Pub Date : 2025-07-09DOI: 10.1016/j.bbamem.2025.184434
Will Linthicum , Qi Wen , Nancy A. Burnham , Svetla D. Petrova , Konstantin Balashev
The investigation of how drugs or toxins alter cell mechanics is gaining significant traction in biomedical science, driven by the dual objectives of elucidating their mechanisms of action and enhancing drug screening processes. In this study, Atomic Force Microscopy (AFM), a prominent experimental technique in recent years, was employed to examine and analyze the mechanical responses of cells exposed to the neurotoxin Vipoxin. This method enables the precise measurement of key mechanical parameters such as cell stiffness and viscoelasticity before and after toxin introduction in the cell culture. It was demonstrated that the cells' stiffness and viscosity decreased with increasing Vipoxin concentration. Additionally, Total Internal Reflection Fluorescence Microscopy (TIRFM) was utilized to monitor morphological changes in the cells over time. These morphological changes were quantitatively analyzed using fractal analysis of the acquired images. The observed changes in cell shapes implied the reorganization of the cell cytoskeleton, thus providing insight into a comprehensive understanding of cell mechanics under the influence of Vipoxin.
{"title":"The phospholipolytically active neurotoxin Vipoxin induces changes of the mechanical properties of breast epithelial cells","authors":"Will Linthicum , Qi Wen , Nancy A. Burnham , Svetla D. Petrova , Konstantin Balashev","doi":"10.1016/j.bbamem.2025.184434","DOIUrl":"10.1016/j.bbamem.2025.184434","url":null,"abstract":"<div><div>The investigation of how drugs or toxins alter cell mechanics is gaining significant traction in biomedical science, driven by the dual objectives of elucidating their mechanisms of action and enhancing drug screening processes. In this study, Atomic Force Microscopy (AFM), a prominent experimental technique in recent years, was employed to examine and analyze the mechanical responses of cells exposed to the neurotoxin Vipoxin. This method enables the precise measurement of key mechanical parameters such as cell stiffness and viscoelasticity before and after toxin introduction in the cell culture. It was demonstrated that the cells' stiffness and viscosity decreased with increasing Vipoxin concentration. Additionally, Total Internal Reflection Fluorescence Microscopy (TIRFM) was utilized to monitor morphological changes in the cells over time. These morphological changes were quantitatively analyzed using fractal analysis of the acquired images. The observed changes in cell shapes implied the reorganization of the cell cytoskeleton, thus providing insight into a comprehensive understanding of cell mechanics under the influence of Vipoxin.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 7","pages":"Article 184434"},"PeriodicalIF":2.8,"publicationDate":"2025-07-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144616113","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}
(−)-Epigallocatechin gallate (EGCg), a tea catechin, exhibits antimicrobial activity. EGCg induces burst of giant unilamellar vesicles (GUVs), resulting in leakage of their internal contents. Here, we examined the effect of membrane tension on the EGCg-induced burst of dioleoylphosphatidylcholine (DOPC)-GUVs. The observation of the EGCg-induced burst of GUVs without membrane tension indicated that first a micropore was formed in the membrane and then its radius rapidly increased with time within ~10 ms without a change in the GUV diameter, and then the GUV diameter decreased to a lipid membrane aggregate. Next, we examined the effect of membrane tension on the EGCg-induced fractional area change (δ) of GUVs and its burst. During the interaction of EGCg with a GUV with low tension (≤ 0.5 mN/m), the δ initially increased slightly and then decreased to negative values, concomitant with the appearance of invaginated structures such as dense particles and narrow tubes in the GUV membrane and lumen, resulting in pore formation and subsequent GUV burst. By contrast, at higher tension, δ increased with time. The fraction of burst GUV after 5 min of interaction, Pburst (5 min), decreased with increasing tension up to 3.0 mN/m, indicating that membrane tension suppressed the burst. The Pburst (5 min) increased with increasing the fraction of GUVs in which invaginated structures appeared within 5 min of interaction, suggesting that the formation of invaginated structures may cause the initial EGCg-induced pore formation. We have proposed a mechanism of the tension effect on the EGCg-induced pore formation.
