Pub Date : 2025-02-25DOI: 10.1016/j.bbrep.2025.101961
Shaohua Ning , Enfu Liu , Fanju Meng , Fei Chen , James Hartnett , Zhihong Lin , Bailin Tu , David J. Hawksworth , Bryan C. Tieman , De Yu Mao , Ryan Piktel , Amit Kumar , You Pan , Philip M. Hemken
Golgi protein 73 (GP73) is a serum marker with potential applications in the assessment of chronic liver disease progression. We developed a research use only (RUO) GP73 immunoassay to detect serum GP73 concentration. A pair of internal antibodies that most effectively capture and detect GP73 were used to develop a prototype assay. An internal recombinant GP73 standard was used to prepare the assay calibrator and controls. The stability performance of the antibodies and GP73 calibrator were evaluated. We analyzed assay performance on the automated Alinity i system, including precision, sensitivity, linearity, endogenous interference, and HAMA/RF interference. The RUO Alinity i GP73 immunoassay showed good stability when the reagent kit was stored on board the instrument for more than 30 days without recalibration. The internal GP73 calibrator was stable at 2–8 °C for 28 days. Total %CV for 20 days was ≤3 %. The limit of quantitation (LoQ) at 20 % CV was 0.20 ng/mL or lower. Dilution analysis yielded a linear result within the range of 2.1 ng/mL – 1000.0 ng/mL. No interference was observed from common endogenous interferents at each test interference level. These findings support the reliability and robustness of the RUO Alinity i GP73 immunoassay for measuring serum GP73 concentration.
{"title":"Development and analytical performance of a new research use only (RUO) GP73 automated immunoassay","authors":"Shaohua Ning , Enfu Liu , Fanju Meng , Fei Chen , James Hartnett , Zhihong Lin , Bailin Tu , David J. Hawksworth , Bryan C. Tieman , De Yu Mao , Ryan Piktel , Amit Kumar , You Pan , Philip M. Hemken","doi":"10.1016/j.bbrep.2025.101961","DOIUrl":"10.1016/j.bbrep.2025.101961","url":null,"abstract":"<div><div>Golgi protein 73 (GP73) is a serum marker with potential applications in the assessment of chronic liver disease progression. We developed a research use only (RUO) GP73 immunoassay to detect serum GP73 concentration. A pair of internal antibodies that most effectively capture and detect GP73 were used to develop a prototype assay. An internal recombinant GP73 standard was used to prepare the assay calibrator and controls. The stability performance of the antibodies and GP73 calibrator were evaluated. We analyzed assay performance on the automated Alinity i system, including precision, sensitivity, linearity, endogenous interference, and HAMA/RF interference. The RUO Alinity i GP73 immunoassay showed good stability when the reagent kit was stored on board the instrument for more than 30 days without recalibration. The internal GP73 calibrator was stable at 2–8 °C for 28 days. Total %CV for 20 days was ≤3 %. The limit of quantitation (LoQ) at 20 % CV was 0.20 ng/mL or lower. Dilution analysis yielded a linear result within the range of 2.1 ng/mL – 1000.0 ng/mL. No interference was observed from common endogenous interferents at each test interference level. These findings support the reliability and robustness of the RUO Alinity i GP73 immunoassay for measuring serum GP73 concentration.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101961"},"PeriodicalIF":2.3,"publicationDate":"2025-02-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143480166","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-24DOI: 10.1016/j.bbrep.2025.101962
Zhiqing Huang , Taoli Ding , Zixuan Ni , Yujie Zheng , Na Liu , Tuan Li , Wenchang Lian , Beihong Zheng , Yan Sun
Safety and ethical issues are the primary concerns for assisted reproductive technology (ART). However, confusion and contamination of samples are common problems in embryo laboratories, preimplantation genetic test (PGT) laboratories, and third-party medical testing laboratories due to large sample numbers and complex procedures. Once these problems occur, they are often difficult to trace, posing risks and ethical challenges to hospital reproductive centers, third-party medical testing laboratories, and patient families. Therefore, it is necessary to establish an effective and feasible tracing system to ensure sample safety. In this study, we designed an exogenous encoding sequence (EES) based on DNA data storage technology, which provide a unique identification code for each in vitro cultured embryo, effectively avoiding potential risks and ethical problems caused by sample confusion and contamination. This exogenous encoding sequence is a DNA molecule that is non-toxic and structurally stable. We verified that a small amount of exogenous encoding sequence (6∗109 copies/uL) can be amplified together with embryo biopsy cells and detected by various sequencing methods without affecting copy number variants (CNVs). Furthermore, if there is contamination from other samples at a proportion of more than 5 %, it can also be identified through the encoding information of the exogenous encoding sequence. Our study proves that the exogenous encoding sequence designed based on DNA data storage technology is effective and reliable, and can be applied in hospital reproductive centers and third-party medical testing laboratories to improve the safety of in vitro cultured embryos and avoid potential ethical problems in the future.
