patients with LRTI, especially for M. tuberculosis, fungi, and viruses, by reducing the report time from three days to six hours. The clinical application prospect of nanopore sequencing technology is promising in the pathogen diagnosis of LRTI.

  • Review
    Ilya Ulasov, Vaishali Singh, Anastasia Laevskaya, Peter Timashev, Rajesh Kumar Kharwar
    Discovery Medicine. 2023, 35(177): 458-475. https://doi.org/10.24976/Discov.Med.202335177.47

    Glioblastoma multiforme is one of the most widespread and dangerous forms of brain tumor with high inflammation. The tumor microenvironment comprises diverse tumor cells, different types of immune cells, and the extracellular matrix. Inflammatory mediators like chemokines, cytokines, and growth factors possibly serve as a capable therapeutic target to quash their tumor-promoting properties in glioblastoma multiforme (GBM). Cytokines are a heterogeneous group of soluble functional proteins which are also associated with the induction and progression of tumors. These are supposed to have both pro-inflammatory (such as tumor necrosis factor-α (TNF-α), interleukin-17A (IL-17A), interferon-γ (IFN-γ), IL-4, IL-2, IL-6, IL-12, IL-13) and anti-inflammatory (such as transforming growth factor-β (TGF-β), IL-10, and granulocyte-macrophage colony-stimulating factor (GM-CSF)) actions and are the crucial communications channels in the tumor microenvironment. In the present minireview we discuss the tumor microenvironment and inflammatory mediators and focus on the involvement of cytokines in establishing communication with the tumor microenvironment. The presented data highlight the possible roles of cytokines in communication between glioblastoma cells and tumor microenvironment. Cytokines formed by immune cells protect the host organs while cytokines secreted by tumor cells are used for their advantage. Though the clinical trials with a number of immunotherapeutic agents are going on around the globe, there is still a requirement for thorough investigation of the regulatory mechanism managing GBM growth, recurrence, and tumor response to the therapy.

  • Article
    Xuan Zhou, Wenting Wang, Li Liu
    Discovery Medicine. 2023, 35(179): 1086-1092. https://doi.org/10.24976/Discov.Med.202335179.105

    Background: Pancreatic cancer (PC), a commonly recognized malignancy, arises within the digestive tract. Somatostatin (SOM) is a regulatory peptide that acts on secretion in vivo. Several studies have shown that SOM has inhibitory effects on various cancers. This work aims to probe the inhibitory effect, and mechanism of SOM action, on the epithelial–mesenchymal transition (EMT) of PC cells.

    Methods: First, the effects of SOM and transforming growth factor-β (TGF-β) on the proliferation of PC cells was determined by Cell Counting Kit-8 (CCK-8) assay. Next, we assessed the impact of SOM and TGF-β on the metastasis and apoptosis of PC cells using transwell assays and flow cytometry. Finally, we evaluated the effects of SOM and TGF-β on the expression of EMT-related proteins, apoptosis-related proteins, and proteins related to the TGF-β/Smad signaling pathway in PC cells using western blot analysis.

    Results: SOM suppressed the growth and metastasis of PC cells, and facilitated their apoptosis (p < 0.05). Moreover, SOM reversed pro-apoptotic effects of TGF-β (p < 0.05). Specifically, SOM increased the expression of Cysteine-aspartic acid protease 3 (Caspase-3) and Bcl-2-associated X protein (Bax) proteins while reducing the expression of B-cell lymphoma 2 (Bcl-2) protein (p < 0.05). SOM also reversed the TGF-β-induced EMT process. The TGF-β1, Smad2, and Smad3 proteins in PC cells treated with SOM were significantly down-regulated (p < 0.05).

    Conclusions: SOM suppressed the EMT progression in PC cells through its regulation of the TGF-β/Smad signaling pathway.

  • Article
    Guoyu Wang, Jie Zhu, Zengxian Wang, Zuliang Xu, Yiming Shi, Lingjie Luo
    Discovery Medicine. 2023, 35(176): 418-428. https://doi.org/10.24976/Discov.Med.202335176.42

    Objectives: To study the effects of curcumin on the proliferation, invasion, apoptosis, and radiosensitivity of the radioresistant nasopharyngeal carcinoma (NPC) C6661-IR strain as well as the potential radiosensitization mechanism.

    Methods: NPC cells were continuously irradiated with different intensities of radiation to induce radiation-resistant cell lines. A plate clone formation assay was used to evaluate the effect of curcumin on the radiosensitivity of NPC cells. 3-(4,5-Dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide thiazolyl blue (MTT) assay was conducted to detect changes in cell viability. Flow cytometry was employed to analyze apoptosis percentage as well as Transwell® assay and immunofluorescence assay to observe cell invasion. Western blotting was applied to detect the expression levels of Bax, Bcl-2, and pro/cleaved-caspase 3. MiR-205-5p mimics and si-TP53INP1 were synthesized and transfected into C6661-IR cells, and the cells were then incubated with 10 μm/L curcumin. Real-time quantitative reverse transcription PCR (RT-qPCR) was used to measure miR-205-5p levels and western blotting was conducted to detect the expression of TP53INP1.

    Results: The optimal radiation dose of X-ray was 6 Gy, and this dose was used in all subsequent experiments. Curcumin treatment significantly inhibited the proliferation and invasion of C6661-IR cells, promoted apoptosis and enhanced radiosensitivity. Compared to the 0 Gy+Cur group and the 6 Gy+Cur group, the miR-205-5p levels were higher in the C6661-IR cells of the 0 Gy and 6 Gy groups. Moreover, miR-204-5p was found to directly target TP53INP1. Curcumin downregulated miR-205-5p levels and upregulated TP53INP1 expression (p < 0.05). Thus, modulation of miR-205-5p or TP53INP1 expression attenuates the biological effects of curcumin on C6661-IR cells.

    Conclusions: Curcumin inhibited the proliferation and invasion of C6661-IR, promoted apoptosis, and enhanced its radiosensitivity to X-rays by mediating miR-205-5p/TP53INP1 expression.

  • Article
    Yun Shi, Fengying Hu, Hao Fu, Shaojie Li, Chengzhi Lu, Chunxiu Hu
    Discovery Medicine. 2023, 35(177): 483-491. https://doi.org/10.24976/Discov.Med.202335177.49

    Background: Lenvatinib is an oral tyrosine kinase inhibitor (TKI), and has been applied in the clinical trials for the treatment of hepatocellular carcinoma (HCC). The function of 5-aminolevulinic acid hydrochloride (ALA) treatment in protecting cardiomyocytes under lenvatinib stimulation was investigated.

    Methods: H9c2 cells were treated with 2 mg/mL lenvatinib for 48 h and 1 mM ALA in the lenvatinib with low dose 5-aminolevulinic acid treatment group (LL) group, 10 mM ALA in the lenvatinib with high-dose 5-aminolevulinic acid treatment group (LH) group and cells without treatment were used as an internal control. C57/BL mice were treated with 10 mg/kg lenvatinib and 200 mg/kg ALA in the LL group and 400 mg/kg ALA in the LH group by gavage once per day for 4 weeks. The proliferation ability of cells was detected using the methyl thiazolyl tetrazolium (MTT) assay. Target gene expression was calculated through real-time quantitative PCR (qPCR), and target protein expression was calculated through Western blotting analysis. The concentrations of cardiovascular protective factors were detected using enzyme linked immunosorbent assay (ELISA).