{"title":"Effect of membrane tension on (−)-epigallocatechin gallate-induced burst of single giant unilamellar vesicles","authors":"Yukihiro Tamba , Naoya Sugita , Mika Terada , Masahito Yamazaki","doi":"10.1016/j.bbamem.2025.184427","DOIUrl":"10.1016/j.bbamem.2025.184427","url":null,"abstract":"<div><div>(−)-Epigallocatechin gallate (EGCg), a tea catechin, exhibits antimicrobial activity. EGCg induces burst of giant unilamellar vesicles (GUVs), resulting in leakage of their internal contents. Here, we examined the effect of membrane tension on the EGCg-induced burst of dioleoylphosphatidylcholine (DOPC)-GUVs. The observation of the EGCg-induced burst of GUVs without membrane tension indicated that first a micropore was formed in the membrane and then its radius rapidly increased with time within ~10 ms without a change in the GUV diameter, and then the GUV diameter decreased to a lipid membrane aggregate. Next, we examined the effect of membrane tension on the EGCg-induced fractional area change (<em>δ</em>) of GUVs and its burst. During the interaction of EGCg with a GUV with low tension (≤ 0.5 mN/m), the <em>δ</em> initially increased slightly and then decreased to negative values, concomitant with the appearance of invaginated structures such as dense particles and narrow tubes in the GUV membrane and lumen, resulting in pore formation and subsequent GUV burst. By contrast, at higher tension, <em>δ</em> increased with time. The fraction of burst GUV after 5 min of interaction, <em>P</em><sub>burst</sub> (5 min), decreased with increasing tension up to 3.0 mN/m, indicating that membrane tension suppressed the burst. The <em>P</em><sub>burst</sub> (5 min) increased with increasing the fraction of GUVs in which invaginated structures appeared within 5 min of interaction, suggesting that the formation of invaginated structures may cause the initial EGCg-induced pore formation. We have proposed a mechanism of the tension effect on the EGCg-induced pore formation.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184427"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144172125","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 : 2025-06-01DOI: 10.1016/j.bbamem.2025.184428
Hallie N. Pennington, Jinwoo Lee
Lassa virus (LASV) is the most prevalent arenavirus afflicting humans and has high pandemic potential. The genetic material of LASV is delivered into the host cell via membrane fusion initiated by the LASV fusion domain (FD). However, the molecular details of the LASV FD, particularly its structure after association with the host cell, remain poorly characterized. This can be attributed to a lack of a viable membrane mimic to effectively stabilize the LASV FD for structural studies. Here, we demonstrate that the structure of the LASV FD widely varies based on the class of membrane mimic. In particular, through CD spectroscopy, we found that the LASV FD required a charged membrane mimic, such as zwitterionic or anionic detergent micelles, to adopt a helical conformation at low pH, but has the highest helical content in the presence of anionic lipids, particularly the detergent micelle LMPG and acidic bicelles. Moreover, we reveal that the LASV FD was well resolved on NMR spectra in CHAPS, DPC, LDAO, LMPG, and acidic bicelles, where LMPG and acidic bicelles had the sharpest peak resolution, but more defined peaks were noted in acidic bicelles over LMPG. In conclusion, our findings indicate that acidic bicelles are the optimal membrane mimic for the stabilization of the LASV FD such that structural studies can be conducted.
{"title":"Acidic bicelles are a suitable membrane mimic for structural studies of the Lassa virus fusion domain","authors":"Hallie N. Pennington, Jinwoo Lee","doi":"10.1016/j.bbamem.2025.184428","DOIUrl":"10.1016/j.bbamem.2025.184428","url":null,"abstract":"<div><div>Lassa virus (LASV) is the most prevalent arenavirus afflicting humans and has high pandemic potential. The genetic material of LASV is delivered into the host cell via membrane fusion initiated by the LASV fusion domain (FD). However, the molecular details of the LASV FD, particularly its structure after association with the host cell, remain poorly characterized. This can be attributed to a lack of a viable membrane mimic to effectively stabilize the LASV FD for structural studies. Here, we demonstrate that the structure of the LASV FD widely varies based on the class of membrane mimic. In particular, through CD spectroscopy, we found that the LASV FD required a charged membrane mimic, such as zwitterionic or anionic detergent micelles, to adopt a helical conformation at low pH, but has the highest helical content in the presence of anionic lipids, particularly the detergent micelle LMPG and acidic bicelles. Moreover, we reveal that the LASV FD was well resolved on NMR spectra in CHAPS, DPC, LDAO, LMPG, and acidic bicelles, where LMPG and acidic bicelles had the sharpest peak resolution, but more defined peaks were noted in acidic bicelles over LMPG. In conclusion, our findings indicate that acidic bicelles are the optimal membrane mimic for the stabilization of the LASV FD such that structural studies can be conducted.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184428"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144231702","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}
TRPV5 and TRPV6 are members of the TRP superfamily of ion channels and are present in almost all vertebrates as linked-genes with high homology and functional similarities. Abnormalities in the regulation or function of these two channels cause multiple pathological conditions, making these highly relevant for several diseases and pharmacological applications. In this context, how these two channels differ from each other is largely unknown. Here we analysed ∼250 protein sequences from vertebrates and critically analysed the conservation of signature motifs, such as different domains, TM-regions, loop-regions, cholesterol-binding regions and lipid-water-interface (LWI) regions. We demonstrate the subtle differences in the motifs and residues that are present at the LWI in TRPV5 and TRPV6. The ratio of hydrophobic-to-hydrophilic residues (but not of positive-to-negative residues) remains conserved for TRPV5 and TRPV6 throughout the vertebrate evolution. We also found motifs where cholesterol and/or sphingolipid can interact with TRPV5 and TRPV6 yet with different energies, both in open- and close-conformation. Accordingly, experimental evidence suggest that partial depletion of membrane cholesterol lowers the agonist-mediated opening of TRPV6, confirming the importance of membrane cholesterol for channel function. Further analysis of somatic mutations suggests that the specific inner LWI regions of TRPV5 (first 3) and TRPV6 (first 5) impose mutational hot-spots that are linked with different cancers. These findings may have broad significance in designing pharmacological agents for targeting TRPV5 and TRPV6 separately or simultaneously.
