Pub Date : 2025-11-12DOI: 10.1016/j.bbagen.2025.130886
Yulia V. Bertsova, Marina V. Serebryakova, Alexander A. Baykov, Alexander V. Bogachev
Bacteria coping with oxygen deficiency can switch to alternative terminal electron acceptors, which can be normal metabolic intermediates or products of dedicated coupled reactions. In the latter case, the genes for the respective terminal reductase and coupling enzyme are expected to cluster in the genome. Here, we determined the roles of two uncharacterized periplasmic proteins encoded by the swoo_3912–swoo_3913 gene cluster in the facultatively anaerobic marine bacterium Shewanella woodyi. We confirmed the current database annotation of the former protein as “urocanate reductase” but identified the latter protein as a histidine betaine trimethylammonia-lyase (HBTL). HBTL converts histidine betaine into urocanate and trimethylamine and is remarkably specific for histidine betaine as substrate. HBTL requires Mg2+ for activity and undergoes slow reversible inactivation at low Mg2+ concentrations. HBTL activity was not evident in S. woodyi cells grown aerobically but was induced in cells grown anaerobically. Both histidine betaine and urocanate supported anaerobic S. woodyi growth and, hence, respiration. Similar gene clusters are found in many anaerobic bacteria, suggesting a wide occurrence of the anaerobic respiration pathway discovered in this work in the bacterial world.
{"title":"Histidine betaine trimethylammonia-lyase, enzyme coupled with terminal urocanate reductase in Shewanella woodyi grown anaerobically","authors":"Yulia V. Bertsova, Marina V. Serebryakova, Alexander A. Baykov, Alexander V. Bogachev","doi":"10.1016/j.bbagen.2025.130886","DOIUrl":"10.1016/j.bbagen.2025.130886","url":null,"abstract":"<div><div>Bacteria coping with oxygen deficiency can switch to alternative terminal electron acceptors, which can be normal metabolic intermediates or products of dedicated coupled reactions. In the latter case, the genes for the respective terminal reductase and coupling enzyme are expected to cluster in the genome. Here, we determined the roles of two uncharacterized periplasmic proteins encoded by the <em>swoo_3912</em>–<em>swoo_3913</em> gene cluster in the facultatively anaerobic marine bacterium <em>Shewanella woodyi</em>. We confirmed the current database annotation of the former protein as “urocanate reductase” but identified the latter protein as a histidine betaine trimethylammonia-lyase (HBTL). HBTL converts histidine betaine into urocanate and trimethylamine and is remarkably specific for histidine betaine as substrate. HBTL requires Mg<sup>2+</sup> for activity and undergoes slow reversible inactivation at low Mg<sup>2+</sup> concentrations. HBTL activity was not evident in <em>S. woodyi</em> cells grown aerobically but was induced in cells grown anaerobically. Both histidine betaine and urocanate supported anaerobic <em>S. woodyi</em> growth and, hence, respiration. Similar gene clusters are found in many anaerobic bacteria, suggesting a wide occurrence of the anaerobic respiration pathway discovered in this work in the bacterial world.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130886"},"PeriodicalIF":2.2,"publicationDate":"2025-11-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145522421","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-11DOI: 10.1016/j.bbagen.2025.130885
Neha Attal , Trenton A. Pritt , Melissa Stair , Tony E. Reeves , Iain H. McKillop
Background
Early alcohol-dependent liver disease (ALD) is characterized by increased hepatic fat storage (hepatosteatosis). Fatty acid binding protein 4 (FABP4), a protein not normally expressed in liver, becomes highly expressed in ALD. This study developed a hepatocyte-specific FABP4 mouse knockout (HS-Fabp4−/−) to study liver responses to alcohol.
Methods
An HS-Fabp4−/− mouse was created using a Cre/loxP embryonic stem cell approach. Male and female HS-Fabp4−/− and wildtype (WT; C57Bl/6) mice were maintained on ethanol-drinking water (EtOH-DW) for 4-weeks. Liver damage, triglyceride content and pathology were assessed. Hepatic FABP1–9 mRNA and FABP4 and FABP5 protein were measured. Human hepatoma cell proliferation in response to exogenous FABP4 or FABP5 was analyzed.
Results
Hepatocyte-specific FABP4 deletion was confirmed in HS-Fabp4−/− mice. No gross phenotypic differences were observed between HS-Fabp4−/− and WT. Maintenance on EtOH-DW resulted in microsteatosis, increased hepatic triglycerides, and elevated aspartate and alanine transaminases, with no differences detected between pair-matched HS-Fabp4−/− and WT mice. Hepatic FABP1–9 mRNA analysis revealed increased FABP4 and FABP5 mRNA expression in WT mice, and elevated FABP5 mRNA in HS-Fabp4−/− mice in response to EtOH-DW, effects that were mirrored in serum FABP4/5 protein. Exposure of hepatoma cells to FABP4 or FABP5 revealed FABP4, but not FABP5, stimulated cell proliferation.