    Results: In these experiments, 10 mM ALA significantly increased the viability rate of cardiomyocytes (105.4 ± 8.0%) compared with the single lenvatinib treatment group (73.2 ± 6.5%). We also noticed that activation of nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase-1 (HO-1) and phosphatidylinositol-3-kinase (PI3K)/protein kinase B (AKT)/mammalian target of rapamycin (mTOR) pathways were activated after low-dose ALA treatment. 5-ALA treatment led to the downregulation of intercellular cell adhesion molecule-1 (ICAM-1) (0.81- and 0.71-fold), vascular cell adhesion molecule (VCAM) (0.63- and 0.66-fold), angiotensin I (ANGI) (0.88- and 0.66-fold), ANGII (0.66- and 0.48-fold) and upregulation of endothelial nitric oxide synthases (eNOS) (1.25- and 1.89-fold) compared with non 5-ALA treatment group.

    Conclusions: With more experiments on animal models, low-dose of ALA treatment might be a therapeutic strategy to alleviate the damage to cardiomyocytes induced by lenvatinib.

  • Article
    Mikayel Ginovyan, Anush Babayan, Anahit Shirvanyan, Alvard Minasyan, Meri Qocharyan, Barbara Kusznierewicz, Izabela Koss-Mikołajczyk, Nikolay Avtandilyan, Anne Vejux, Agnieszka Bartoszek, Naira Sahakyan
    Discovery Medicine. 2023, 35(177): 590-611. https://doi.org/10.24976/Discov.Med.202335177.59

    Background: Herbal medicinal products containing Vaccinium myrtillus L. (bilberry) fruits and fruit extracts are widely available in the market. Although bilberry leaves and stems are considered as bio-waste, they contain much higher levels of phenolic compounds than fruits. The study aimed to investigate the antimicrobial and anticancer potential of aerial part extracts from Vaccinium myrtillus L. (V. myrtillus, VM) plants harvested at high altitudes in Armenian landscape and characterize the bioactive phytochemicals.

    Material and Methods: For evaluation of antioxidant properties, chemical-based tests (total phenolic and flavonoid content, and antiradical activity in 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and 2,2-diphenyl-1-picrylhydrazyl (DPPH) tests) and cellular antioxidant activity (CAA) assay were applied. Genotoxicity and anticancer properties of the extract alone and in combination with fluorouracil were explored in human cancer and normal cell lines. Antibacterial properties of V. myrtillus extract alone and in combination with antibiotics, as well as their effect on proton-flux rate through cell membrane were explored on bacterial strains. The characterization of active phytochemicals was done using Liquid Chromatography-Quadrupole-Orbitrap High-Resolution Mass Spectrometry (LC-Q-Orbitrap HRMS).

    Results: The V. myrtillus aerial part extract demonstrated promising antioxidant properties in all tests. The selective cytotoxic activity was documented against various cancer cell lines (human colon adenocarcinoma (HT29), human breast cancer (MCF-7) and human cervical carcinoma (HeLa)), while it did not inhibit the growth of tested human normal primary renal mixed epithelial cells (HREC) even at 10-fold higher concentrations. The extract did not have genotoxic properties in comet assay making it a potential source for the development of anticancer preparations. The investigated extract did not directly inhibit the growth of Escherichia coli (E. coli) and Salmonella typhimurium (S. typhimurium) strains at up to 1 mg/mL concentration. However, V. myrtillus extract enhanced the kanamycin intake and increased its efficiency against E. coli strain. The phytochemical characterization of the extract showed the presence of different groups of phenolics.

    Conclusions: Based on obtained data, we suggest the aerial parts of the V. myrtillus plant as an alternative source of bioactive natural products for food supplements, nutraceuticals, functional foods and medicine.

  • Article
    Wanruchada Katchamart, Suppavich Kieattisaksopon, Pongthorn Narongroeknawin, Weerasak Muangpaisan, Narittaya Varothai
    Discovery Medicine. 2023, 35(176): 436-443. https://doi.org/10.24976/Discov.Med.202335176.44

    Background: Sarcopenia is a common condition that can occur in people with chronic inflammatory diseases, including rheumatoid arthritis (RA). The aim of this study was to determine the prevalence and factors associated with this condition in patients with RA.

    Methods: This prospective cross-sectional study was conducted on 182 adult patients with RA. They were diagnosed with sarcopenia using the Asian Working Group's 2019 update on sarcopenia diagnosis. The body composition was estimated using a body impedance analyzer. Physical performance and muscle strength were evaluated with six-meter walk test and hand grip dynamometer, respectively. The Disease Activity Score (DAS) 28 and the Health Assessment Questionnaire (HAQ) were used to assess disease activity and functional status, respectively.

    Results: The majority (87.4%) were female with a mean age (SD) of 59.2 (10.2) years. They had been suffering from RA for a long time (median disease duration [Interquartile range (IQR)] 11 [6–16] years) and had mildly active disease [mean DAS28 (SD) 2.61 (0.83)] with slightly functional disability [median HAQ (IQR) 0.34 (0–0.65)]. Of these, 26.4% had sarcopenia. Advanced age [relative risk (RR) 1.07 (95% confidence interval (CI) 1.02–1.11), p = 0.002], low body mass index (BMI) [RR (95% CI) 0.81 (0.72–0.90), p < 0.001], high disease activity [RR (95% CI) 1.64 (1.22–2.12), p = 0.045], and depression [RR (95% CI) 1.18 (1.01–1.37), p = 0.04] were independently associated with sarcopenia.

    Conclusions: Sarcopenia was found to be common in Thai RA, and its independent risk factors are age, disease activity, BMI, and depression. Well-controlled disease activity may be beneficial for preventing or minimizing sarcopenia and improving patient outcomes.

  • Article
    Xinghai Liu, Tong Liu, Kunli Yang, Hong Liu
    Discovery Medicine. 2023, 35(176): 275-282. https://doi.org/10.24976/Discov.Med.202335176.28

    Background: Asiaticoside is one of the main components of triterpenoid saponins extracted from Centella asiatica. Asiaticoside has shown the effects of wound healing, osteoclastogenesis, anti-inflammatory, anti-cancer, and improving cognition in multiple human disease models. However, studies on the antifatigue effects of asiaticoside have not been explored. Therefore, the aim of this study was to investigate the potential antifatigue effect and underlying mechanism of asiaticoside administration on exhaustive exercise performance.

    Methods: Male Kunming mice were divided into four groups randomly (n = 20/group). Saline (10 mL/kg) was administered to the model control group and the other three experimental groups were fed with low (10 mg/kg), medium (20 mg/kg) and high (40 mg/kg) asiaticoside once/daily for 14 days. The antifatigue effect of asiaticoside on mice was estimated by analyzing changes in body weight, weight-loaded swimming time, rotating time, lactic acid, urea nitrogen, liver/muscle glycogen, serumal superoxide dismutase, superoxide dismutase and the liver tissues of hematoxylin and eosin (H&E) staining.

    Results: The results indicated that no significant differences were observed in the body weight of each group (p > 0.05). Compared with the model control group, supplementation of asiaticoside significantly prolonged the weight-loaded swimming time and rotating time; Decreased the blood lactic acid (LA), blood urea nitrogen (BUN), and serumal malonaldehyde (MDA); And increased the content of liver/muscle glycogen and serumal superoxide dismutase levels (SOD) (p < 0.05). Furthermore, the pathological results of the liver were improved greatly. The maximal effect was observed in the medium group of 20 mg/kg.

    Conclusions: Asiaticoside is capable of reducing the fatigue effect by regulating energy consumption, energy metabolism and improving antioxidant activity after exercise. While there are still some shortcomings in this study, our findings provide a scientific basis for developing an asiaticoside-based antifatigue supplement.