{"title":"Amino acids critical for lipid/s-interaction at the lipid-water-interface of TRPV5/TRPV6 remain different during vertebrate radiation: Relevance in cancer, bone disorders and other pathophysiologies","authors":"Sweta Agarwal , Vikash Kumar , Anupriya Chattapadhya , Shamit Kumar , Luna Goswami , Chandan Goswami","doi":"10.1016/j.bbamem.2025.184433","DOIUrl":"10.1016/j.bbamem.2025.184433","url":null,"abstract":"<div><div>TRPV5 and TRPV6 are members of the TRP superfamily of ion channels and are present in almost all vertebrates as linked-genes with high homology and functional similarities. Abnormalities in the regulation or function of these two channels cause multiple pathological conditions, making these highly relevant for several diseases and pharmacological applications. In this context, how these two channels differ from each other is largely unknown. Here we analysed ∼250 protein sequences from vertebrates and critically analysed the conservation of signature motifs, such as different domains, TM-regions, loop-regions, cholesterol-binding regions and lipid-water-interface (LWI) regions. We demonstrate the subtle differences in the motifs and residues that are present at the LWI in TRPV5 and TRPV6. The ratio of hydrophobic-to-hydrophilic residues (but not of positive-to-negative residues) remains conserved for TRPV5 and TRPV6 throughout the vertebrate evolution. We also found motifs where cholesterol and/or sphingolipid can interact with TRPV5 and TRPV6 yet with different energies, both in open- and close-conformation. Accordingly, experimental evidence suggest that partial depletion of membrane cholesterol lowers the agonist-mediated opening of TRPV6, confirming the importance of membrane cholesterol for channel function. Further analysis of somatic mutations suggests that the specific inner LWI regions of TRPV5 (first 3) and TRPV6 (first 5) impose mutational hot-spots that are linked with different cancers. These findings may have broad significance in designing pharmacological agents for targeting TRPV5 and TRPV6 separately or simultaneously.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184433"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144551838","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 : 2025-06-01DOI: 10.1016/j.bbamem.2025.184432
Lidia Ślusarczyk , Klaudia Rząd , Daniel Kamiński , Mariusz Gagoś , Małgorzata Figiel , Arkadiusz Matwijczuk
The paper presents the results of the first spectroscopic, microcalorimetric, and crystallographic study analyzing in great detail the strongly synergistic composition containing a selected 1,3,4-thiadiazole derivative: 4-(5-methyl-1,3,4-thiadiazol-2-ylo)benzene-1,3-diol (C1) and the antibiotic amphotericin B (AmB) in model biological DPPC films additionally modified with sterols: cholesterol (Chol) and ergosterol (Erg). The spectroscopic properties of the analyzed composition were studied with the use of spectroscopic methods, including: measurements of electronic fluorescence and absorption spectra with the technique of resonance light scattering (RLS), measurements of stationary fluorescence anisotropy and time-resolved fluorescence lifetimes with the method of single photon counting (TCSPC), circular dichroism spectra (CD), and infrared FTIR spectra. The mentioned methods were further complemented by including microcalorimetric DSC and crystallographic XRD analyses. All the measurements were taken in model biological systems formed as liposomal films and liposomal multilayers composed of DPPC and/or modified with sterols: cholesterol and ergosterol. The absorption spectra measured for C1/AmB and the synergistic composition thereof revealed differences indicating that while the antibiotic forms retained its known tendency to form aggregate in film systems, the synergistic composition had certain effects on the aggregational equilibrium. This was also reflected in the fluorescence emission measured for said systems, in particular the RLS and CD spectra, as well as, to an extent, the results of fluorescence anisotropy measurements. As we proceeded to the analysis of FTIR spectra, it was observed that the synergistic composition of C1 + AmB showed stronger interaction with the hydrophobic layer of the film. In turn, the crystallographic measurements performed for the synergistic composition revealed its impact on the multilayer's thickness. Finally, microcalorimetric measurements indicated that the synergistic composition had a lesser impact on the main phase transition temperature of the lipid, as compared to the respective compounds tested separately. The presented paper is the most detailed report to date pertaining to the synergism observed for the relevant systems analyzed in model biological films modified with sterols.