Conclusions
Hepatocyte-specific FABP4 deletion does not alter hepatic fat accumulation in response to EtOH feeding. Hepatic FABP4 protein produced in response to EtOH is released from hepatocytes and exogenous FABP4 promotes hepatoma cell proliferation in vitro, an effect not observed for FABP5.
{"title":"Hepatic response to ethanol feeding in a hepatocyte-specific fatty acid binding protein-4 knock out mouse model","authors":"Neha Attal , Trenton A. Pritt , Melissa Stair , Tony E. Reeves , Iain H. McKillop","doi":"10.1016/j.bbagen.2025.130885","DOIUrl":"10.1016/j.bbagen.2025.130885","url":null,"abstract":"<div><h3>Background</h3><div>Early alcohol-dependent liver disease (ALD) is characterized by increased hepatic fat storage (hepatosteatosis). Fatty acid binding protein 4 (FABP4), a protein not normally expressed in liver, becomes highly expressed in ALD. This study developed a hepatocyte-specific FABP4 mouse knockout (HS-<em>Fabp4</em><sup>−/−</sup>) to study liver responses to alcohol.</div></div><div><h3>Methods</h3><div>An HS-<em>Fabp4</em><sup>−/−</sup> mouse was created using a Cre/loxP embryonic stem cell approach. Male and female HS-<em>Fabp4</em><sup>−/−</sup> and wildtype (WT; C57Bl/6) mice were maintained on ethanol-drinking water (EtOH-DW) for 4-weeks. Liver damage, triglyceride content and pathology were assessed. Hepatic FABP1–9 mRNA and FABP4 and FABP5 protein were measured. Human hepatoma cell proliferation in response to exogenous FABP4 or FABP5 was analyzed.</div></div><div><h3>Results</h3><div>Hepatocyte-specific FABP4 deletion was confirmed in HS-<em>Fabp4</em><sup>−/−</sup> mice. No gross phenotypic differences were observed between HS-<em>Fabp4</em><sup>−/−</sup> and WT. Maintenance on EtOH-DW resulted in microsteatosis, increased hepatic triglycerides, and elevated aspartate and alanine transaminases, with no differences detected between pair-matched HS-<em>Fabp4</em><sup>−/−</sup> and WT mice. Hepatic FABP1–9 mRNA analysis revealed increased FABP4 and FABP5 mRNA expression in WT mice, and elevated FABP5 mRNA in HS-<em>Fabp4</em><sup>−/−</sup> mice in response to EtOH-DW, effects that were mirrored in serum FABP4/5 protein. Exposure of hepatoma cells to FABP4 or FABP5 revealed FABP4, but not FABP5, stimulated cell proliferation.</div></div><div><h3>Conclusions</h3><div>Hepatocyte-specific FABP4 deletion does not alter hepatic fat accumulation in response to EtOH feeding. Hepatic FABP4 protein produced in response to EtOH is released from hepatocytes and exogenous FABP4 promotes hepatoma cell proliferation <em>in vitro</em>, an effect not observed for FABP5.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130885"},"PeriodicalIF":2.2,"publicationDate":"2025-11-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145511575","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-08DOI: 10.1016/j.bbagen.2025.130880
Patricia Andrea Garavaglia , Sebastián Aduviri , Pablo Trujillo , Laura Mónica Tasso , Joaquín Juan Bautista Cannata , Monica Pickholz , Gabriela Andrea García
Chagas disease, caused by the protozoan Trypanosoma cruzi, has become a global health concern due to increased globalization. Several studies suggest that the aldo-keto reductase from T. cruzi (TcAKR) is involved in resistance to benznidazole, the drug commonly used to treat this infection.
To further support the role of TcAKR in drug resistance, we evaluated its interaction with four compounds —quercetin, flufenamic acid, phenolphthalein, and menadione—previously reported as inhibitors of other AKRs. Molecular docking was performed to assess affinity and molecular specific interactions, and the inhibitory effects of these compounds on both TcAKR activities —aldo-keto reductase and quinone-oxidoreductase— were experimentally determined.
Binding affinities, in decreasing order, were: quercetin > flufenamic acid > phenolphthalein > menadione. Both quercetin and flufenamic acid interacted with amino acid residues located outside the enzyme's active site. Quercetin completely inhibited both TcAKR activities, while flufenamic acid inhibited approximately 50 %. Phenolphthalein and menadione showed low levels of inhibition. The inhibition profiles of quercetin and flufenamic acid were consistent with a noncompetitive mechanism.
The effect of quercetin on benznidazole resistance was evaluated in transfected parasites overexpressing TcAKR, which are 1.8-fold more resistant to this drug. Quercetin treatment restored benznidazole sensitivity in these parasites, reducing the IC₅₀ to levels comparable to those of control parasites. These results provide further evidence of TcAKR's involvement in benznidazole resistance and suggest that its inhibition can enhance treatment efficacy.