  • Review
    Undurti Narasimha Das
    Discovery Medicine. 2023, 35(177): 451-457. https://doi.org/10.24976/Discov.Med.202335177.46

    Seasonal variation in blood pressure that is higher in winter and lower in summer has been attributed to several factors that include changes in the activity of autonomic nervous system, vasopressin and expression of endothelial nitric oxide synthase (eNOS). Transient receptor potential melastatin 8 (TRPM8), a non-selective Ca2+-permeable cationic channel, serves as a molecular transducer to sense cold by the somatosensory system. TRPM8 is sensitive to protein kinase C (PKC) and phosphatidyl inositol-4,5-biphosphate [PI(4,5)P2] suggesting that TRPM8 is stimulated by phospholipase C (PLC)-coupled receptors. Activated PLC inhibits TRPM8 by reducing cellular PI(4,5)P2 levels and by activating PKC via diacyl glycerol. Bradykinin and prostaglandin E2 (PGE2), which are pro-inflammatory molecules, reduce the responses to cold, suggesting that phospholipase A2 (PLA2), which releases polyunsaturated fatty acids (PUFAs), the precursors of various eicosanoids, from the cell membrane lipid pool can modulate the function of TRPM8. TRPM8 functions as a nociceptor and modulates immune response. These and other studies indicate that cold-induced activation of transient receptor potential melastatin 8 (TRPM8) plays a role in the pathobiology of hypertension, preeclampsia and in the regulation of inflammation and immunity.

  • Article
    Lin Zou, Qin Shi, Yingxuan Li, Zhen Yuan, Li Peng, Jiancan Lu, Hongling Zhu, Junhua Ma
    Discovery Medicine. 2023, 35(177): 612-622. https://doi.org/10.24976/Discov.Med.202335177.60

    Background: The function of flavin containing dimethylaniline monooxygenase 1 (FMO1), which is known to play a part in lipid metabolism, remains unclear in the development of nonalcoholic fatty liver disease (NAFLD). This research has the objective of examining the contributions of FMO1 in the progression of NAFLD and the associated mechanisms, particularly the peroxisome proliferator activated receptor alpha (PPARα) and ferroptosis pathways.

    Methods: An in vitro NAFLD model was established by treating L02 cells with free fatty acids (FFAs). The FMO1 and ferroptosis levels were examined in the cellular NAFLD model. FMO1 was knocked down using short-interfering RNA transfection. The effects of FMO1 knockdown on lipid accumulation, PPARα expression, and ferroptosis were examined in the cellular NAFLD model. Additionally, the effects of FMO1 and/or PPARα overexpression on lipid metabolism and ferroptosis were analyzed. Furthermore, L02 cells were pre-treated with GW7647 (PPARα agonist) or RSL3 (ferroptosis activator) and stimulated with FFAs.

    Results: The levels of FMO1 and ferroptosis were upregulated in the in vitro NAFLD model. FMO1 knockdown suppressed the FFA-induced accumulation of lipids in hepatocytes, downregulation of PPARα expression, and upregulation of ferroptosis. In contrast, FMO1 overexpression dysregulated lipid metabolism and downregulated PPARα levels. Meanwhile, PPARα overexpression mitigated the FMO1 overexpression-induced upregulation of ferroptosis and lipid accumulation. Treatment with RSL3 suppressed the effects of PPARα overexpression on lipid accumulation and FMO1 expression.

    Conclusions: FMO1 upregulates ferroptosis by suppressing PPARα in NAFLD, which leads to the dysregulation of lipid metabolism.

  • Article
    Yichao Wang, Yanhong Shao, Juan Yu
    Discovery Medicine. 2023, 35(176): 353-360. https://doi.org/10.24976/Discov.Med.202335176.36

    Background: Noninvasive prenatal testing (NIPT) has been widely adopted in prenatal examination for fetal chromosomal aneuploidy. The present study aimed to evaluate the clinical features of NIPT for both common trisomy and sex chromosome aneuploidy (SCA).

    Methods: A total of 24,164 pregnant women with NIPT testing from July 2020 to June 2022 were recruited at the Linping Maternity and Child Health Care Hospital.

    Results: Ninety cases showed high risk of trisomy 21/18/13 with karyotype results available, and the sensitivity, specificity, and positive predictive value (PPV) were 98.41%, 99.88% and 68.89%, respectively. The three most important reasons for screening were advanced maternal age (AMA, 28.06%), intermediate risk of prenatal screening (20.34%) and Multiple of medium (MoM) abnormality of prenatal screening (17.38%). High risk of NIPT results with Z-score ≥15 have a higher PPV when compared to those with 3 ≤ Z-score < 10, and 10 ≤ Z-score < 15. Meanwhile, 97 pregnant women received positive results for fetal sex chromosome aneuploidy (SCA) in NIPT. In addition, the rate for further diagnostics of SCA was 64.95% and the PPV of SCA was 50.79%.

    Conclusions: Our data show that NIPT has a promising future in prenatal screening for genetic abnormalities of the fetus, and that the accuracy of NIPT is closely related to Z-score.

  • Article
    Jia-qi Song, Xia Wang, Zhi-min Zeng, Ping-an Liang, Cong-ying Zhong, An-wen Liu
    Discovery Medicine. 2023, 35(176): 321-331. https://doi.org/10.24976/Discov.Med.202335176.33

    Objective: Anti-angiogenic therapy has proven effective in non-small-cell lung cancer (NSCLC) patients. The purpose of this study was to evaluate the efficacy of programmed cell death protein 1 (PD-1) inhibitors combined with anti-angiogenic therapy in patients with driver gene mutation negative NSCLC and brain metastases (BMs).

    Methods: A retrospective analysis was performed on NSCLC BMs in patients without driver gene mutations who received PD-1 inhibitors. Two groups, receiving either PD-1 inhibitor monotherapy or PD-1 inhibitor plus anti-angiogenesis therapy, were identified. The primary endpoints were overall survival (OS) and intracranial progression-free survival (iPFS). The secondary endpoints were safety, intracranial objective response rate (iORR) and intracranial disease control rate (iDCR).

    Results: 113 NSCLC patients were included, 51 (45.1%) in the PD-1 inhibitor monotherapy group and 62 (54.9%) in the PD-1 inhibitor plus anti-angiogenesis therapy group. The median follow-up time was 26.2 months. OS was higher in the combination therapy cohort than in the monotherapy cohort (OS: 21.4 vs. 11.8 months; p = 0.004), with no significant difference in iPFS (p = 0.088). Moreover, the PD-1 inhibitor + anti-angiogenic therapeutic regimen exhibited the preferred iDCR (p = 0.005) but not the iORR (p = 0.121). There was no significant difference in the incidence of grade 3–4 adverse events between the two groups. In multivariate Cox regression analysis, PD-1 inhibitor therapy combined with anti-angiogenic treatment (p = 0.003) was an independent prognostic indicator of OS.

    Conclusions: Combining PD-1 inhibitor therapy with anti-angiogenic treatment significantly improves the OS of driver gene mutation negative NSCLC patients with BMs.

  • Review
    Jeffrey Okojie, Sarah McCollum, Jared Barrott
    Discovery Medicine. 2023, 35(179): 921-927. https://doi.org/10.24976/Discov.Med.202335179.87

    The field of oncology is continuously seeking to find effective treatment therapies with limited side effects. Antibody-drug conjugates (ADCs) are a promising class of cancer therapies that have been shown to be effective with limited side effects. Although promising, these therapies experience shortcomings, such as the stability and reproducibility of current conjugation methods. Historically, ADCs have been produced by stochastic conjugation methods; however, new methods of site-specific conjugation have evolved to mitigate current ADC shortcomings. In this article, we highlight the success of ADCs as well as some of their challenges. We also highlight the shortcomings of stochastic conjugation and explore the various site-specific conjugation methods and their advantages over stochastic conjugation.