{"title":"An exogenous encoding sequence based on DNA data storage technology and its application in assisted reproductive technology","authors":"Zhiqing Huang , Taoli Ding , Zixuan Ni , Yujie Zheng , Na Liu , Tuan Li , Wenchang Lian , Beihong Zheng , Yan Sun","doi":"10.1016/j.bbrep.2025.101962","DOIUrl":"10.1016/j.bbrep.2025.101962","url":null,"abstract":"<div><div>Safety and ethical issues are the primary concerns for assisted reproductive technology (ART). However, confusion and contamination of samples are common problems in embryo laboratories, preimplantation genetic test (PGT) laboratories, and third-party medical testing laboratories due to large sample numbers and complex procedures. Once these problems occur, they are often difficult to trace, posing risks and ethical challenges to hospital reproductive centers, third-party medical testing laboratories, and patient families. Therefore, it is necessary to establish an effective and feasible tracing system to ensure sample safety. In this study, we designed an exogenous encoding sequence (EES) based on DNA data storage technology, which provide a unique identification code for each in vitro cultured embryo, effectively avoiding potential risks and ethical problems caused by sample confusion and contamination. This exogenous encoding sequence is a DNA molecule that is non-toxic and structurally stable. We verified that a small amount of exogenous encoding sequence (6∗109 copies/uL) can be amplified together with embryo biopsy cells and detected by various sequencing methods without affecting copy number variants (CNVs). Furthermore, if there is contamination from other samples at a proportion of more than 5 %, it can also be identified through the encoding information of the exogenous encoding sequence. Our study proves that the exogenous encoding sequence designed based on DNA data storage technology is effective and reliable, and can be applied in hospital reproductive centers and third-party medical testing laboratories to improve the safety of in vitro cultured embryos and avoid potential ethical problems in the future.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101962"},"PeriodicalIF":2.3,"publicationDate":"2025-02-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143480165","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Erythropoietin-producing hepatocellular receptor B6 (EphB6) is a member of the largest Eph subfamily of receptor tyrosine kinases. EphB6 is widely expressed in various tissues and regulates cellular homeostasis by interacting with its membrane-bound ephrin ligands and other receptors. EphB6 is involved in cancer pathology despite lacking kinase activity. Developing sensitive monoclonal antibodies (mAbs) for EphB6 has been desired for treatment, diagnosis, and further analysis of EphB6. This study established a novel specific and sensitive anti-human EphB6 mAb clone Eb6Mab-3 (mouse IgG1, kappa) by the Cell-Based Immunization and Screening (CBIS) method. In flow cytometry, Eb6Mab-3 demonstrated reactivity with EphB6-overexpressed Chinese hamster ovary-K1 cells (CHO/EphB6) and endogenously EphB6-expressing DLD-1 colorectal cancer cells. Cross-reactivity of Eb6Mab-3 was not observed. Eb6Mab-3 demonstrated a moderate binding affinity (dissociation constant; KD) for CHO/EphB6 (KD: 2.6 ± 1.0 × 10−8 M) and a high binding affinity for DLD-1 (KD: 3.4 ± 1.3 × 10−9 M). Eb6Mab-3 can detect EphB6 protein in CHO/EphB6 lysate in Western blot. Eb6Mab-3, established by the CBIS method, could be valuable for analyzing the EphB6-associated cellular functions and has potential applications in diagnosis and treatment with specificity and high affinity for cancer cells.