{"title":"Pharmacological inhibition of Trypanosoma cruzi aldo-keto reductase (TcAKR) and its effect on benznidazole resistance","authors":"Patricia Andrea Garavaglia , Sebastián Aduviri , Pablo Trujillo , Laura Mónica Tasso , Joaquín Juan Bautista Cannata , Monica Pickholz , Gabriela Andrea García","doi":"10.1016/j.bbagen.2025.130880","DOIUrl":"10.1016/j.bbagen.2025.130880","url":null,"abstract":"<div><div>Chagas disease, caused by the protozoan <em>Trypanosoma cruzi</em>, has become a global health concern due to increased globalization. Several studies suggest that the aldo-keto reductase from <em>T. cruzi</em> (<em>T</em>cAKR) is involved in resistance to benznidazole, the drug commonly used to treat this infection.</div><div>To further support the role of <em>Tc</em>AKR in drug resistance, we evaluated its interaction with four compounds —quercetin, flufenamic acid, phenolphthalein, and menadione—previously reported as inhibitors of other AKRs. Molecular docking was performed to assess affinity and molecular specific interactions, and the inhibitory effects of these compounds on both <em>Tc</em>AKR activities —aldo-keto reductase and quinone-oxidoreductase— were experimentally determined.</div><div>Binding affinities, in decreasing order, were: quercetin > flufenamic acid > phenolphthalein > menadione. Both quercetin and flufenamic acid interacted with amino acid residues located outside the enzyme's active site. Quercetin completely inhibited both <em>Tc</em>AKR activities, while flufenamic acid inhibited approximately 50 %. Phenolphthalein and menadione showed low levels of inhibition. The inhibition profiles of quercetin and flufenamic acid were consistent with a noncompetitive mechanism.</div><div>The effect of quercetin on benznidazole resistance was evaluated in transfected parasites overexpressing <em>Tc</em>AKR, which are 1.8-fold more resistant to this drug. Quercetin treatment restored benznidazole sensitivity in these parasites, reducing the IC₅₀ to levels comparable to those of control parasites. These results provide further evidence of <em>Tc</em>AKR's involvement in benznidazole resistance and suggest that its inhibition can enhance treatment efficacy.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130880"},"PeriodicalIF":2.2,"publicationDate":"2025-11-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145464715","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-08DOI: 10.1016/j.bbagen.2025.130884
Zohre Eftekhari, Fatemeh Kazemi-Lomedasht
Molecular imaging is a cornerstone of modern medicine, enabling non-invasive visualization of biological processes at the molecular level. The emergence of nanobodies (Nbs), small single-domain antibody fragments derived from camelids, has transformed this field due to their superior tissue penetration, rapid clearance, and high target specificity compared to conventional antibodies. This review focuses on the integration of Nbs with positron emission tomography (PET) and single-photon emission computed tomography (SPECT) two complementary molecular imaging modalities known for their high sensitivity, quantitative potential, and clinical relevance. Nb-based PET and SPECT imaging probes are emerging as powerful tools for detecting disease-associated molecular targets with exceptional precision. Their unique properties, including high affinity, specificity, and stability, make them ideal candidates for developing advanced radiotracers that enable early disease detection, monitoring of therapeutic responses, and evaluation of novel treatment strategies. Despite these advantages, several challenges remain, such as scalable Nb production, reduction of immunogenicity in clinical applications, and optimization of radiolabeling methods that preserve Nb integrity and function. This review highlights recent advances in Nb engineering, radiolabeling strategies, and preclinical and clinical applications of Nb-based PET and SPECT imaging, while outlining critical directions for future research. By addressing current limitations, Nb-based molecular imaging holds great promise for improving diagnostic accuracy, advancing personalized medicine, and expanding its clinical impact across diverse disease areas.
{"title":"Nanobody-based imaging: Advancing precision in molecular diagnostics","authors":"Zohre Eftekhari, Fatemeh Kazemi-Lomedasht","doi":"10.1016/j.bbagen.2025.130884","DOIUrl":"10.1016/j.bbagen.2025.130884","url":null,"abstract":"<div><div>Molecular imaging is a cornerstone of modern medicine, enabling non-invasive visualization of biological processes at the molecular level. The emergence of nanobodies (Nbs), small single-domain antibody fragments derived from camelids, has transformed this field due to their superior tissue penetration, rapid clearance, and high target specificity compared to conventional antibodies. This review focuses on the integration of Nbs with positron emission tomography (PET) and single-photon emission computed tomography (SPECT) two complementary molecular imaging modalities known for their high sensitivity, quantitative potential, and clinical relevance. Nb-based PET and SPECT imaging probes are emerging as powerful tools for detecting disease-associated molecular targets with exceptional precision. Their unique properties, including high affinity, specificity, and stability, make them ideal candidates for developing advanced radiotracers that enable early disease detection, monitoring of therapeutic responses, and evaluation of novel treatment strategies. Despite these advantages, several challenges remain, such as scalable Nb production, reduction of immunogenicity in clinical applications, and optimization of radiolabeling methods that preserve Nb integrity and function. This review highlights recent advances in Nb engineering, radiolabeling strategies, and preclinical and clinical applications of Nb-based PET and SPECT imaging, while outlining critical directions for future research. By addressing current limitations, Nb-based molecular imaging holds great promise for improving diagnostic accuracy, advancing personalized medicine, and expanding its clinical impact across diverse disease areas.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130884"},"PeriodicalIF":2.2,"publicationDate":"2025-11-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145487720","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-05DOI: 10.1016/j.bbagen.2025.130882