  • Review
    Rongping Huang, Zhikun Zhang, Lu Gan, Dianfa Fan, Zhangbo Qian, Xinjun Sun, Yong Huang
    Discovery Medicine. 2023, 35(175): 95-103. https://doi.org/10.24976/Discov.Med.202335175.10

    Hepatocellular carcinoma development and many other tumors are closely related to alpha-fetoprotein (AFP), its determination can be used as a positive test for tumors. It is mainly used clinically as a serum marker to diagnose and monitor the efficacy of primary hepatocellular carcinoma. Therefore, a variety of biosensors have been developed to detect AFP. Electrochemical sensors integrate a variety of detection methods. They have inherent advantages over other types of sensors, they are fast, portable, simple, and highly sensitive. Some meaningful electrochemical biosensors work with nanomaterials acting as signal amplification elements or as signal amplification catalysts. This review introduced the field of biosensors and discuss about the use of nanomaterials in electrochemical sensing, specificity electrochemical biosensing of AFP. The study ends with a discussion about the prospects for nanomaterial-based signal amplification and future research directions.

  • Article
    Jiawen Jiang, Wei Dong, Wen Zhang, Qian Wang, Ruyi Wang, Jiaxu Wang, Hao Wu, Hui Dong, Robert Chunhua Zhao, Jiao Wang, Zhe Li
    Discovery Medicine. 2023, 35(179): 995-1014. https://doi.org/10.24976/Discov.Med.202335179.96

    Background: Hypoxia is a pivotal factor influencing cellular gene expression and contributing to the malignant progression of tumors. Metabolic anomalies under hypoxic conditions are predominantly mediated by mitochondria. Nonetheless, the exploration of hypoxia-induced long noncoding RNAs (lncRNAs) associated with mitochondria remains largely uncharted.

    Methods: We established hypoxia cell models using primary human hepatocytes (PHH) and hepatocellular carcinoma (HCC) cell lines. We isolated mitochondria for high-throughput sequencing to investigate the roles of candidate lncRNAs in HCC progression. We employed in vitro and in vivo assays to evaluate the functions of solute carrier family 1 member 5 antisense lncRNA (SLC1A5-AS). RNA-seq was utilized to scrutinize the comprehensive genome profile regulated by SLC1A5-AS in HCC. Subsequently, quantitative real-time polymerase chain reaction (qRT-PCR) and western blot analysis were utilized to validate the expression of alanine-serine-cysteine transporter 2 (ASCT2, encoded by the SLC1A5 gene), and a glutamine uptake assay was employed to estimate the glutamine uptake capacity of Huh-7 cells after SLC1A5-AS overexpression. To delve into the mechanisms governing the regulation of SLC1A5 expression by SLC1A5-AS, we employed a biotin-labeled SLC1A5-AS probe in conjunction with a western blot assay to confirm the interactions between SLC1A5-AS and candidate transcription factors. Luciferase reporter assays and chromatin immunoprecipitation (ChIP) were utilized to authenticate the effects of the predicted transcription factors on SLC1A5 promoter activity.

    Results: Following the screening, we identified CTB-147N14.6, derived from the antisense strand of the SLC1A5 gene, which we have named SLC1A5-AS. SLC1A5-AS exhibited significantly elevated expression levels in HCC tissue and was associated with poor prognosis in HCC patients. In vitro and in vivo assays revealed that the overexpression of SLC1A5-AS significantly heightened cell invasion and metastasis. RNA-seq data unveiled SLC1A5-AS involvement in glutamine metabolism, left-handed amino (L-amino) acid transmembrane transporter activity, and the nuclear factor kappa-B (NF-κB) signaling pathway. Overexpression of SLC1A5-AS markedly increased ASCT2 mRNA/protein levels, thereby enhancing glutamine uptake and promoting the growth and metastasis of HCC cells. Mechanistically, higher RNA levels of SLC1A5-AS directly bound with myeloid zinc finger 1 (MZF1), acting as a transcriptional repressor, thus diminishing its binding to the SLC1A5 promoter region.

    Conclusions: Our findings unveil a novel role for the lncRNA SLC1A5-AS in glutamine metabolism, suggesting that targeting SLC1A5-AS/MZF1, in conjunction with ASCT2 inhibitor treatment, could be a potential therapeutic strategy for this disease.

  • Article
    Leiting Yang, Han Yan, Yi Liang, Dian Sun, Shijun Lu, Zichun Tang, Ming Ma, Ming Shen
    Discovery Medicine. 2023, 35(176): 383-393. https://doi.org/10.24976/Discov.Med.202335176.39

    Background: Conditioned medium (CM) from human amnion-derived mesenchymal stem cells (hAMSCs) exhibits excellent pro-angiogenic capacity, and circ-100290 participates in this process. Autophagy is involved in the relevant mechanisms of angiogenesis, but it is unclear whether autophagy is related to the pro-angiogenesis effect of hAMSCs. This research sought to determine whether autophagy involved in the process of pro-angiogenesis induced by hAMSCs might be regulated by circ-100290.

    Methods: Upon treatment with CM from hAMSC or 3-methyladenine (3-MA), autophagosomes in human umbilical vein endothelial cells (HUVECs) were observed by transmission electron microscopy. HUVECs' angiogenic ability was evaluated by in vitro assays (transwell, wound healing, tube formation) and an in vivo Matrigel plug assay. Specific small interfering RNAs (siRNA) or inhibitors were used to regulate circ-100290 expression. Additionally, western blot and quantitative reverse transcription-polymerase chain reaction (RT-qPCR) were used to evaluate expression of the following indicators: Beclin-1, LC3-II, matrix metalloproteinase 2 (MMP2), MMP9, vascular endothelial growth factor (VEGF)-A, and endothelial nitric oxide synthase (eNOS).

    Results: Incubation with hAMSC-CM increased autophagy, angiogenesis and the expressions of VEGF-A and eNOS in HUVECs, all of which were inhibited by 3-MA. Knocking down circ-100290 in hAMSC-CM-treated HUVECs reduced Beclin-1 expression and inhibited autophagy. This resulted in lower angiogenesis in the Matrigel plug assay showing that reduced angiogenesis occurred after circ-100290 silencing in hAMSC-CM-treated HUVECs.

    Conclusions: Circ-100290 promotes autophagy-mediated angiogenesis in hAMSC-CM-treated HUVECs.

  • Article
    Zhu Liang, Guoxiong Zeng, Wang Wan, Biao Deng, Chunyuan Chen, Fasheng Li, Guanzhou Lin, Yuying Lin, Haitao Lin, Guixi Mo, Huilai Miao
    Discovery Medicine. 2023, 35(175): 131-143. https://doi.org/10.24976/Discov.Med.202335175.14

    Background: With the wide application of multislice spiral computed tomography (CT), the frequency of detection of multiple lung cancer is increasing. This study aimed to analyze gene mutations characteristics in multiple primary lung cancers (MPLC) using large panel next-generation sequencing (NGS) assays.

    Methods: Patients with MPLC surgically removed from the Affiliated Hospital of Guangdong Medical University from Jan 2020 to Dec 2021 enrolled the study. NGS sequencing of large panels of 425 tumor-associated genes was performed.