{"title":"Development of a novel anti-erythropoietin-producing hepatocellular receptor B6 monoclonal antibody Eb6Mab-3 for flow cytometry","authors":"Tomohiro Tanaka , Yu Kaneko , Haruto Yamamoto , Guanjie Li, Shiori Fujisawa, Hiroyuki Satofuka, Keisuke Shinoda, Takuya Nakamura, Mika K. Kaneko, Hiroyuki Suzuki, Yukinari Kato","doi":"10.1016/j.bbrep.2025.101960","DOIUrl":"10.1016/j.bbrep.2025.101960","url":null,"abstract":"<div><div>Erythropoietin-producing hepatocellular receptor B6 (EphB6) is a member of the largest Eph subfamily of receptor tyrosine kinases. EphB6 is widely expressed in various tissues and regulates cellular homeostasis by interacting with its membrane-bound ephrin ligands and other receptors. EphB6 is involved in cancer pathology despite lacking kinase activity. Developing sensitive monoclonal antibodies (mAbs) for EphB6 has been desired for treatment, diagnosis, and further analysis of EphB6. This study established a novel specific and sensitive anti-human EphB6 mAb clone Eb<sub>6</sub>Mab-3 (mouse IgG<sub>1</sub>, kappa) by the Cell-Based Immunization and Screening (CBIS) method. In flow cytometry, Eb<sub>6</sub>Mab-3 demonstrated reactivity with EphB6-overexpressed Chinese hamster ovary-K1 cells (CHO/EphB6) and endogenously EphB6-expressing DLD-1 colorectal cancer cells. Cross-reactivity of Eb<sub>6</sub>Mab-3 was not observed. Eb<sub>6</sub>Mab-3 demonstrated a moderate binding affinity (dissociation constant; <em>K</em><sub>D</sub>) for CHO/EphB6 (<em>K</em><sub>D</sub>: 2.6 ± 1.0 × 10<sup>−8</sup> M) and a high binding affinity for DLD-1 (<em>K</em><sub>D</sub>: 3.4 ± 1.3 × 10<sup>−9</sup> M). Eb<sub>6</sub>Mab-3 can detect EphB6 protein in CHO/EphB6 lysate in Western blot. Eb<sub>6</sub>Mab-3, established by the CBIS method, could be valuable for analyzing the EphB6-associated cellular functions and has potential applications in diagnosis and treatment with specificity and high affinity for cancer cells.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101960"},"PeriodicalIF":2.3,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143454154","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-21DOI: 10.1016/j.bbrep.2025.101947
Yuanzhao Wu, Cong Chen, Zao Jin, Kesi Zheng
Aims
To explore the potential functions and impacts of anoikis-related genes (ARGs) in breast cancer chemotherapy and to construct a prognosis model for HER2-negative breast cancer (HNBC) based on drug resistance-related ARGs.
Background
Breast cancer remains a leading cause of cancer-related mortality, with HER2-negative subtypes exhibiting high rates of metastasis and recurrence. Standard treatments for HNBC include taxane- and anthracycline-based chemotherapies, which aim to mitigate recurrence and metastasis. Anoikis, a specialized form of programmed cell death, plays a pivotal role in maintaining tissue homeostasis by eliminating detached cells. Cancer cells often develop resistance to anoikis, enabling survival in adverse conditions and promoting tumor progression.
Objective
To investigate the intersection of breast cancer drug resistance-related genes and anoikis-related genes (ARGs) and to assess their potential as biomarkers for HNBC. The study also aims to analyze differences in immune microenvironment and drug sensitivity among different prognosis score groups.
Method
A bioinformatics approach was employed to identify the intersection of breast cancer drug resistance-related genes and ARGs. A prognosis model for HNBC was developed based on these identified drug resistance-related ARGs. The study further examined differences in the immune microenvironment and drug sensitivity among different prognosis score groups.
Result
A prognosis model for HNBC was successfully constructed based on drug resistance-related ARGs. The study identified significant differences in immune microenvironment and drug sensitivity across different prognosis score groups.
Conclusion
The findings suggest that ARGs could be key in tailoring more effective therapeutic approaches for patients with HER2-negative breast cancer.
{"title":"Exploration of the role of drug resistance-associated anoikis-related genes in HER2-Negative breast cancer through bioinformatics analysis","authors":"Yuanzhao Wu, Cong Chen, Zao Jin, Kesi Zheng","doi":"10.1016/j.bbrep.2025.101947","DOIUrl":"10.1016/j.bbrep.2025.101947","url":null,"abstract":"<div><h3>Aims</h3><div>To explore the potential functions and impacts of anoikis-related genes (ARGs) in breast cancer chemotherapy and to construct a prognosis model for HER2-negative breast cancer (HNBC) based on drug resistance-related ARGs.</div></div><div><h3>Background</h3><div>Breast cancer remains a leading cause of cancer-related mortality, with HER2-negative subtypes exhibiting high rates of metastasis and recurrence. Standard treatments for HNBC include taxane- and anthracycline-based chemotherapies, which aim to mitigate recurrence and metastasis. Anoikis, a specialized form of programmed cell death, plays a pivotal role in maintaining tissue homeostasis by eliminating detached cells. Cancer cells often develop resistance to anoikis, enabling survival in adverse conditions and promoting tumor progression.</div></div><div><h3>Objective</h3><div>To investigate the intersection of breast cancer drug resistance-related genes and anoikis-related genes (ARGs) and to assess their potential as biomarkers for HNBC. The study also aims to analyze differences in immune microenvironment and drug sensitivity among different prognosis score groups.</div></div><div><h3>Method</h3><div>A bioinformatics approach was employed to identify the intersection of breast cancer drug resistance-related genes and ARGs. A prognosis model for HNBC was developed based on these identified drug resistance-related ARGs. The study further examined differences in the immune microenvironment and drug sensitivity among different prognosis score groups.</div></div><div><h3>Result</h3><div>A prognosis model for HNBC was successfully constructed based on drug resistance-related ARGs. The study identified significant differences in immune microenvironment and drug sensitivity across different prognosis score groups.</div></div><div><h3>Conclusion</h3><div>The findings suggest that ARGs could be key in tailoring more effective therapeutic approaches for patients with HER2-negative breast cancer.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101947"},"PeriodicalIF":2.3,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143454155","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-21DOI: 10.1016/j.bbrep.2025.101955