The study provides an in-depth spectroscopic, microcalorimetric, and crystallographic description of the molecular interactions analyzed for said molecular systems in model films formed of DPPC with the addition of sterols. Its results are particularly significant given the fact that AmB continues to be a clinically relevant antibiotic employed in the treatment of particularly severe internal mycoses and against otherwise resistant fungal strains.
{"title":"An advanced spectroscopic and crystallographic study on a synergistic composition of a 1,3,4-thiadiazole derivative and amphotericin B, in model biological liposomal system","authors":"Lidia Ślusarczyk , Klaudia Rząd , Daniel Kamiński , Mariusz Gagoś , Małgorzata Figiel , Arkadiusz Matwijczuk","doi":"10.1016/j.bbamem.2025.184432","DOIUrl":"10.1016/j.bbamem.2025.184432","url":null,"abstract":"<div><div>The paper presents the results of the first spectroscopic, microcalorimetric, and crystallographic study analyzing in great detail the strongly synergistic composition containing a selected <em>1,3,4</em>-thiadiazole derivative: <em>4-</em>(<em>5-methyl-1,3,4-thiadiazol-2-ylo</em>)<em>benzene-1,3-diol</em> (<strong>C1</strong>) and the antibiotic amphotericin B (<strong>AmB</strong>) in model biological DPPC films additionally modified with sterols: cholesterol (<strong>Chol</strong>) and ergosterol (<strong>Erg</strong>). The spectroscopic properties of the analyzed composition were studied with the use of spectroscopic methods, including: measurements of electronic fluorescence and absorption spectra with the technique of resonance light scattering (RLS), measurements of stationary fluorescence anisotropy and time-resolved fluorescence lifetimes with the method of single photon counting (TCSPC), circular dichroism spectra (CD), and infrared FTIR spectra. The mentioned methods were further complemented by including microcalorimetric DSC and crystallographic XRD analyses. All the measurements were taken in model biological systems formed as liposomal films and liposomal multilayers composed of DPPC and/or modified with sterols: cholesterol and ergosterol. The absorption spectra measured for C1/AmB and the synergistic composition thereof revealed differences indicating that while the antibiotic forms retained its known tendency to form aggregate in film systems, the synergistic composition had certain effects on the aggregational equilibrium. This was also reflected in the fluorescence emission measured for said systems, in particular the RLS and CD spectra, as well as, to an extent, the results of fluorescence anisotropy measurements. As we proceeded to the analysis of FTIR spectra, it was observed that the synergistic composition of C1 + AmB showed stronger interaction with the hydrophobic layer of the film. In turn, the crystallographic measurements performed for the synergistic composition revealed its impact on the multilayer's thickness. Finally, microcalorimetric measurements indicated that the synergistic composition had a lesser impact on the main phase transition temperature of the lipid, as compared to the respective compounds tested separately. The presented paper is the most detailed report to date pertaining to the synergism observed for the relevant systems analyzed in model biological films modified with sterols.</div><div>The study provides an in-depth spectroscopic, microcalorimetric, and crystallographic description of the molecular interactions analyzed for said molecular systems in model films formed of DPPC with the addition of sterols. Its results are particularly significant given the fact that AmB continues to be a clinically relevant antibiotic employed in the treatment of particularly severe internal mycoses and against otherwise resistant fungal strains.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184432"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144534809","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 : 2025-06-01DOI: 10.1016/j.bbamem.2025.184431
Justyna Kapral-Piotrowska , Agata Wawrzyniak , Jarosław Pawelec , Barbara Zarzyka , Roman Paduch , Jerzy Żuchowski , Adrianna Sławińska-Brych , Barbara Zdzisińska , Bartłomiej Pawlęga , Alicja Wójcik-Załuska , Ewa Baranowska-Wójcik , Joanna Jakubowicz-Gil , Wiesław I. Gruszecki , Bożena Pawlikowska-Pawlęga
Lensoside Aβ (LAβ) is a quercetin derivative isolated from the leaves and stems of the Lens culinaris cultivar Tina. Flavonoid-membrane interactions are crucial for their physiological and pharmacological activity. We have demonstrated the impact of LAβ on EYPC liposomes resembling the lipid phase of tumor cell membranes with the use of the 1H NMR technique and have examined its activity on HeLa cells for the first time. To study the interactions of the tested compound with lipids and proteins at the molecular level, the FTIR technique was applied. To reveal changes in morphology and ultrastructure as well as examine its effect on apoptosis induction and cell viability, SEM, TEM, light, and fluorescence microscopy, flow cytometry analysis, LIVE/DEAD assays were employed. The ability of LAβ to induce oxidative stress was determined by staining with DHR123. The FTIR analyses indicated that LAβ interacts with the PO2− groups in the polar head region. Moreover, a decrease in the relative protein concentration and changes in protein spectral profile in the amide I region were noted. Flavonoid reduced the viability of HeLa cells, which was correlated with the induction of apoptosis supported by SEM and TEM observations. Moreover, the addition of lensoside Aβ induced oxidative stress. These results confirm that lensoside Aβ may be used in novel therapeutic approaches for treating cervical cancer.