    Results: The 425 panel sequencing of 114 nodules in 36 patients showed that epidermal growth factor receptor (EGFR) accounted for the largest proportion (55.3%), followed by Erb-B2 Receptor Tyrosine Kinase 2 (ERBB2) (9.6%), v-Raf murine sarcoma viral oncogene homolog B1 (BRAF), and Kirsten rat sarcoma viral oncogene (KRAS) (8.8%). Fusion target variation was rare (only 2, 1.8%). ERBB2 Y772_A775dup accounted for 73%, KRAS G12C for about 18%, and BRAF V600E for only 10%. AT-rich interaction domain 1A (ARID1A) mutations were significantly higher in invasive adenocarcinoma (IA) which contained solid/micro-papillary malignant components (p = 0.008). The tumor mutation burden (TMB) distribution was low, with a median TMB of 1.1 MUTS/Mb. There were no differences in the TMB distribution of different driver genes. In addition, 97.2% of MPLC patients (35/36) had driver gene mutations, and 47% had co-mutations, mainly in IA (45%) and invasive adenocarcinoma (MIA) (37%) nodule, with EGFR (39.4%), KRAS (9.1%), ERBB2 (6.1%), tumor protein 53 (TP53) (6.1%) predominately.

    Conclusions: MPLC has a unique genetic mutation characteristic that differs from advanced patients and usually presents with low TMB. Comprehensive NGS helps to diagnose MPLC and guides the MPLC clinical treatment. ARID1A is significantly enriched in IA nodules containing micro-papillary/solid components, suggesting that these MPLC patients may have a poor prognosis.

  • Article
    Meng Yin, Xiaosong Qin
    Discovery Medicine. 2023, 35(175): 104-115. https://doi.org/10.24976/Discov.Med.202335175.11

    Background: At present, there is no comprehensive evaluation of the efficacy and safety of low molecular weight heparin (LMWH) for the treatment of thrombophilia during pregnancy in clinical practice. This study aimed to systematically evaluate the efficacy of LMWH in the treatment of patients and its effects on coagulation function, thereby providing a reference for the clinical treatment and prognosis evaluation of thrombophilia during pregnancy.

    Methods: Database PubMed, Web of Science and Embase as well as China National Knowledge Infrastructure and Wanfang Database were applied for the search of data. A comparative study on the efficacy of LMWH in the treatment of gestational thrombophilia was enrolled. Stata 16.0 software (Stata, College Station, TX, USA) was utilized to conduct the meta-analysis.

    Results: A total of 487 relevant articles were retrieved and 14 studies were finally included. Patients in the LMWH combined with the low-dose aspirin group had a significantly higher live birth rate than those in the aspirin or LMWH treat group (OR (odds ratio) = 4.54, 95% CI (confidence interval): 2.76, 7.45). The adverse effects rate was lower in the LMWH combined with the low-dose aspirin group than in the aspirin or LMWH treatment group (OR = 0.40, 95% CI: 0.29, 0.56). After treatment, patients in the LMWH combined with the low-dose aspirin group had significantly lower D-dimer (SMD (standardized mean differences) = –1.50, 95% CI: –2.19, 0.80) and platelet count (PLT; SMD = –0.13, 95% CI: –0.35, 0.09) than those in the aspirin or LMWH treatment group. However, activated partial thromboplastin time (APTT; SMD = 0.16, 95% CI: –0.10, 0.42), thrombin time (TT; SMD = 0.60, 95% CI: –0.14, 1.34), plasma prothrombin time (PT; SMD = 0.42, 95% CI: –0.71, 1.56), and fibrin values (FIB; SMD = –0.92, 95% CI: –2.12, 0.28) were significantly higher in the LMWH combined with low-dose aspirin group than those in the aspirin or LMWH treatment group.

    Conclusions: LMWH heparin combined with low-dose aspirin can effectively correct coagulation function in pregnant women, improve prothrombotic state and increase the live birth rate, which has high clinical value.

  • Article
    Daojun Hu, Li Zhang, Bing Qin, Ning Wang, Xingjun Li, Wenjie Shi
    Discovery Medicine. 2023, 35(179): 1177-1189. https://doi.org/10.24976/Discov.Med.202335179.114

    Background: Previous studies have explored the relationship between serum lead levels and the risk of female breast cancer (FBC). However, it is still uncertain whether urinary lead levels are associated with FBC. This study aimed to investigate the potential association between urinary lead and FBC.

    Methods: A cross-sectional case-control study was conducted using the National Health and Nutrition Examination Survey (NHANES), which is a series of cross-sectional, nationally representative surveys of the United States population consisting of 10 survey waves from 1999 to 2018. This study analyzed a total of 2795 female participants (≥20 years), consisting of 210 participants with FBC and 2585 healthy controls. Urinary lead was detected using Inductively Coupled Plasma-Mass Spectrometry, which was divided into four levels by using quartiles-defining cut points. Multivariate logistic regression was used to analyze the association between urinary lead and FBC.

    Results: Multivariate logistic regression revealed that urinary lead was positively correlated with FBC (Odds ratio [OR], 2.16; 95% confidence interval [CI]: [1.18, 3.95], p < 0.05) in a fully adjusted model. There were significantly increased ORs of FBC in quartile 4 (Q4) and quartile 3 (Q3), compared with the lowest quartile 1 (Q1) (Q4, OR = 1.48, 95% CI [0.89, 2.48]; Q3: OR = 1.01, 95% CI [0.59, 1.73], p for trend = 0.021). No significant interaction effects were observed between urinary lead levels and FBC between the subgroups (age, race, educational status, body mass index (BMI), marital status, family income to poverty ratio, hypertension status, diabetes status, renal function status, smoking history, ever been pregnant, oral contraceptive use, occupation classification, etc.) (All interaction p-value > 0.05).

    Conclusions: Urinary lead is likely positively associated with FBC in the US population.

  • Article
    Yanzhu Wang, Xue Li, Kadiya Abudukeyimu, Wenzheng Du, Yuxian Ning, Xiaoli Qi, Ning Hua, Nan Wei, Gang Ding, Jing Li, Linlin Song, Ying Zhang, Xuehan Qian
    Discovery Medicine. 2023, 35(174): 11-18. https://doi.org/10.24976/Discov.Med.202335174.2

    Background: There are some uncertainties about the effect of low-power red laser treatment on myopia control for anisometropic myopia in children. To evaluate the effect and safety of low-power red laser treatment on refractive development for anisometropic myopia in children, a contralateral comparison study was conducted.

    Methods: The more myopic eye of child with anisometropic myopia was treated with low-power red laser treatment (LRL group), the other eye received no treatment other than the wearing of single-focus spectacles (SFS) (SFS Group). The LRL treatment was given at home under parental guidance for 3 minutes each time, twice daily with a minimal interval of 4 hours, 7 days per week, using an equipment that produces red laser of 650 nm wavelength at an illuminance range of roughly 1200–1800 lux and an energy of 0.60 mw for a 4-mm pupil (class I classification).

    Results: Among 51 included children, 44 (86.27%) completed the 3-months study, consisting of 15 girls (34.1%) and 29 boys (65.9%). After 3-months axial length (AL) and spherical equivalent refraction (SER) progression were –0.08 mm [95% CI (confidence interval), 0.11 to 0.06 mm] and +0.23 diopter (D) (95% CI, 0.13–0.33 D) for LRL group and +0.08 mm (95% CI, 0.05–0.11 mm) and –0.07 D (95% CI, –0.16–0.03 D) for SFS group. AL and SER progression between the groups varied by 0.17 mm (95% CI, 0.13–0.20 mm) and –0.30 D (95% CI, –0.42 to –0.18 D). There was no visible structural damage on optical coherence tomography (OCT) scans.