Pooreum Lim, Sang Woo Woo, Jihye Han, Young Lim Lee, Jae Ho Shim, Hyeon Soo Kim
Sarcopenia is an age-related muscle atrophy characterized by decreased muscle mass and function. However, potential treatments to alleviate sarcopenia remain limited. In this study, we investigated the effects of α-ketoisocaproate (KIC) on C2C12 differentiation and reactive oxygen species (ROS)-induced atrophy in C2C12 myotubes. We demonstrated that KIC upregulates the expression of myogenic differentiation factors, including myoblast determination protein 1 (MyoD) and myogenin (MyoG), during C2C12 differentiation. Additionally, KIC enhanced the expression of myosin heavy chain (MHC) isoforms MHC1, MHC2b, and MHC2x in C2C12 myotubes. KIC suppressed the decreased MyoG expression and the increase in the muscle atrophy-related factor, muscle atrophy F-box (MAFbx), in ROS-induced C2C12 myotubes. In addition, it restored the reduced expression of MHC and the diameter of C2C12 myotubes. We showed that KIC alleviated muscle atrophy by inhibiting mitogen-activated protein kinase (MAPK) signaling pathways, such as p38 MAPK and extracellular signal-regulated kinase 1/2 (ERK1/2). These findings suggest that KIC may serve as a potential therapeutic agent for ameliorating sarcopenia by inhibiting MAPK signaling in ROS-induced skeletal muscle cells.
{"title":"Effects of alpha-ketoisocaproate in oxidative stress-induced C2C12 myotubes via inhibition of p38 MAPK and ERK1/2","authors":"Pooreum Lim, Sang Woo Woo, Jihye Han, Young Lim Lee, Jae Ho Shim, Hyeon Soo Kim","doi":"10.1016/j.bbrep.2025.101955","DOIUrl":"10.1016/j.bbrep.2025.101955","url":null,"abstract":"<div><div>Sarcopenia is an age-related muscle atrophy characterized by decreased muscle mass and function. However, potential treatments to alleviate sarcopenia remain limited. In this study, we investigated the effects of α-ketoisocaproate (KIC) on C2C12 differentiation and reactive oxygen species (ROS)-induced atrophy in C2C12 myotubes. We demonstrated that KIC upregulates the expression of myogenic differentiation factors, including myoblast determination protein 1 (MyoD) and myogenin (MyoG), during C2C12 differentiation. Additionally, KIC enhanced the expression of myosin heavy chain (MHC) isoforms MHC1, MHC2b, and MHC2x in C2C12 myotubes. KIC suppressed the decreased MyoG expression and the increase in the muscle atrophy-related factor, muscle atrophy F-box (MAFbx), in ROS-induced C2C12 myotubes. In addition, it restored the reduced expression of MHC and the diameter of C2C12 myotubes. We showed that KIC alleviated muscle atrophy by inhibiting mitogen-activated protein kinase (MAPK) signaling pathways, such as p38 MAPK and extracellular signal-regulated kinase 1/2 (ERK1/2). These findings suggest that KIC may serve as a potential therapeutic agent for ameliorating sarcopenia by inhibiting MAPK signaling in ROS-induced skeletal muscle cells.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101955"},"PeriodicalIF":2.3,"publicationDate":"2025-02-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143454156","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1016/j.bbrep.2025.101958
Bahman Panahi , Rasmieh Hamid
Fungal infections pose a considerable threat to the cultivation of barley (Hordeum vulgare) and often limit the crop yield. During infection, the transcriptome undergoes extensive reprogramming involving several regulatory pathways. To address this complexity, we performed a comprehensive meta-analysis and co-expression network analysis using rigorously curated RNA-seq datasets from three different fungal diseases. Pre-processing of the data, including batch effect correction, ensured high-quality integration of the datasets. Module-trait relationship (MTR) analysis identified functional modules associated with fungal disease response. Hub genes within these modules were prioritized by multi-model centrality analyses using Cytoscape, which considered the metrics Degree, Closeness, Betweenness and Maximum Clique Centrality together with the MCODE algorithm to detect densely connected subclusters. These hub genes were further validated by cross-validation and receiver operating characteristic (ROC) curve analysis and achieved AUC values greater than 0.7, confirming their robustness. A total of 6688 consistently expressed genes were identified, including 879 upregulated and 701 downregulated genes. Co-expression networks revealed 19 different gene modules, six of which were significantly associated with the response of barley to fungal infection. The blue module in particular was associated with immune responses such as activation of the MAPK cascade and pathogen recognition, while the green module correlated with defence mechanisms and secondary metabolism. The hub genes within these modules showed high predictive power for fungal resistance, as shown by the AUC values of the ROC curve of over 0.7, emphasizing their potential as biomarkers. This study uniquely integrates multiple RNA-seq datasets to identify novel regulatory networks and hub genes, including 345 transcription factors (TFs) from different families, with MYB and bHLH being particularly abundant. The results provide valuable insights into regulatory networks associated with fungal disease response in barley. These results can support genomic selection and marker-assisted breeding programs and accelerate the development of resistant varieties.