Lensoside a β (LAβ)是一种槲皮素衍生物，从植物天娜（Tina）的叶和茎中分离得到。黄酮类化合物与膜的相互作用对其生理和药理活性至关重要。我们利用1H NMR技术证明了LAβ对类似肿瘤细胞膜脂质期的EYPC脂质体的影响，并首次检测了其对HeLa细胞的活性。为了在分子水平上研究被测化合物与脂质和蛋白质的相互作用，应用了FTIR技术。采用扫描电镜（SEM）、透射电镜（TEM）、荧光显微镜（light and fluorescence microscopy）、流式细胞术（flow cytometry）、LIVE/DEAD等方法观察其形态学和超微结构的变化以及对细胞凋亡诱导和细胞活力的影响。DHR123染色法测定LAβ诱导氧化应激的能力。FTIR分析表明，LAβ与极性头区PO2 -基团相互作用。此外，还发现了相对蛋白浓度的降低和酰胺I区蛋白谱的变化。SEM和TEM观察显示，黄酮类化合物降低HeLa细胞活力与诱导凋亡有关。此外，晶状体皂苷Aβ的加入诱导了氧化应激。这些结果证实了透镜糖苷Aβ可能用于治疗宫颈癌的新方法。
{"title":"Interaction of lensoside Aβ with lipids and proteins of HeLa cells","authors":"Justyna Kapral-Piotrowska , Agata Wawrzyniak , Jarosław Pawelec , Barbara Zarzyka , Roman Paduch , Jerzy Żuchowski , Adrianna Sławińska-Brych , Barbara Zdzisińska , Bartłomiej Pawlęga , Alicja Wójcik-Załuska , Ewa Baranowska-Wójcik , Joanna Jakubowicz-Gil , Wiesław I. Gruszecki , Bożena Pawlikowska-Pawlęga","doi":"10.1016/j.bbamem.2025.184431","DOIUrl":"10.1016/j.bbamem.2025.184431","url":null,"abstract":"<div><div>Lensoside Aβ (LAβ) is a quercetin derivative isolated from the leaves and stems of the <em>Lens culinaris</em> cultivar Tina. Flavonoid-membrane interactions are crucial for their physiological and pharmacological activity. We have demonstrated the impact of LAβ on EYPC liposomes resembling the lipid phase of tumor cell membranes with the use of the <sup>1</sup>H NMR technique and have examined its activity on HeLa cells for the first time. To study the interactions of the tested compound with lipids and proteins at the molecular level, the FTIR technique was applied. To reveal changes in morphology and ultrastructure as well as examine its effect on apoptosis induction and cell viability, SEM, TEM, light, and fluorescence microscopy, flow cytometry analysis, LIVE/DEAD assays were employed. The ability of LAβ to induce oxidative stress was determined by staining with DHR123. The FTIR analyses indicated that LAβ interacts with the PO<sub>2</sub><sub>−</sub> groups in the polar head region. Moreover, a decrease in the relative protein concentration and changes in protein spectral profile in the amide I region were noted. Flavonoid reduced the viability of HeLa cells, which was correlated with the induction of apoptosis supported by SEM and TEM observations. Moreover, the addition of lensoside Aβ induced oxidative stress. These results confirm that lensoside Aβ may be used in novel therapeutic approaches for treating cervical cancer.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184431"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144330690","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 : 2025-06-01DOI: 10.1016/j.bbamem.2025.184429
Vanessa Gallego-Londoño , Gloria A. Santa-González , Juan M. Giraldo-Lorza , Mauricio Rojas , G. Bea A. Wisman , Steven de Jong , Marcela Manrique-Moreno
Cationic peptides offer a promising alternative for cancer treatment due to their ability to target cancer cells via standard membrane features, thereby overcoming intratumoral heterogeneity. This study investigates the cytotoxic activity and the membrane-disruptive effects of two snake venom-derived peptides, Crotalicidin (Ctn) and NA-CATH-ATRA-1-ATRA-1 (NA) in human breast cancer cells. Cell viability assays showed that both Ctn and NA significantly diminished the viability of MCF-7 and MDA-MB-231 cells, with NA showing greater potency, as indicated by lower IC50 values of 13.4 μM for MCF-7 and 6.4 μM for MDA-MB-231. Microscopy and flow cytometry revealed size reduction and increased granularity in treated cells. Further analyses indicated that the peptides induced membrane permeabilization, as evidenced by significant propidium iodide uptake, without significantly altering mitochondrial membrane potential. Apoptosis markers such as cleaved caspase-9 and PARP, were not detected by western blot.