    Conclusions: AL growth, myopia progression, and anisometropia of the binoculars can all be slowed down by LRL treatment. Compared to SER progression, axial elongation is more accurate and simpler to monitor. LRL treatment unrecorded functional and structural damage of binoculus.

  • Article
    Jie Chen, Lingmin Hu, Yingchun Ling, Yuhua Xu, Xiao Yu, Luping Ma, Mengna Shou
    Discovery Medicine. 2023, 35(175): 168-177. https://doi.org/10.24976/Discov.Med.202335175.17

    Background: Polycystic ovary syndrome (PCOS) is an endocrine disorder that occurs frequently in women of childbearing age and is associated with insulin resistance. Serum visfatin can affect insulin resistance by binding to insulin receptors and further affect the occurrence and development of PCOS. In this study, we investigated the current status of serum visfatin levels in patients with PCOS through a literature search and meta-analysis.

    Methods: We searched online Pubmed, Embase, Web of Science, the Cochrane Library, CNKI (China National Knowledge Infrastructure), CBMdisc (China Biology Medicine disc) databases and registered websites such as the ICTRP (International Clinical Trial Registration Platform) and clinicaltrials.gov (https://clinicaltrials.gov/) for case-control studies on PCOS and visfatin levels, assessed the quality of the included articles with the Newcastle-Ottawa Scale (NOS scale), and combined the comparison of serum visfatin levels between patients with PCOS and healthy individuals from high-quality studies.

    Results: 20 research papers were included in the quantitative analysis of this study. The combined analysis showed that obese patients with PCOS had statistically significantly higher visfatin levels than healthy people [MD (mean difference) = 12.94, 95% CI (confidence interval) (6.52–19.37), Z = 3.95, p < 0.0001]. Visfatin levels were higher in non-obese patients with PCOS than in healthy people and are statistically significant [MD = 14.98, 95% CI (5.80–24.16), Z = 3.20, p = 0.001]. Heterogeneity in the combined analysis was not related to study location, the publication year of the literature, source of serum samples, but was influenced by the quality of the literature. After excluding the most influential papers, the combined analysis was conducted again, and the conclusion was consistent with that before the exclusion. The results of Egger’s test showed no significant publication bias.

    Conclusions: High serum visfatin levels are a natural feature of PCOS and are not associated with obesity; Serum visfatin levels may be a potential marker for the diagnosis of PCOS, but their relationship with PCOS and insulin resistance remains worthy of in-depth investigation.

  • Article
    Wei Li, Yun Cheng, Junchi Cheng, Jincao Yao, Mei Song, Meiying Yan, Yajun Qi
    Discovery Medicine. 2023, 35(174): 19-27. https://doi.org/10.24976/Discov.Med.202335174.3

    Background: The long intergenic non-coding RNA 01614 (LINC01614) is aberrantly expressed in various malignancies, suggesting its role in oncogenesis. However, it has not been well studied in breast cancer.

    Methods: The cancer genome atlas databases (TCGA) and public database of breast cancer gene-expression miner (bc-GenExMiner) were utilized to analyze the prognostic role of LINC01614 in breast cancer. Kaplan–Meier, and Cox regression analyses were conducted for survival analysis. Nomograms were built to predict survival. We used deconvolution-based methods, such as TIMER (Tumor Immune Estimation Resource) and CIBERSORT (cell-type identification by estimating relative subsets of RNA transcripts), to explore the relationship between LINC01614 and immune cell characteristics.

    Results: The very abnormal expression of LINC01614 was found in 14 types of malignancy, including breast cancer. The LINC01614 was significantly overexpressed in human epidermal growth factor receptor 2 (HER2)+, estrogen receptor (ER)+, progesterone receptor (PR)+, and non-triple negative breast cancer (non-TNBC). According to survival analysis, the higher expression of LINC01614 was related with poor survival. The co-expressed genes analysis exhibited that LINC01614 was closely associated with the collagen-associated process and phosphoinositide 3-kinases-protein kinase B (PI3K-Akt) signaling pathway. Moreover, this study has explored the association among LINC01614 expression, tumor-infiltrating immune cells, and the efficacy of chemotherapeutics.

    Conclusions: Our data reveal the expression pattern of LINC01614 in breast carcinoma with different molecular subtypes. The results also indicated that the LINC01614 could be a novel diagnostic and prognostic marker for breast carcinoma.

  • Article
    Chunli Ma, Qing Gao, Li Zhang, Chao Li, Geng Wu, Lei Yang
    Discovery Medicine. 2023, 35(179): 1123-1133. https://doi.org/10.24976/Discov.Med.202335179.109

    Background: Ischemic stroke is an acute cerebrovascular disease with high mortality rates and poor prognoses. The influence of ischemic stroke includes a heavy economic burden to patients and society, making the exploration of new therapeutic targets for preventing and treating ischemic stroke urgent. This study aimed to explore the effect of phosphoglycerate mutase family member 5 (PGAM5) on oxidative stress and mitochondrial dysfunction in ischemic stroke.

    Methods: The model of ischemic neuronal brain injury was established through culturing purchased human neuroblastoma cells (SH-SY5Y) by oxygen-glucose deprivation/reoxygenation (OGD/R). There were six experimental groups, including the OGD/R model group (SH-cells of OGD/R model), OE-NC group (cells of OGD/R model transfected with scramble cDNA), OE-PGAM5 group (cells of OGD/R model transfected with full-length sequence of PGAM5), si-NC group (cells of OGD/R model transfected with negative control small interference (si)RNA), si-PGAM5 group (cells of OGD/R model transfected with siRNA for PGAM5 knockdown), and a control group (cells cultured normally). Cell counting kit-8 (CCK-8) and flow cytometry were used to determine the activity and apoptosis of cells. Subsequently, the effects of PGAM5 expression on oxidative stress and mitochondrial dysfunction were analyzed. Mitochondrial morphology was observed by transmission electron microscopy (TEM), and mitochondrial membrane potential (MMP) was determined by JC-1 fluorescent probe. The levels of reactive oxygen species (ROS) were measured by flow cytometry, and levels of malondialdehyde (MDA) and superoxide dismutase (SOD) were measured by enzyme-linked immunosorbent assay (ELISA) assay. The expression of light chain (LC)3-II/I and autophagy-related gene 5 (ATG5) proteins were measured, and the regulation of PGAM5 expression on PTEN-induced putative protein kinase 1 (PINK1)/Parkin pathway was also explored.

    Results: PGAM5 overexpression in OGD/R cells decreased the cell viability (p < 0.001) while increasing cell apoptosis (p < 0.01) compared to the OGD/R group. Inhibition of PGAM5 expression reversed the decreased cell viability (p < 0.001) and the increased cell apoptosis (p < 0.01). The JC-1 fluorescence showed that OGD/R treatment reduced mitochondrial membrane potential (p < 0.001) and TEM showed an obvious increase in phagosomes. In addition, OGD/R treatment enhanced oxidative stress (increased ROS, p < 0.01; increased MDA, p < 0.001; decreased SOD, p < 0.001), which could be further enhanced by overexpression of PGAM5 (ROS, p < 0.001; MDA, p < 0.001; SOD, p < 0.001) while reversed by the inhibition of PGAM5 (ROS, p < 0.01; MDA, p < 0.001; SOD, p < 0.001). The OGD/R-activated PINK1/Parkin pathway was inhibited by the knockdown of PGAM5 (p < 0.01) but promoted by the overexpression of PGAM5 (p < 0.05).