{"title":"Unveiling functional module associated with fungal disease stress in barley (Hordeum vulgare)","authors":"Bahman Panahi , Rasmieh Hamid","doi":"10.1016/j.bbrep.2025.101958","DOIUrl":"10.1016/j.bbrep.2025.101958","url":null,"abstract":"<div><div>Fungal infections pose a considerable threat to the cultivation of barley (<em>Hordeum vulgare</em>) and often limit the crop yield. During infection, the transcriptome undergoes extensive reprogramming involving several regulatory pathways. To address this complexity, we performed a comprehensive meta-analysis and co-expression network analysis using rigorously curated RNA-seq datasets from three different fungal diseases. Pre-processing of the data, including batch effect correction, ensured high-quality integration of the datasets. Module-trait relationship (MTR) analysis identified functional modules associated with fungal disease response. Hub genes within these modules were prioritized by multi-model centrality analyses using Cytoscape, which considered the metrics Degree, Closeness, Betweenness and Maximum Clique Centrality together with the MCODE algorithm to detect densely connected subclusters. These hub genes were further validated by cross-validation and receiver operating characteristic (ROC) curve analysis and achieved AUC values greater than 0.7, confirming their robustness. A total of 6688 consistently expressed genes were identified, including 879 upregulated and 701 downregulated genes. Co-expression networks revealed 19 different gene modules, six of which were significantly associated with the response of barley to fungal infection. The blue module in particular was associated with immune responses such as activation of the MAPK cascade and pathogen recognition, while the green module correlated with defence mechanisms and secondary metabolism. The hub genes within these modules showed high predictive power for fungal resistance, as shown by the AUC values of the ROC curve of over 0.7, emphasizing their potential as biomarkers. This study uniquely integrates multiple RNA-seq datasets to identify novel regulatory networks and hub genes, including 345 transcription factors (TFs) from different families, with MYB and bHLH being particularly abundant. The results provide valuable insights into regulatory networks associated with fungal disease response in barley. These results can support genomic selection and marker-assisted breeding programs and accelerate the development of resistant varieties.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101958"},"PeriodicalIF":2.3,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143454289","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-20DOI: 10.1016/j.bbrep.2025.101956
Gabriel Lingotti, Mark R. Jones
Ratiometric and single-wavelength fluorophores are limited in their ability to provide basal level activity prior to treatment. This study presents a novel approach to characterise cell-wide basal activity of calcium using first and second harmonic spectral phasor analysis. Cells stained with the single-wavelength calcium fluorophore Oregon Green™ BAPTA 1-AM exhibited significant differences in wavelength or width in the nucleus, cytoplasm and membrane under basal conditions. Large and small cursor analysis was applied in the first and second harmonic, with smaller cursors revealing a region of interest enveloping and protruding from the nucleus in a structure akin to the sarcoplasmic reticulum. The first harmonic was found to be more sensitive in measurements of λmax, while the second harmonic showed increased sensitivity in measurements of spectral width. The results of this study indicate that first and second harmonic frequencies should be used in conjunction with phasor analysis of fluorophore microenvironments, rather than the first harmonic alone. Use of this approach may provide more insight into the cellular microenvironment under basal activity and treatment responses.