Additionally, LDH release and confocal microscopic analysis supported the findings of membrane disruption. Finally, infrared spectroscopy (FT-IR) on lipid extracts revealed peptide-membrane interactions, resulting in phase transitions consistent with membrane disruption. These findings highlight the potent cytotoxic effects of Ctn and NA on breast cancer cells and their potential as novel therapeutic agents.
{"title":"Crotalicidin and NA-CATH-ATRA-1-ATRA-1 peptide-induced membrane disruption in human breast cancer cells","authors":"Vanessa Gallego-Londoño , Gloria A. Santa-González , Juan M. Giraldo-Lorza , Mauricio Rojas , G. Bea A. Wisman , Steven de Jong , Marcela Manrique-Moreno","doi":"10.1016/j.bbamem.2025.184429","DOIUrl":"10.1016/j.bbamem.2025.184429","url":null,"abstract":"<div><div>Cationic peptides offer a promising alternative for cancer treatment due to their ability to target cancer cells via standard membrane features, thereby overcoming intratumoral heterogeneity. This study investigates the cytotoxic activity and the membrane-disruptive effects of two snake venom-derived peptides, Crotalicidin (Ctn) and NA-CATH-ATRA-1-ATRA-1 (NA) in human breast cancer cells. Cell viability assays showed that both Ctn and NA significantly diminished the viability of MCF-7 and MDA-MB-231 cells, with NA showing greater potency, as indicated by lower IC<sub>50</sub> values of 13.4 μM for MCF-7 and 6.4 μM for MDA-MB-231. Microscopy and flow cytometry revealed size reduction and increased granularity in treated cells. Further analyses indicated that the peptides induced membrane permeabilization, as evidenced by significant propidium iodide uptake, without significantly altering mitochondrial membrane potential. Apoptosis markers such as cleaved caspase-9 and PARP, were not detected by western blot.</div><div>Additionally, LDH release and confocal microscopic analysis supported the findings of membrane disruption. Finally, infrared spectroscopy (FT-IR) on lipid extracts revealed peptide-membrane interactions, resulting in phase transitions consistent with membrane disruption. These findings highlight the potent cytotoxic effects of Ctn and NA on breast cancer cells and their potential as novel therapeutic agents.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184429"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144257297","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 : 2025-06-01DOI: 10.1016/j.bbamem.2025.184430
Veronika Osten , Dirk Schneider
The structure and the function of membrane proteins can be affected by the lipid bilayer environment, yet its impact is often neglected in in vitro studies where proteins are typically analyzed in membrane mimetics, mostly liposomal systems. It has been observed that the activity of the bacterial ATP-binding cassette (ABC) transporter BmrA (Bacillus multidrug resistance ATP) differs when measured in detergent vs. a model membrane environment, indicating that the physico-chemical properties of the membrane environment crucially affect the protein's activity. We now performed a systematic analysis to elucidate the impact of individual lipid/membrane properties on the activity of BmrA and identified three parameters controlling the BmrA activity in lipid bilayers: (i) the hydrophobic thickness of the membrane, (ii) a negative surface charge, and (iii) the packing of lipids in the acyl-chain and head group regions. Our study provides valuable insights into how a specific lipid composition can influence the basal ATPase activity of BmrA and emphasizes that the lipid composition should be carefully selected in in vitro studies of membrane proteins.