Ghizlane Loubane , Gabriel Robert , Syed Benazir Firdaus , Raphaël Robidas , Christian Comeau , Pierre-Luc Boudreault , Jeampy E. Komba , Hugo Gagnon , J Richard Wagner , Stephen Naylor , Klaus Klarskov
Homocysteine thiolactone (HTL) has been implicated in cardiovascular and neurological pathologies. While homocysteine can S-homocysteinylate thiol groups in proteins, the chemical properties of HTL facilitates covalent binding to protein ε-amino groups on lysine residues, which can initiate protein aggregation. Ascorbate plays an important role in the prevention of oxidative stress. Ascorbate is easily oxidized by the loss of two electrons to dehydroascorbate (DHA), which can be reduced back to ascorbate by thiol-containing smaller and larger molecules. In the present study, reaction products between two aldehydes, formaldehyde and propionaldehyde as well as DHA with the physiological relevant isomer of homocysteine thiolactone i.e. L-HTL were purified, and their structure was determined by 1D and 2D-nuclear magnetic resonance. In all three cases the reaction products are likely formed by initial imine condensation, subsequent formation of a hemiaminal product followed by HTL ring opening and intramolecular nucleophilic attack of the resulting thiol anion to form a six-member thiazinane ring with a carboxylic acid group. The structure of the DHA, L-HTL reaction product was confirmed by high resolution accurate ESIMS/MS in negative mode. Formation of the reaction product between DHA and HTL prevented N-homocysteinylation of cytochrome c by HTL, confirming earlier observations. The reaction product is formed in human neuroblastoma cells (SH-SY5Y) when exposed to HTL and DHA or ascorbate, potentially preventing protein aggregation. The consequences associated with formation of a reaction product between DHA and HTL suggest that DHA could protect against protein N-homocysteinylation.
{"title":"Structural characterization of a L-dehydroascorbic acid–L-homocysteine thiolactone reaction product: Intracellular formation in neuronal cells","authors":"Ghizlane Loubane , Gabriel Robert , Syed Benazir Firdaus , Raphaël Robidas , Christian Comeau , Pierre-Luc Boudreault , Jeampy E. Komba , Hugo Gagnon , J Richard Wagner , Stephen Naylor , Klaus Klarskov","doi":"10.1016/j.bbagen.2025.130882","DOIUrl":"10.1016/j.bbagen.2025.130882","url":null,"abstract":"<div><div>Homocysteine thiolactone (HTL) has been implicated in cardiovascular and neurological pathologies. While homocysteine can S-homocysteinylate thiol groups in proteins, the chemical properties of HTL facilitates covalent binding to protein ε-amino groups on lysine residues, which can initiate protein aggregation. Ascorbate plays an important role in the prevention of oxidative stress. Ascorbate is easily oxidized by the loss of two electrons to dehydroascorbate (DHA), which can be reduced back to ascorbate by thiol-containing smaller and larger molecules. In the present study, reaction products between two aldehydes, formaldehyde and propionaldehyde as well as DHA with the physiological relevant isomer of homocysteine thiolactone i.e. L-HTL were purified, and their structure was determined by 1D and 2D-nuclear magnetic resonance. In all three cases the reaction products are likely formed by initial imine condensation, subsequent formation of a hemiaminal product followed by HTL ring opening and intramolecular nucleophilic attack of the resulting thiol anion to form a six-member thiazinane ring with a carboxylic acid group. The structure of the DHA, L-HTL reaction product was confirmed by high resolution accurate ESIMS/MS in negative mode. Formation of the reaction product between DHA and HTL prevented N-homocysteinylation of cytochrome <em>c</em> by HTL, confirming earlier observations. The reaction product is formed in human neuroblastoma cells (SH-SY5Y) when exposed to HTL and DHA or ascorbate, potentially preventing protein aggregation. The consequences associated with formation of a reaction product between DHA and HTL suggest that DHA could protect against protein N-homocysteinylation.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130882"},"PeriodicalIF":2.2,"publicationDate":"2025-11-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145464716","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-05DOI: 10.1016/j.bbagen.2025.130883
Sara Trzos , Sylwia Orzechowska , Paweł Link-Lenczowski , Grzegorz Sokołowski , Malgorzata Baranska , Ewa Pocheć
Hashimoto's thyroiditis (HT) and Graves' disease (GD), autoimmune thyroid diseases (AITDs), are among the most commonly reported autoimmune disorders. Early and unambiguous diagnosis and monitoring of the disease development are crucial to obtain the most effective treatment results. Matrix-assisted laser desorption/ionization with a time-of-flight analyzer mass spectrometry (MALDI-ToF MS) and Raman spectroscopy are promising analytical methods in this regard. These non-invasive and sensitive techniques provide information on specific features of proteins, including their post-translational modifications and the level of a wide range of biomolecules in the bloodstream, which are useful for assessing the health status of patients. This study screened for potential biomarkers in AITDs using both methods. Sera from HT patients at two stages of disease progression and GD patients before and after normalization of thyrotropic hormone following immunosuppressive treatment were used in the study. Serum biomolecule changes were analyzed using Raman spectroscopy. N-glycans released from serum glycoproteins were detected by MALDI-ToF mass spectrometry. We observed that the levels of phenylalanine, carotenoids, and phospholipids in HT sera correlate with increased inflammation accompanying this disease. The GD group showed a lower amount of serum collagen compared to the HT patients. Quantitative comparison of N-glycans revealed several differences between the study groups and healthy donors. The higher galactosylation and α2,6-sialylation of serum proteins in HT2 relative to GD patients are the main differences in N-glycan profiles of AITDs. The obtained preliminary results demonstrate that these analytical techniques have potential in diagnosing and monitoring AITDs, and can be a good alternative to the currently used methods. Analysis of a larger number of samples, as well as a more detailed MS methodology for precise decoding of glycan structures, is necessary for further research.