{"title":"Applying first & second harmonic spectral phasor analysis on a single-wavelength calcium fluorophore","authors":"Gabriel Lingotti, Mark R. Jones","doi":"10.1016/j.bbrep.2025.101956","DOIUrl":"10.1016/j.bbrep.2025.101956","url":null,"abstract":"<div><div>Ratiometric and single-wavelength fluorophores are limited in their ability to provide basal level activity prior to treatment. This study presents a novel approach to characterise cell-wide basal activity of calcium using first and second harmonic spectral phasor analysis. Cells stained with the single-wavelength calcium fluorophore Oregon Green™ BAPTA 1-AM exhibited significant differences in wavelength or width in the nucleus, cytoplasm and membrane under basal conditions. Large and small cursor analysis was applied in the first and second harmonic, with smaller cursors revealing a region of interest enveloping and protruding from the nucleus in a structure akin to the sarcoplasmic reticulum. The first harmonic was found to be more sensitive in measurements of <em>λ<sub>max</sub></em>, while the second harmonic showed increased sensitivity in measurements of spectral width. The results of this study indicate that first and second harmonic frequencies should be used in conjunction with phasor analysis of fluorophore microenvironments, rather than the first harmonic alone. Use of this approach may provide more insight into the cellular microenvironment under basal activity and treatment responses.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101956"},"PeriodicalIF":2.3,"publicationDate":"2025-02-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143454288","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-19DOI: 10.1016/j.bbrep.2025.101954
Nafiseh Shokri , Mohammad Elahimanesh , Masoomeh Bakhshandeh , Mohammad Najafi
Background
The atherosclerosis process relates to the dysfunction of different cells in the plaque microenvironment. The vascular smooth muscle cells (VSMCs) play an important role in atherosclerosis. Since the FoxO family is reported to control cell growth, thus the aim of this study was to investigate the effects of Heparin, Betulinic acid, and Ibrutinib on the FoxO1/pFoxO1 axis in vascular smooth muscle cells.
Methods
The VSMCs were cultured in DMEM-F12 medium and, were treated with Heparin (30IU), Betulinic acid (60 μM), and Ibrutinib (2 μM) in the 24 and 48-h periods. The FoxO1 gene expression level was identified using the RT-qPCR technique. The FoxO1 and pFoxO1 protein values were measured by the Western blot technique.
Results
The FoxO1 gene expression levels were reduced significantly in the cellular groups treated with Heparin, Betulinic acid, and Ibrutinib (P < 0.05) in both periods. Moreover, the pFoxO1 and FoxO1 protein values were decreased by Heparin, and Betulinic acid in the VSMCs.
Conclusion
Heparin suppressed the FoxO1/pFoxO1 signaling axis in vascular smooth muscle cells (VSMCs). Furthermore, it modulated the effects of Betulinic acid and Ibrutinib.
{"title":"Heparin suppresses FoxO1/pFoxO1 signaling axis in vascular smooth muscle cells","authors":"Nafiseh Shokri , Mohammad Elahimanesh , Masoomeh Bakhshandeh , Mohammad Najafi","doi":"10.1016/j.bbrep.2025.101954","DOIUrl":"10.1016/j.bbrep.2025.101954","url":null,"abstract":"<div><h3>Background</h3><div>The atherosclerosis process relates to the dysfunction of different cells in the plaque microenvironment. The vascular smooth muscle cells (VSMCs) play an important role in atherosclerosis. Since the FoxO family is reported to control cell growth, thus the aim of this study was to investigate the effects of Heparin, Betulinic acid, and Ibrutinib on the FoxO1/pFoxO1 axis in vascular smooth muscle cells.</div></div><div><h3>Methods</h3><div>The VSMCs were cultured in DMEM-F12 medium and, were treated with Heparin (30IU), Betulinic acid (60 μM), and Ibrutinib (2 μM) in the 24 and 48-h periods. The FoxO1 gene expression level was identified using the RT-qPCR technique. The FoxO1 and pFoxO1 protein values were measured by the Western blot technique.</div></div><div><h3>Results</h3><div>The FoxO1 gene expression levels were reduced significantly in the cellular groups treated with Heparin, Betulinic acid, and Ibrutinib (P < 0.05) in both periods. Moreover, the pFoxO1 and FoxO1 protein values were decreased by Heparin, and Betulinic acid in the VSMCs.</div></div><div><h3>Conclusion</h3><div>Heparin suppressed the FoxO1/pFoxO1 signaling axis in vascular smooth muscle cells (VSMCs). Furthermore, it modulated the effects of Betulinic acid and Ibrutinib.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101954"},"PeriodicalIF":2.3,"publicationDate":"2025-02-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143437811","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Inducing antigen peptide-specific cytotoxic T cells is challenging, partly due to the difficulty of maintaining the quality of antigen-presenting cells, such as dendritic cells. Consequently, artificial antigen-presenting cells (aAPCs) derived from the erythroleukemia cell line K562 have been employed for T cell stimulation. K562-based aAPCs can be utilized for both non-specific and antigen-specific T cell stimulation. Antigen peptide-pulsed aAPCs are commonly used to stimulate T cells with known specific antigenic peptides, which require identifying antigenic peptides from cognate antigen proteins. Therefore, antigen gene-overexpressing aAPCs might be useful for detecting unknown antigenic peptides. In this study, we evaluated the efficacy of cytotoxic T lymphocyte (CTL) induction using antigen gene-overexpressing aAPCs. To enhance antigen presentation efficiency, we assessed the signal peptide (SP) fused with MHC class I trafficking signal (MITD) sequences (SP-MITD). SP-MITD, fused with epitopes from the neoantigen AKF9 and the viral antigen CMVpp65, was transduced into aAPCs. We compared the CTL induction ability of peptide-pulsed aAPCs, only mini-gene-overexpressing aAPCs [SP-MITD (−)], and mini-gene fused with SP-MITD overexpressing aAPCs [SP-MITD (+)]. The SP-MITD (+) gene-overexpressing aAPCs exhibited the highest CTL induction efficiency compared to both peptide-pulsed and SP-MITD (−) gene-overexpressing aAPCs. These findings suggest that antigen gene-fused with SP-MITD transduced aAPCs are highly effective for inducing CTLs specific to both known and unknown antigenic peptides.