{"title":"Membrane properties control the ATPase activity of the ABC transporter BmrA","authors":"Veronika Osten , Dirk Schneider","doi":"10.1016/j.bbamem.2025.184430","DOIUrl":"10.1016/j.bbamem.2025.184430","url":null,"abstract":"<div><div>The structure and the function of membrane proteins can be affected by the lipid bilayer environment, yet its impact is often neglected in <em>in vitro</em> studies where proteins are typically analyzed in membrane mimetics, mostly liposomal systems. It has been observed that the activity of the bacterial ATP-binding cassette (ABC) transporter BmrA (<em>Bacillus</em> multidrug resistance ATP) differs when measured in detergent <em>vs.</em> a model membrane environment, indicating that the physico-chemical properties of the membrane environment crucially affect the protein's activity. We now performed a systematic analysis to elucidate the impact of individual lipid/membrane properties on the activity of BmrA and identified three parameters controlling the BmrA activity in lipid bilayers: (i) the hydrophobic thickness of the membrane, (ii) a negative surface charge, and (iii) the packing of lipids in the acyl-chain and head group regions. Our study provides valuable insights into how a specific lipid composition can influence the basal ATPase activity of BmrA and emphasizes that the lipid composition should be carefully selected in <em>in vitro</em> studies of membrane proteins.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184430"},"PeriodicalIF":2.8,"publicationDate":"2025-06-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144322078","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 : 2025-05-23DOI: 10.1016/j.bbamem.2025.184426
Valeriya Trusova, Uliana Malovytsia, Kateryna Vus, Olga Zhytniakivska, Galyna Gorbenko
The feasibility of engineering the sophisticated hybrid drug delivery platforms through the integration of phospholipid vesicles within a matrix of amyloid suspensions has been evaluated. Utilizing the equilibrium dialysis methodology and spectrofluorometric technique, the quantitative analysis of doxorubicin (DOX) encapsulation capacity of diverse phospholipid assemblies, amyloid suspensions, and their corresponding composite systems has been performed. Our findings revealed that the incorporation of negatively charged cardiolipin (CL) into phosphatidylcholine (PC) lipid vesicles significantly enhances DOX encapsulation and retention, while the addition of amyloid fibrils to charged liposomes has minimal impact on the drug binding. The neutral PC liposomes modified with insulin and lysozyme fibrillar suspensions exhibited improved doxorubicin encapsulation and retention compared to unmodified liposomes, thereby displaying a potential for reduced toxicity and prolonged drug action in vivo. Notably, amyloid fibrils alone were found to demonstrate the lower degree of DOX encapsulation and retention as compared to liposomes. Fluorimetric analysis suggests that the presence of insulin and lysozyme fibrils alters the microenvironment of DOX towards a more hydrophobic which is consistent with deeper bilayer penetration. Cumulative data from release kinetics and retention studies along with fluorescence measurements suggest that PC liposome-insulin fibril composites represent the most promising DOX nanocarriers, combining enhanced drug encapsulation, structural stability, and optimal drug location within the bilayer. The results obtained provide valuable insights into the design of protein-lipid nanomaterials for enhanced drug delivery, offering promising avenues for the development of more effective and targeted therapeutic strategies.
{"title":"Amyloid-liposome composites as hybrid platforms for doxorubicin delivery","authors":"Valeriya Trusova, Uliana Malovytsia, Kateryna Vus, Olga Zhytniakivska, Galyna Gorbenko","doi":"10.1016/j.bbamem.2025.184426","DOIUrl":"10.1016/j.bbamem.2025.184426","url":null,"abstract":"<div><div>The feasibility of engineering the sophisticated hybrid drug delivery platforms through the integration of phospholipid vesicles within a matrix of amyloid suspensions has been evaluated. Utilizing the equilibrium dialysis methodology and spectrofluorometric technique, the quantitative analysis of doxorubicin (DOX) encapsulation capacity of diverse phospholipid assemblies, amyloid suspensions, and their corresponding composite systems has been performed. Our findings revealed that the incorporation of negatively charged cardiolipin (CL) into phosphatidylcholine (PC) lipid vesicles significantly enhances DOX encapsulation and retention, while the addition of amyloid fibrils to charged liposomes has minimal impact on the drug binding. The neutral PC liposomes modified with insulin and lysozyme fibrillar suspensions exhibited improved doxorubicin encapsulation and retention compared to unmodified liposomes, thereby displaying a potential for reduced toxicity and prolonged drug action <em>in vivo</em>. Notably, amyloid fibrils alone were found to demonstrate the lower degree of DOX encapsulation and retention as compared to liposomes. Fluorimetric analysis suggests that the presence of insulin and lysozyme fibrils alters the microenvironment of DOX towards a more hydrophobic which is consistent with deeper bilayer penetration. Cumulative data from release kinetics and retention studies along with fluorescence measurements suggest that PC liposome-insulin fibril composites represent the most promising DOX nanocarriers, combining enhanced drug encapsulation, structural stability, and optimal drug location within the bilayer. The results obtained provide valuable insights into the design of protein-lipid nanomaterials for enhanced drug delivery, offering promising avenues for the development of more effective and targeted therapeutic strategies.