桥本甲状腺炎（HT）和格雷夫斯病（GD），自身免疫性甲状腺疾病（AITDs），是最常报道的自身免疫性疾病。早期和明确的诊断和监测疾病发展是至关重要的，以获得最有效的治疗结果。基质辅助激光解吸/电离与飞行时间分析仪质谱（MALDI-ToF MS）和拉曼光谱是这方面有前途的分析方法。这些非侵入性和敏感的技术提供了蛋白质的特定特征信息，包括它们的翻译后修饰和血液中各种生物分子的水平，这对评估患者的健康状况很有用。本研究使用这两种方法筛选AITDs的潜在生物标志物。本研究采用HT患者两期病程和GD患者经免疫抑制治疗后促甲状腺激素恢复正常前后的血清。用拉曼光谱分析血清生物分子变化。采用MALDI-ToF质谱法检测血清糖蛋白释放的n -聚糖。我们观察到HT血清中苯丙氨酸、类胡萝卜素和磷脂的水平与此病伴随的炎症增加相关。GD组血清胶原蛋白含量低于HT组。n -聚糖的定量比较揭示了研究组与健康供者之间的一些差异。与GD患者相比，HT2患者血清蛋白的半乳糖基化和α2,6-唾液基化水平较高是AITDs n -聚糖谱的主要差异。初步结果表明，这些分析技术在诊断和监测AITDs方面具有一定的潜力，可以作为现有方法的一个很好的替代方案。进一步的研究需要分析大量的样品，以及更详细的质谱方法来精确解码多糖结构。
{"title":"Screening of biomarkers in autoimmune thyroid diseases (AITDs): preliminary study based on Raman spectroscopy and MALDI-ToF mass spectrometry","authors":"Sara Trzos , Sylwia Orzechowska , Paweł Link-Lenczowski , Grzegorz Sokołowski , Malgorzata Baranska , Ewa Pocheć","doi":"10.1016/j.bbagen.2025.130883","DOIUrl":"10.1016/j.bbagen.2025.130883","url":null,"abstract":"<div><div>Hashimoto's thyroiditis (HT) and Graves' disease (GD), autoimmune thyroid diseases (AITDs), are among the most commonly reported autoimmune disorders. Early and unambiguous diagnosis and monitoring of the disease development are crucial to obtain the most effective treatment results. Matrix-assisted laser desorption/ionization with a time-of-flight analyzer mass spectrometry (MALDI-ToF MS) and Raman spectroscopy are promising analytical methods in this regard. These non-invasive and sensitive techniques provide information on specific features of proteins, including their post-translational modifications and the level of a wide range of biomolecules in the bloodstream, which are useful for assessing the health status of patients. This study screened for potential biomarkers in AITDs using both methods. Sera from HT patients at two stages of disease progression and GD patients before and after normalization of thyrotropic hormone following immunosuppressive treatment were used in the study. Serum biomolecule changes were analyzed using Raman spectroscopy. <em>N</em>-glycans released from serum glycoproteins were detected by MALDI-ToF mass spectrometry. We observed that the levels of phenylalanine, carotenoids, and phospholipids in HT sera correlate with increased inflammation accompanying this disease. The GD group showed a lower amount of serum collagen compared to the HT patients. Quantitative comparison of <em>N</em>-glycans revealed several differences between the study groups and healthy donors. The higher galactosylation and α2,6-sialylation of serum proteins in HT2 relative to GD patients are the main differences in <em>N</em>-glycan profiles of AITDs. The obtained preliminary results demonstrate that these analytical techniques have potential in diagnosing and monitoring AITDs, and can be a good alternative to the currently used methods. Analysis of a larger number of samples, as well as a more detailed MS methodology for precise decoding of glycan structures, is necessary for further research.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130883"},"PeriodicalIF":2.2,"publicationDate":"2025-11-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145470321","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-04DOI: 10.1016/j.bbagen.2025.130881
Yan-Yan Guo, Xia-Nan Chu, Pei-Hua Wang, Ya-Qian Li, Li Xing
DNA damage response (DDR) signaling not only maintains genomic integrity but also plays a role in the activation of immune cells, including macrophages. In response to the stimuli, macrophages can be polarized into a proinflammatory phenotype, M1. In the monocytoid THP-1 cell-derived macrophage model, sodium butyrate was found to inhibit the expression of M1 biomarkers TNF-α, IL-6, IL-1β, and CXCL10, and downregulate DDR-associated proteins, including the apical Ataxia-Telangiectasia mutated protein kinase (ATM). However, siRNA-mediated knockdown of ATM did not reduce the expression of M1 biomarkers, but still downregulated the expression of DDR-associated proteins such as RAD50, p53, CHK1, NBS1, and γH2AX. Moreover, ATM knockdown modulated the expression of innate immunity regulatory genes, including sialic acid binding immunoglobulin type lectins 14 (Siglec14), Siglec15, signaling lymphocyte activation molecule family 1 (Siamf1), Siamf7, and guanylate binding protein 2 in response to the infection of Mycobacterium tuberculosis H37Ra. The results suggest that ATM may serve as a regulator to couple the DDR and innate immune response of macrophages, but barely contributes to the sodium butyrate-mediated inhibition of certain M1 biomarkers.