诱导抗原肽特异性细胞毒性 T 细胞具有挑战性，部分原因是难以保持树突状细胞等抗原递呈细胞的质量。因此，人们采用从红细胞白血病细胞系 K562 提取的人工抗原递呈细胞（aAPCs）来刺激 T 细胞。基于 K562 的人工抗原递呈细胞可用于非特异性和抗原特异性 T 细胞刺激。抗原肽脉冲 aAPC 通常用于用已知特异性抗原肽刺激 T 细胞，这需要从同源抗原蛋白中识别抗原肽。因此，抗原基因外表达的 aAPCs 可能有助于检测未知的抗原肽。在这项研究中，我们评估了使用抗原基因外表达 aAPCs 诱导细胞毒性 T 淋巴细胞（CTL）的效果。为了提高抗原呈递效率，我们评估了与 MHC I 类转运信号（MITD）序列融合的信号肽（SP）（SP-MITD）。SP-MITD融合了新抗原AKF9和病毒抗原CMVpp65的表位，被转导到aAPCs中。我们比较了多肽pulsed aAPCs、只表达迷你基因的aAPCs [SP-MITD（-）]和迷你基因与SP-MITD融合的过表达aAPCs [SP-MITD（+）]的CTL诱导能力。与肽脉冲和 SP-MITD (-) 基因表达的 aAPCs 相比，SP-MITD (+) 基因表达的 aAPCs 的 CTL 诱导效率最高。这些研究结果表明，抗原基因融合 SP-MITD 转导的 aAPCs 对诱导已知和未知抗原肽特异性 CTL 都非常有效。
{"title":"MHC class I trafficking signal improves induction of cytotoxic T lymphocyte using artificial antigen presenting cells","authors":"Kenta Sasaki , Kenji Murata , Tomoyuki Minowa , Naoki Shijubou , Hiroki Kobayashi , Munehide Nakatsugawa , Aiko Murai , Yuka Mizue , Terufumi Kubo , Takayuki Kanaseki , Tomohide Tsukahara , Hisashi Uhara , Akemi Ishida-Yamamoto , Yoshihiko Hirohashi , Toshihiko Torigoe","doi":"10.1016/j.bbrep.2025.101946","DOIUrl":"10.1016/j.bbrep.2025.101946","url":null,"abstract":"<div><div>Inducing antigen peptide-specific cytotoxic T cells is challenging, partly due to the difficulty of maintaining the quality of antigen-presenting cells, such as dendritic cells. Consequently, artificial antigen-presenting cells (aAPCs) derived from the erythroleukemia cell line K562 have been employed for T cell stimulation. K562-based aAPCs can be utilized for both non-specific and antigen-specific T cell stimulation. Antigen peptide-pulsed aAPCs are commonly used to stimulate T cells with known specific antigenic peptides, which require identifying antigenic peptides from cognate antigen proteins. Therefore, antigen gene-overexpressing aAPCs might be useful for detecting unknown antigenic peptides. In this study, we evaluated the efficacy of cytotoxic T lymphocyte (CTL) induction using antigen gene-overexpressing aAPCs. To enhance antigen presentation efficiency, we assessed the signal peptide (SP) fused with MHC class I trafficking signal (MITD) sequences (SP-MITD). SP-MITD, fused with epitopes from the neoantigen AKF9 and the viral antigen CMVpp65, was transduced into aAPCs. We compared the CTL induction ability of peptide-pulsed aAPCs, only mini-gene-overexpressing aAPCs [SP-MITD (−)], and mini-gene fused with SP-MITD overexpressing aAPCs [SP-MITD (+)]. The SP-MITD (+) gene-overexpressing aAPCs exhibited the highest CTL induction efficiency compared to both peptide-pulsed and SP-MITD (−) gene-overexpressing aAPCs. These findings suggest that antigen gene-fused with SP-MITD transduced aAPCs are highly effective for inducing CTLs specific to both known and unknown antigenic peptides.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101946"},"PeriodicalIF":2.3,"publicationDate":"2025-02-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143437810","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-18DOI: 10.1016/j.bbrep.2025.101943
Dengqin Ma , Bing Li , Bang Xin , Bingfang Xie , Enpen Zhu , Zihao Zhang , Xiaoqin Ha
Objective
To investigate the associations between metabolic changes and functions, including energy metabolism, immune response, and redox balance, under short-term hypobaric hypoxia exposure. Non-targeted metabolomics and bioinformatics analysis were applied to explore the adaptive mechanisms of organisms in hypobaric hypoxia.