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184426"},"PeriodicalIF":2.8,"publicationDate":"2025-05-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144130922","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 : 2025-05-22DOI: 10.1016/j.bbamem.2025.184424
Evelyn A. Okorafor , Emma A. Gordon , Indra D. Sahu , Muhammad Zeeshan Shah , Dominik Konkolewicz , Gary A. Lorigan
Membrane bilayers are complex three-dimensional structures whose molecular events in the deep dimensions of membrane lipids are crucial for understanding membrane function. This study investigates the interaction of coexisting membrane domains in terms of hydrophobicity, alkyl chain order, and fluidity using Styrene Maleic Acid (SMA) copolymers as membrane mimics. We employed continuous wave electron paramagnetic resonance spectroscopy (CW-EPR) to characterize the structural dynamic properties of membrane domains without separation. Lipid-spin probe vesicles were prepared using phospholipids with varying degrees of saturation (DOPC, POPC, DMPC, and DSPC) and doxyl spin-labeled phospholipids at different depths (5, 12, and 16-doxyl PC) as membrane probes. These vesicles were titrated with two SMA polymers of different hydrophobic tail lengths. Dynamic light scattering (DLS) confirmed the formation of Styrene Maleic Acid lipid nanoparticles (SMALPs). CW-EPR spectroscopy was used to characterize the dynamic properties of vesicles incorporated into the SMALP systems. Analysis of the CW-EPR spectral line shape data revealed that the hydrophobic tail of SMA, the degree of lipid saturation, and the length of phospholipids significantly affect membrane fluidity and alkyl chain ordering, as well as lipid interactions. Notably, samples containing DSPC, a fully saturated longer-chain phospholipid, and those containing SMA exhibited increased rigidity of motion, reduced fluidity, and improved ordering of the alkyl chain in the membrane. This study provides crucial insights into the molecular dynamics of membrane bilayers and the impact of SMA copolymers on membrane properties, contributing to our understanding of fundamental membrane functions such as lateral movement of proteins and lipids.
{"title":"Influence of lipid saturation on the structural properties of styrene maleic acid lipid nanoparticles (SMALPs)","authors":"Evelyn A. Okorafor , Emma A. Gordon , Indra D. Sahu , Muhammad Zeeshan Shah , Dominik Konkolewicz , Gary A. Lorigan","doi":"10.1016/j.bbamem.2025.184424","DOIUrl":"10.1016/j.bbamem.2025.184424","url":null,"abstract":"<div><div>Membrane bilayers are complex three-dimensional structures whose molecular events in the deep dimensions of membrane lipids are crucial for understanding membrane function. This study investigates the interaction of coexisting membrane domains in terms of hydrophobicity, alkyl chain order, and fluidity using Styrene Maleic Acid (SMA) copolymers as membrane mimics. We employed continuous wave electron paramagnetic resonance spectroscopy (CW-EPR) to characterize the structural dynamic properties of membrane domains without separation. Lipid-spin probe vesicles were prepared using phospholipids with varying degrees of saturation (DOPC, POPC, DMPC, and DSPC) and doxyl spin-labeled phospholipids at different depths (5, 12, and 16-doxyl PC) as membrane probes. These vesicles were titrated with two SMA polymers of different hydrophobic tail lengths. Dynamic light scattering (DLS) confirmed the formation of Styrene Maleic Acid lipid nanoparticles (SMALPs). CW-EPR spectroscopy was used to characterize the dynamic properties of vesicles incorporated into the SMALP systems. Analysis of the CW-EPR spectral line shape data revealed that the hydrophobic tail of SMA, the degree of lipid saturation, and the length of phospholipids significantly affect membrane fluidity and alkyl chain ordering, as well as lipid interactions. Notably, samples containing DSPC, a fully saturated longer-chain phospholipid, and those containing SMA exhibited increased rigidity of motion, reduced fluidity, and improved ordering of the alkyl chain in the membrane. This study provides crucial insights into the molecular dynamics of membrane bilayers and the impact of SMA copolymers on membrane properties, contributing to our understanding of fundamental membrane functions such as lateral movement of proteins and lipids.</div></div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":"1867 5","pages":"Article 184424"},"PeriodicalIF":2.8,"publicationDate":"2025-05-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144141407","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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