{"title":"The role of ATM in sodium butyrate-mediated inhibition of macrophage polarization","authors":"Yan-Yan Guo, Xia-Nan Chu, Pei-Hua Wang, Ya-Qian Li, Li Xing","doi":"10.1016/j.bbagen.2025.130881","DOIUrl":"10.1016/j.bbagen.2025.130881","url":null,"abstract":"<div><div>DNA damage response (DDR) signaling not only maintains genomic integrity but also plays a role in the activation of immune cells, including macrophages. In response to the stimuli, macrophages can be polarized into a proinflammatory phenotype, M1. In the monocytoid THP-1 cell-derived macrophage model, sodium butyrate was found to inhibit the expression of M1 biomarkers TNF-α, IL-6, IL-1β, and CXCL10, and downregulate DDR-associated proteins, including the apical Ataxia-Telangiectasia mutated protein kinase (ATM). However, siRNA-mediated knockdown of ATM did not reduce the expression of M1 biomarkers, but still downregulated the expression of DDR-associated proteins such as RAD50, p53, CHK1, NBS1, and γH2AX. Moreover, ATM knockdown modulated the expression of innate immunity regulatory genes, including sialic acid binding immunoglobulin type lectins 14 (Siglec14), Siglec15, signaling lymphocyte activation molecule family 1 (Siamf1), Siamf7, and guanylate binding protein 2 in response to the infection of <em>Mycobacterium tuberculosis</em> H37Ra. The results suggest that ATM may serve as a regulator to couple the DDR and innate immune response of macrophages, but barely contributes to the sodium butyrate-mediated inhibition of certain M1 biomarkers.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130881"},"PeriodicalIF":2.2,"publicationDate":"2025-11-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145457084","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-02DOI: 10.1016/j.bbagen.2025.130876
Yanpeng Ma , Longlong Liu , Jiayue Duan , Xiaojian Wang
Background
Cholangiocarcinoma (CCA) is characterized by poor prognosis and a lack of effective biomarkers and therapeutic targets. Although copper dysregulation has been implicated in CCA, the role of the copper transporter SLC31A1 (CTR1) in its progression remains poorly understood.
Methods
The mRNA and protein levels of SLC31A1 in tumor and adjacent non-tumor tissues was evaluated by RT-PCR and enzyme-linked immunosorbent assay, respectively. Tissue copper concentrations were determined using a colorimetric complexation assay. Correlations between SLC31A1 expression, copper accumulation, and clinicopathological parameters were evaluated using Pearson's analysis. Cell proliferation and colony formation were assessed by the cell counting kit 8 and colony formation assays, respectively. Mitochondrial function was assessed by measuring intracellular ATP levels, reactive oxygen species (ROS) production, and the glutathione redox ratio.
Results
Elevated SLC31A1 expression was significantly associated with microvascular invasion, larger tumor size (≥5 cm), and advanced AJCC stage (III–IV). Both SLC31A1 mRNA and protein levels were significantly upregulated in CCA tissues compared with adjacent non-tumor tissues and correlated with copper accumulation. In vitro, SLC31A1 was highly expressed across multiple CCA cell lines. Silencing SLC31A1 in RBE cells impaired cell proliferation and colony formation. Mechanistically, depletion of SLC31A1 increased intracellular ROS, reduced the GSH/GSSG ratio, and decreased ATP production, indicating disruption of mitochondrial function and redox homeostasis.
Conclusions
SLC31A1 promotes cholangiocarcinoma progression by maintaining copper homeostasis, mitochondrial integrity, and redox balance. These findings suggest that SLC31A1 may serve as a potential prognostic biomarker and a candidate therapeutic target.