    Conclusions: PGAM5 stimulates oxidative stress and impairs mitochondrial function in ischemic stroke, and regulates the PINK1/Parkin signaling pathway. Therefore, PGAM5 is likely to be a target for the therapy of ischemic stroke.

  • Article
    Yun Cheng, Xiaoqing Yi, Shuang Fu, Junchi Cheng, Wei Li, Hongliang Xu
    Discovery Medicine. 2023, 35(174): 28-35. https://doi.org/10.24976/Discov.Med.202335174.4

    Background: Long non-coding RNA (lncRNA) AP000695.2 (ENSG00000248538) expresses abnormally in various malignancies, what shows its role as oncogene. However, it has not been extensively studied in gastric cancer. The aim of the current study was to explore the clinical value of AP000695.2 to prognose gastric cancer.

    Methods: The cancer genome atlas (TCGA) and the gene expression profiling interactive analysis (GEPIA) online tool were used to analyze AP000695.2 expression pattern, diagnostic and prognostic role in gastric cancer. Kaplan–Meier and Cox regression analyses were used to assess survival in patients with gastric cancer. Receiver operating curve (ROC) analysis was used to assess AP000695.2 diagnostic capacity. Nomograms were created to predict overall survival (OS) and progression free survival (PFS).

    Results: LncRNA AP000695.2 was abnormally upregulated in 19 types of malignancy, including gastric cancer. Survival analysis indicated that high expression of AP000695.2 was associated with poor survival of gastric cancer. Multivariate Cox regression analysis verified the independent prognostic value of AP000695.2 to predict OS (HR (hazard ratio): 1.104, 95% CI (confidence interval): 1.035–1.178, p = 0.003) and PFS (HR: 1.170, 95% CI: 1.090–1.256, p < 0.001). ROC analysis indicated a favorable AP000695.2 diagnostic capacity (area under the curve (AUC) = 0.890). Nomograms were also constructed for OS and PFS based on AP000695.2 expression-related risk score. Additionally, AP000695.2 was found to be positively associated with tumor-infiltrating immune cells, including classically activated (M1) macrophages, neutrophils, alternatively activated (M2) macrophages, and natural killer (NK) cells.

    Conclusions: It was observed that AP000695.2 can be used as a novel biomarker to diagnose or predict survival of gastric patient.

  • Article
    Shu Xu, Wenjun Zhang, Yan Zhang, Zhou Xu, Ting Wu
    Discovery Medicine. 2023, 35(175): 185-192. https://doi.org/10.24976/Discov.Med.202335175.19

    Background: The therapeutic outcomes for acute ischemic stroke (AIS) with early neurological deterioration (END) are adverse. Argatroban is a novel direct thrombin inhibitor, which is safe in the treatment of AIS, but its efficacy is controversial. This study sought to assess the therapeutic effect of argatroban as an adjunct to aspirin in the treatment of AIS patients with END. Patients’ prognosis for the presence of END was also evaluated.

    Methods: Overall, 166 AIS patients with END were included in the study from June 2018 to June 2021 in The Affiliated Zhangjiagang Hospital of Soochow University. Patients were divided in the control group (aspirin alone) and the study group (aspirin combined with argatroban). General data of the patients were collected. Clinical indexes such as the modified Edinburgh-Scandinavian stroke scale (MESSS), and the serum fibrinogen (FIB) and neuropeptide Y (NPY) levels before and after treatment were also collected. Correlations between prognosis and general data, and FIB and NPY levels in AIS patients with END were analyzed by multivariate logistic regression. The performance of FIB and NPY levels in predicting patients’ prognosis was further analyzed using receiver operating characteristic (ROC) curves.

    Results: There was no significant difference in the general data, such as sex, age, course of disease and basic diseases between the 2 groups. After treatment, the MESSS score (13.08 ± 3.24 vs. 16.48 ± 3.32, p < 0.001), serum FIB level (2.72 ± 0.81 vs. 3.52 ± 0.71, p < 0.001), and NPY level (121.28 ± 17.34 vs. 152.09 ± 18.25, p < 0.001) of the study group was significantly lower than that of the control group. A further analysis revealed that the serum FIB (OR, odds ratio = 2.296, 95% confidence interval, CI: 1.437–3.669, p = 0.001) and NPY (OR = 1.020, 95% CI: 1.002–1.039, p = 0.031) levels were independent risk factors of patients’ prognosis for the presence of END.

    Conclusions: Aspirin combined with argatroban significantly improved neurological impairment of AIS patients with END, which is worthy of clinical application.

  • Article
    Dandan Lang, Zhengmin Liu, Dejun Li
    Discovery Medicine. 2023, 35(176): 233-241. https://doi.org/10.24976/Discov.Med.202335176.24

    Background: Omalizumab is a recombinant humanized monoclonal antibody against immunoglobulin E., which can specifically bind to IgE in blood and inhibit the release of inflammatory mediators to improve the symptoms of IgE-mediated asthma effectively. This meta-analysis was used to retrieve the studies in recent years to provide a clinical reference for the omalizumab in treating allergic asthma (AA).

    Methods: The databases Ovid, Embase, Pubmed, the Cochrane Library of clinical trials, CNKI (China National Knowledge Infrastructure) (China), and Wangfang Data (China) were searched for all studies on omalizumab involvement in treating allergic childhood asthma up to January 2022. Effectiveness, rate of exacerbation within 24 weeks (and 52 weeks), and the incidence of adverse reactions and serious adverse reactions were used as the primary data analysis indicators.

    Results: Seven eligible pieces of literature were included. Meta-analysis indicated that omalizumab could significantly improve the treatment efficacy in children with asthma [RR (Risk Ratio) = 1.24, 95% CI (Confidential Interval) (1.09, 1.41), Z = 3.30, p = 0.001], reduced the incidence of significant clinical exacerbation in children with asthma within 24 weeks [RR = 0.55, 95% CI (0.35, 0.85), Z = –2.67, p = 0.001], reduced the incidence of significant clinical exacerbation in children with asthma within 52 weeks [RR = 0.52, 95% CI (0.39, 0.71), Z = –4.2, p < 0.0001], and the incidence of total serious adverse reactions was not statistically different from placebo [RR = 1.00, 95% CI (0.98, 1.03), Z = 0.71, p = 0.479], the incidence of serious adverse reactions was significantly decreased [RR = 0.53, 95% CI (0.36, 0.77), Z = –3.35, p = 0.001].

    Conclusions: In treating IgE (immunoglobulin E)-mediated asthma in children, adding oral (or subcutaneous) omalizumab to a glucocorticoid regimen can enhance the effectiveness of treatment, reduce the probability of significant exacerbation during treatment, and reduce the incidence of serious adverse reactions.

  • Article
    Wei Zhang, Jianjian Zhang
    Discovery Medicine. 2023, 35(178): 853-860. https://doi.org/10.24976/Discov.Med.202335178.80

    Background: Sepsis-induced myocardial dysfunction (SIMD) confers substantial morbidity and mortality. Semaglutide treatment has demonstrated efficacy in ameliorating sepsis-related organ damage via attenuation of inflammation, oxidative stress, and apoptotic cell death. In this study, we constructed a mouse SIMD model using cecal ligation and puncture (CLP) to explore whether semaglutide preconditioning can modulate autophagy levels and attenuate myocardial injury.