Methods
Healthy adult male Sprague—Dawley rats were placed in environments simulating altitudes of 6500 m (HC group) and 1588 m (Control group). After 14 days, arterial serum samples were analyzed using Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS). Significant metabolites (P < 0.05, VIP >1) were identified, and KEGG enrichment analysis was conducted. Differential metabolites were globally analyzed with MetaboAnalyst 5.0.
Results
A total of 117 significantly altered metabolites were identified. In the HC group, 84 metabolites significantly increased, while 33 metabolites significantly decreased compared to the Control group. KEGG enrichment analysis revealed significant metabolic pathways, including the PPAR signaling pathway, bile secretion, arginine biosynthesis, alcoholism, and cholesterol metabolism (P < 0.05). Global analysis indicated that these differential metabolites were involved in various pathways, such as energy metabolism, amino acid metabolism, nucleotide metabolism, lipid metabolism, vitamin and cofactor metabolism, steroid metabolism, neurotransmitter metabolism, and heme metabolism, all of which play crucial roles in multiple biological processes.
Conclusion
Short-term hypobaric hypoxia exposure significantly altered the metabolite profiles in the arterial serum samples of rats, revealing adaptive metabolic reprogramming in energy metabolism, redox balance, immune function, endocrine regulation, and neurological systems.
{"title":"Metabolomic analysis of rat arterial serum under hypobaric hypoxia: Adaptive regulation of physiological systems by metabolic reprogramming","authors":"Dengqin Ma , Bing Li , Bang Xin , Bingfang Xie , Enpen Zhu , Zihao Zhang , Xiaoqin Ha","doi":"10.1016/j.bbrep.2025.101943","DOIUrl":"10.1016/j.bbrep.2025.101943","url":null,"abstract":"<div><h3>Objective</h3><div>To investigate the associations between metabolic changes and functions, including energy metabolism, immune response, and redox balance, under short-term hypobaric hypoxia exposure. Non-targeted metabolomics and bioinformatics analysis were applied to explore the adaptive mechanisms of organisms in hypobaric hypoxia.</div></div><div><h3>Methods</h3><div>Healthy adult male Sprague—Dawley rats were placed in environments simulating altitudes of 6500 m (HC group) and 1588 m (Control group). After 14 days, arterial serum samples were analyzed using Liquid Chromatography-Tandem Mass Spectrometry (LC-MS/MS). Significant metabolites (P < 0.05, VIP >1) were identified, and KEGG enrichment analysis was conducted. Differential metabolites were globally analyzed with MetaboAnalyst 5.0.</div></div><div><h3>Results</h3><div>A total of 117 significantly altered metabolites were identified. In the HC group, 84 metabolites significantly increased, while 33 metabolites significantly decreased compared to the Control group. KEGG enrichment analysis revealed significant metabolic pathways, including the PPAR signaling pathway, bile secretion, arginine biosynthesis, alcoholism, and cholesterol metabolism (P < 0.05). Global analysis indicated that these differential metabolites were involved in various pathways, such as energy metabolism, amino acid metabolism, nucleotide metabolism, lipid metabolism, vitamin and cofactor metabolism, steroid metabolism, neurotransmitter metabolism, and heme metabolism, all of which play crucial roles in multiple biological processes.</div></div><div><h3>Conclusion</h3><div>Short-term hypobaric hypoxia exposure significantly altered the metabolite profiles in the arterial serum samples of rats, revealing adaptive metabolic reprogramming in energy metabolism, redox balance, immune function, endocrine regulation, and neurological systems.</div></div>","PeriodicalId":8771,"journal":{"name":"Biochemistry and Biophysics Reports","volume":"41 ","pages":"Article 101943"},"PeriodicalIF":2.3,"publicationDate":"2025-02-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143428835","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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