{"title":"Copper transporter 1 contributes to the progression of cholangiocarcinoma","authors":"Yanpeng Ma , Longlong Liu , Jiayue Duan , Xiaojian Wang","doi":"10.1016/j.bbagen.2025.130876","DOIUrl":"10.1016/j.bbagen.2025.130876","url":null,"abstract":"<div><h3>Background</h3><div>Cholangiocarcinoma (CCA) is characterized by poor prognosis and a lack of effective biomarkers and therapeutic targets. Although copper dysregulation has been implicated in CCA, the role of the copper transporter SLC31A1 (CTR1) in its progression remains poorly understood.</div></div><div><h3>Methods</h3><div>The mRNA and protein levels of SLC31A1 in tumor and adjacent non-tumor tissues was evaluated by RT-PCR and enzyme-linked immunosorbent assay, respectively. Tissue copper concentrations were determined using a colorimetric complexation assay. Correlations between SLC31A1 expression, copper accumulation, and clinicopathological parameters were evaluated using Pearson's analysis. Cell proliferation and colony formation were assessed by the cell counting kit 8 and colony formation assays, respectively. Mitochondrial function was assessed by measuring intracellular ATP levels, reactive oxygen species (ROS) production, and the glutathione redox ratio.</div></div><div><h3>Results</h3><div>Elevated SLC31A1 expression was significantly associated with microvascular invasion, larger tumor size (≥5 cm), and advanced AJCC stage (III–IV). Both SLC31A1 mRNA and protein levels were significantly upregulated in CCA tissues compared with adjacent non-tumor tissues and correlated with copper accumulation. In vitro, SLC31A1 was highly expressed across multiple CCA cell lines. Silencing SLC31A1 in RBE cells impaired cell proliferation and colony formation. Mechanistically, depletion of SLC31A1 increased intracellular ROS, reduced the GSH/GSSG ratio, and decreased ATP production, indicating disruption of mitochondrial function and redox homeostasis.</div></div><div><h3>Conclusions</h3><div>SLC31A1 promotes cholangiocarcinoma progression by maintaining copper homeostasis, mitochondrial integrity, and redox balance. These findings suggest that SLC31A1 may serve as a potential prognostic biomarker and a candidate therapeutic target.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130876"},"PeriodicalIF":2.2,"publicationDate":"2025-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145443536","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-02DOI: 10.1016/j.bbagen.2025.130877
Layza Sá Rocha , Ana Cristina Jacobowski , Eduarda Thiburcio , Rafael Araujo Pereira , Claudiane Vilharroel Almeida , Camila De Oliveira Gutierrez , Thaís de Andrade Farias Rodrigues , Rodrigo Juliano Oliveira , Gabriel B. Taveira , Priscila Aiko Hiane , Ana Paula de Araújo Boleti , Octávio Luiz Franco , Marlon Henrique Cardoso , Maria Ligia Rodrigues Macedo
The ineffectiveness of conventional therapies against melanoma necessitates the development of novel approaches characterized by high selectivity and low systemic toxicity. Using computational optimization, KW18, a synthetic α-helical peptide, was rationally engineered. It incorporates conserved bioactive motifs from patented peptides. The Joker algorithm was used to refine the peptide. This process enhanced its positive charge, hydrophobicity, and α-helical stability. In vitro, KW18 displayed potent cytotoxicity against melanoma cells (IC₅₀ = 11.41–21.75 μM) with 84–91 % inhibition at 128 μM, minimal toxicity to FN1 fibroblasts, hemolysis below 5 %, and stability exceeding 80 %. Functional assays revealed mitochondrial damage and caspase-dependent apoptosis (86.5 % late apoptosis at 24 h). Cell cycle analysis in B16F10-Nex2 cells revealed a moderate accumulation in G0/G1 phase (62.9 %) alongside a substantial S-phase population (25.8 %), with minimal Sub-G0 (5.2 %) and reduced G2/M (6.1 %). This profile suggests a cytostatic-like effect through partial cell cycle slowdown. Structural characterization via helical wheel projections and circular dichroism confirmed its α-helical conformation, consistent with membrane-targeting activity. In vivo, Galleria mellonella larvae exposed to KW18 exhibited 100 % survival, supporting a favorable safety profile. Collectively, KW18 combines apoptosis induction with cell cycle modulation, offering a dual mechanism against melanoma while sparing healthy cells. These findings designate KW18 as a stable, selective, and safe therapeutic option for drug-resistant melanoma and a beneficial element for combined treatments.