    Methods: C57BL/6 mice were randomly divided into six groups: sham, CLP (including CLP-6 h, CLP-12 h and CLP-24 h subgroups), semaglutide, and semaglutide+Compound-C, with five mice in each group. The latter two groups were given daily intraperitoneal injections of semaglutide for 14 days. The semaglutide+Compound-C group was given the autophagy inhibitor Compound-C intraperitoneally 1-hour before CLP surgery. After the last injection of semaglutide, SIMD mouse models were constructed by CLP surgery, while the sham group underwent a sham operation. All mice were sacrificed after surgery, and blood and myocardial specimens were collected. Enzyme-linked immunosorbent assay (ELISA) was used to measure the levels of inflammatory factors and myocardial injury markers in the serum, while quantitative real-time polymerase chain reaction (qRT-PCR) and western blot was used to detect the expression of autophagic markers [microtubule-associated protein 1A/1B-light chain 3B (LC3B), Beclin-1, p62] and AMP-activated protein kinase (AMPK) in myocardial tissue. Hematoxylin and eosin (H&E) staining was used to observe pathological changes in myocardial tissue.

    Results: The myocardial fibers in the sham group were normal, while those in the CLP group showed disordered arrangement, interstitial edema, and a large number of infiltrating inflammatory cells. A few vacuolar changes were observed locally in the semaglutide group, and more vacuolar changes were observed in the semaglutide+Compound-C group. Autophagy was inhibited in the CLP group mice. Compared with the CLP group, the semaglutide group showed a decreased levels of inflammatory factors (tumor necrosis factor-α, interleukin-1β) and myocardial injury markers (creatine kinase isoenzyme, cardiac troponin T) in the serum, a reduced expression of autophgic substrate p62, and an increased expression of LC3II (the lipidated form of LC3I)/LC3I (microtubule-associated protein 1A/1B-light chain 3), Beclin-1, and p-AMPK (phosphorylated AMP-activated protein kinase)/AMPK in the injured myocardial tissues of mice (p < 0.05). And the protective effects of semaglutide against SIMD were partially reversed by the treatment of AMPK inhibitor Compound-C (p < 0.05).

    Conclusions: Taken together, these data indicate that semaglutide provides protection against CLP-triggered myocardial inflammation and injury, potentially by reactivating myocardial autophagy pathways via activation of AMPK signaling. Further mechanistic studies are needed to definitively elucidate the functional significance of AMPK signaling in mediating the beneficial cardiac effects of semaglutide during sepsis.

  • Article
    Qiaoying Chai, Wei Zhang, Lijuan Gao, Yingtao Yang, Mengdan Miao, Da Liu, Lixia Chen, Mingqi Zheng, Shuanli Xin
    Discovery Medicine. 2023, 35(176): 394-404. https://doi.org/10.24976/Discov.Med.202335176.40

    Objective: To probe the effect of trehalose on myocardial hypertrophy and its specific molecular mechanism.

    Methods: C57BL/6J male mice were divided into four subgroups: Sham operation subgroup (Sham), negative sham subgroup (Sham+Trehalose), transverse aortic constriction (TAC), and trehalose treatment subgroup (TAC+Trehalose). Immediately after the TAC operation, trehalose at a dose of 10 mg/kg was given daily via gavage. After four weeks, changes in cardiac function were evaluated using ultrasound to measure EF (ejection fraction), FS (fractional shortening), IVRT (isovolumic relaxation time), MPI (myocardial performance index), Tau (isovolumic relaxation time constant), LVESP (left ventricular end-systolic pressure), and EDPVR (end-diastolic pressure-volume relationship). The profiles of autophagy-associated proteins (p62, LC3II/I, and Beclin-1) and GATA4 protein in mice myocardial tissues were assessed by Western blotting (WB). Myocardial cells were classified from TAC mice into five groups: Control, Trehalose, Phenylephrine (PE), PE+Trehalose, and PE+Trehalose+autophagy inhibitor chloroquine groups. In the PE group, cardiomyocytes were treated with 50 μmol/L PE. Then, the cells were treated with trehalose (100 μmol/L), trehalose (100 μmol/L)+autophagy (20 μmol/L) for 24 hours respectively. The Control group was treated with the same amount of normal saline. Flow cytometry was utilized to detect myocardial cell apoptosis in each subgroup. The alterations in apoptosis and autophagy-correlated proteins (p62, LC3II/I, and Beclin-1) were assessed by WB. Additionally, the level of GATA4 protein upstream of autophagy was estimated. Furthermore, the expression levels of pro-apoptotic proteins Bad, BAX, Cleaved-caspase-3, and anti-apoptotic protein Bcl-2 were examined by WB.

    Results: The TAC operation significantly augmented myocardial hypertrophy, heart weight-to-body weight ratio, and myocardial cell apoptosis in mice (p < 0.05). Trehalose significantly improved cardiac hypertrophy, cardiomyocyte apoptosis, and cardiac function decline in mice. Additionally, it also significantly enhanced autophagy in mouse cardiac tissues (p < 0.05). At the cellular level, trehalose significantly decreased PE-elicited apoptosis and promoted the protein expressions of Beclin-1 and LC3 II/I in cardiomyocytes while significantly dampening the profiles of p62 and GATA4 in cells. The effect of trehalose and chloroquine treatment was significantly greater than that of the trehalose group.

    Conclusions: Trehalose significantly abates myocardial hypertrophy and pressure overload-induced cardiomyocyte apoptosis in mice. The cardioprotective effect of trehalose on enhanced autophagy is attributed, at least in part, to the promotion of autophagic degradation of GATA4.

  • Article
    Jiaxuan Liu, Yaqin Xu, Yong Q. Chen, Dan Su
    Discovery Medicine. 2023, 35(176): 343-352. https://doi.org/10.24976/Discov.Med.202335176.35

    Background: Immune dysregulation contributes to the development of ulcerative colitis (UC). The research on the inflammatory response of UC is mainly focused on T cells, with less understanding of the role of B cells. Pax transactivation domain-interacting protein (PTIP) is essential for the development of B cell subpopulations and humoral immunity. The purpose of this study was to elucidate the role of PTIP in B cells of mice with dextran sodium sulfate (DSS)-induced colitis.

    Methods: The B-cell-specific PTIP knockout (PTIP-/-) mice were established by crossbreeding cluster of differentiation (CD)19cre/cre mice with PTIPflox/flox mice. The UC mice were induced by drinking water supplemented with 3.8% Dextran Sulfate Sodium (DSS) (PTIP-/- + DSS). The histological analysis was performed using hematoxylin and eosin staining. The immune cells were isolated using a fluorescence-activated cell sorter. The serum antibodies (immunoglobulin M (IgM) or immunoglobulin G (IgG)) and tumor necrosis factor (TNF)-α were determined by Enzyme linked immunosorbent assay (ELISA).

    Results: Interestingly, our findings demonstrate that PTIP deficiency in B cells significantly ameliorates UC. In contrast to PTIP-/- + DSS, the wild type (WT) + DSS group showed a more robust increase in disease activity index (DAI) scores (p < 0.05), a substantially shortened colon (p < 0.001) and a decrease of mucous-producing goblet cells and the complete destruction of crypts. Moreover, PTIP-deficient mice manifested markedly altered neutrophil and T-cell distribution in UC (p < 0.05). Although anti-commensal IgG exacerbates UC, we demonstrated, for the first time, that serum natural IgG does not aggravate the pathology of UC. Furthermore, PTIP regulates UC by controlling B-2 cells independently from T cells.

    Conclusions: Transplantation of splenic B-2 cells from PTIP-deficient mice protected recipient NOD/ShiltJGpt-Prkdcem26Cd52Il2rgem26Cd22/Gpt (NCG) mice from severe UC.