{"title":"Rationally designed peptide induces apoptosis and cell cycle modulation in resistant melanoma","authors":"Layza Sá Rocha , Ana Cristina Jacobowski , Eduarda Thiburcio , Rafael Araujo Pereira , Claudiane Vilharroel Almeida , Camila De Oliveira Gutierrez , Thaís de Andrade Farias Rodrigues , Rodrigo Juliano Oliveira , Gabriel B. Taveira , Priscila Aiko Hiane , Ana Paula de Araújo Boleti , Octávio Luiz Franco , Marlon Henrique Cardoso , Maria Ligia Rodrigues Macedo","doi":"10.1016/j.bbagen.2025.130877","DOIUrl":"10.1016/j.bbagen.2025.130877","url":null,"abstract":"<div><div>The ineffectiveness of conventional therapies against melanoma necessitates the development of novel approaches characterized by high selectivity and low systemic toxicity. Using computational optimization, KW18, a synthetic α-helical peptide, was rationally engineered. It incorporates conserved bioactive motifs from patented peptides. The Joker algorithm was used to refine the peptide. This process enhanced its positive charge, hydrophobicity, and α-helical stability. In vitro, KW18 displayed potent cytotoxicity against melanoma cells (IC₅₀ = 11.41–21.75 μM) with 84–91 % inhibition at 128 μM, minimal toxicity to FN1 fibroblasts, hemolysis below 5 %, and stability exceeding 80 %. Functional assays revealed mitochondrial damage and caspase-dependent apoptosis (86.5 % late apoptosis at 24 h). Cell cycle analysis in B16F10-Nex2 cells revealed a moderate accumulation in G0/G1 phase (62.9 %) alongside a substantial S-phase population (25.8 %), with minimal Sub-G0 (5.2 %) and reduced G2/M (6.1 %). This profile suggests a cytostatic-like effect through partial cell cycle slowdown. Structural characterization via helical wheel projections and circular dichroism confirmed its α-helical conformation, consistent with membrane-targeting activity. In vivo, <em>Galleria mellonella</em> larvae exposed to KW18 exhibited 100 % survival, supporting a favorable safety profile. Collectively, KW18 combines apoptosis induction with cell cycle modulation, offering a dual mechanism against melanoma while sparing healthy cells. These findings designate KW18 as a stable, selective, and safe therapeutic option for drug-resistant melanoma and a beneficial element for combined treatments.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130877"},"PeriodicalIF":2.2,"publicationDate":"2025-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145443628","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-11-01DOI: 10.1016/j.bbagen.2025.130878
Weichen Zhan , Xiaowei Ding , Zhongrui Cui , Yizhuo Wu , Yiwen Gu , Hanxiao Cheng , Xinxin Ge , Yun Wang , Jiangyun Luo , Bing Xiao
Despite significant advancements in liposome-mediated transfection technology over the past decades, achieving optimal transfection efficiency with lipoplex remains challenging in certain primary cells, such as vascular smooth muscle cells, endothelial cells, and suspension cells. Here, we present an innovative approach to significantly enhance Lipofectamine-based transient transfection efficiency in hard-to-transfect cells by applying elevated cyclic hydrostatic pressure (CHP). The plasmids encoding the enhanced green fluorescent protein (EGFP) were transfected using Lipofectamine 3000 reagent, and the transfection efficiency was evaluated by Western blot or flow cytometry. Our results demonstrate that CHP (0.0083 Hz, 0–100 mmHg) significantly enhanced the transfection efficiency of lipoplex in primary human aortic smooth muscle cells (HASMCs) and other difficult-to-transfect cell types. Mechanistic studies revealed that the enhancement of liposome-mediated transfection by CHP was dependent on the activation of clathrin-dependent endocytic pathways. Importantly, this mechanical stimulation did not affect the proliferative or migratory capacities of HASMCs, despite the identification of significantly modulated proteins (5.8 % of the total proteome) by proteomic analysis. This study establishes a novel, safe strategy to enhance lipoplex-mediated nucleic acid delivery in challenging-to-transfect cell types.
{"title":"Elevated cyclic hydrostatic pressure enhances the transfection activity of lipoplexes by activating clathrin-mediated endocytosis","authors":"Weichen Zhan , Xiaowei Ding , Zhongrui Cui , Yizhuo Wu , Yiwen Gu , Hanxiao Cheng , Xinxin Ge , Yun Wang , Jiangyun Luo , Bing Xiao","doi":"10.1016/j.bbagen.2025.130878","DOIUrl":"10.1016/j.bbagen.2025.130878","url":null,"abstract":"<div><div>Despite significant advancements in liposome-mediated transfection technology over the past decades, achieving optimal transfection efficiency with lipoplex remains challenging in certain primary cells, such as vascular smooth muscle cells, endothelial cells, and suspension cells. Here, we present an innovative approach to significantly enhance Lipofectamine-based transient transfection efficiency in hard-to-transfect cells by applying elevated cyclic hydrostatic pressure (CHP). The plasmids encoding the enhanced green fluorescent protein (EGFP) were transfected using Lipofectamine 3000 reagent, and the transfection efficiency was evaluated by Western blot or flow cytometry. Our results demonstrate that CHP (0.0083 Hz, 0–100 mmHg) significantly enhanced the transfection efficiency of lipoplex in primary human aortic smooth muscle cells (HASMCs) and other difficult-to-transfect cell types. Mechanistic studies revealed that the enhancement of liposome-mediated transfection by CHP was dependent on the activation of clathrin-dependent endocytic pathways. Importantly, this mechanical stimulation did not affect the proliferative or migratory capacities of HASMCs, despite the identification of significantly modulated proteins (5.8 % of the total proteome) by proteomic analysis. This study establishes a novel, safe strategy to enhance lipoplex-mediated nucleic acid delivery in challenging-to-transfect cell types.</div></div>","PeriodicalId":8800,"journal":{"name":"Biochimica et biophysica acta. General subjects","volume":"1870 1","pages":"Article 130878"},"PeriodicalIF":2.2,"publicationDate":"2025-11-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145437017","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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