High-resolution label-free 3D mapping of extracellular pH of single living cells

Abstract: Dynamic mapping of extracellular pH (pHe) at the single-cell level is critical for understanding the role of H+ in cellular and subcellular processes, with particular importance in cancer. While several pHe sensing techniques have been developed, accessing this information at the single-cell level requires improvement in sensitivity, spatial and temporal resolution. We report on a zwitterionic label-free pH nanoprobe that addresses these long-standing challenges. The probe has a sensitivity > 0.01 units, 2 ms response time, and 50 nm spatial resolution. The platform was integrated into a double-barrel nanoprobe combining pH sensing with feedback-controlled distance dependance via Scanning Ion Conductance Microscopy. This allows for the simultaneous 3D topographical imaging and pHe monitoring of living cancer cells. These classes of nanoprobes were used for real-time high spatiotemporal resolution pHe mapping at the subcellular level and revealed tumour heterogeneity of the peri-cellular environments of melanoma and breast cancer cells

New strategies for detailed visualisation and analysis of stem cell-material interactions

The success of stem cell therapy and regenerative medicine relies on providing bio- chemical and biophysical cues to safely and reliably control cell fate transitions. It has now been recognised that recapitulating the physical attributes of the stem cell microenvironment is a key feature in controlling cell behaviour. Of particular interest is the ability of the extracellular microenvironment to present surface topographical features that contribute to control of cell attachment and morphology, which have a direct impact on maintenance of cell phenotype and gene expression. This realisation has led to the development of materials with surface topography to probe and control the stem cell behaviour. In this thesis, the influence of material topography on cell behaviour is investigated through the use of both microscale and nanoscale topography, with a particular focus on visualisation strategies to characterise the features of the cell-material interface. Firstly, microscale topography in the form of microgrooves, is used to study the impact of topography at the level of the nucleus in neural stem cells. Through the development of new visualisation methods, incorporating state-of-the-art focused-ion beam scanning electron microscopy and scanning ion conductance microscopy, the nuclei of cells cul- tured on microgrooved topography are shown to correlatively remodel their nuclear architecture, epigenetics and mechanical properties. Secondly, using nanoscale topography in the form of high aspect ratio nanoneedle arrays, the cell-material interface is investigated at the level of the plasma membrane in human mesenchymal stem cells. Nanoneedles, typically used to deliver a variety of therapeutically relevant payloads, are shown to create a tight interface with the cell membrane, while triggering changes in plasma membrane architecture and inducing the activation of endocytosis pathways, which contributes significantly to payload delivery at the interface.Open Acces

Perspectives of patients and healthcare professionals on metabolic monitoring of adult prescribed second-generation antipsychotics for severe mental illness: A meta-synthesis.

ObjectivesWe conducted a meta-synthesis of qualitative studies to synthesize the views of psychiatric patients on second-generation antipsychotics (SGAs) and the healthcare providers about the metabolic monitoring of adult-prescribed SGAs.MethodsA systematic search was conducted in four databases through SCOPUS, PubMed, EMBASE, and CINAHL to identify qualitative studies of patients' and healthcare professionals' perspectives on the metabolic monitoring of SGAs. Initially, titles and abstracts were screened to exclude articles that were not relevant followed by full-text reading. Study quality was assessed by using Critical Appraisal Skills Program (CASP) criteria. Themes were synthesized and presented as per the Interpretive data synthesis process (Evans D, 2002).ResultsA total of 15 studies met the inclusion criteria and were analyzed in meta-synthesis. Four themes were identified: 1. Barriers to metabolic monitoring; 2. Patient related concerns to metabolic monitoring; 3. Support system by mental health services to promote metabolic monitoring; and 4. Integrating physical health with mental health services. From the participants' perspectives, barriers to metabolic monitoring were accessibility of services, lack of education and awareness, time/resource constraints, financial hardship, lack of interest on metabolic monitoring, patient capacity and motivation to maintain physical health and role confusion and impact on communication. Education and training on monitoring practices as well as integrated mental health services for metabolic monitoring to promote quality and safe use of SGAs are the most likely approaches to promote adherence to best practices and minimize treatment-related metabolic syndrome in this highly vulnerable cohort.ConclusionThis meta-synthesis highlights key barriers from the perspectives of patients and healthcare professionals regarding the metabolic monitoring of SGAs. These barriers and suggested remedial strategies are important to pilot in the clinical setting and to assess the impact of the implementation of such strategies as a component of pharmacovigilance to promote the quality use of SGAs as well as to prevent and/or manage SGAs-induced metabolic syndrome in severe and complex mental health disorders

Perspectives of patients and healthcare professionals on metabolic monitoring of adult prescribed second-generation antipsychotics for severe mental illness: A meta-synthesis

Objectives We conducted a meta-synthesis of qualitative studies to synthesize the views of psychiatric patients on second-generation antipsychotics (SGAs) and the healthcare providers about the metabolic monitoring of adult-prescribed SGAs. Methods A systematic search was conducted in four databases through SCOPUS, PubMed, EMBASE, and CINAHL to identify qualitative studies of patients’ and healthcare professionals’ perspectives on the metabolic monitoring of SGAs. Initially, titles and abstracts were screened to exclude articles that were not relevant followed by full-text reading. Study quality was assessed by using Critical Appraisal Skills Program (CASP) criteria. Themes were synthesized and presented as per the Interpretive data synthesis process (Evans D, 2002). Results A total of 15 studies met the inclusion criteria and were analyzed in meta-synthesis. Four themes were identified: 1. Barriers to metabolic monitoring; 2. Patient related concerns to metabolic monitoring; 3. Support system by mental health services to promote metabolic monitoring; and 4. Integrating physical health with mental health services. From the participants’ perspectives, barriers to metabolic monitoring were accessibility of services, lack of education and awareness, time/resource constraints, financial hardship, lack of interest on metabolic monitoring, patient capacity and motivation to maintain physical health and role confusion and impact on communication. Education and training on monitoring practices as well as integrated mental health services for metabolic monitoring to promote quality and safe use of SGAs are the most likely approaches to promote adherence to best practices and minimize treatment-related metabolic syndrome in this highly vulnerable cohort. Conclusion This meta-synthesis highlights key barriers from the perspectives of patients and healthcare professionals regarding the metabolic monitoring of SGAs. These barriers and suggested remedial strategies are important to pilot in the clinical setting and to assess the impact of the implementation of such strategies as a component of pharmacovigilance to promote the quality use of SGAs as well as to prevent and/or manage SGAs-induced metabolic syndrome in severe and complex mental health disorders

Immunogold FIB-SEM: combining volumetric ultrastructure visualization with 3D biomolecular analysis to dissect cell-environment interactions

Volumetric imaging techniques capable of correlating structural and functional information with nanoscale resolution are necessary to broaden the insight into cellular processes within complex biological systems. The recent emergence of focused ion beam scanning electron microscopy (FIB‐SEM) has provided unparalleled insight through the volumetric investigation of ultrastructure; however, it does not provide biomolecular information at equivalent resolution. Here, immunogold FIB‐SEM, which combines antigen labeling with in situ FIB‐SEM imaging, is developed in order to spatially map ultrastructural and biomolecular information simultaneously. This method is applied to investigate two different cell–material systems: the localization of histone epigenetic modifications in neural stem cells cultured on microstructured substrates and the distribution of nuclear pore complexes in myoblasts differentiated on a soft hydrogel surface. Immunogold FIB‐SEM offers the potential for broad applicability to correlate structure and function with nanoscale resolution when addressing questions across cell biology, biomaterials, and regenerative medicine

High-resolution label-free 3D mapping of extracellular pH of single living cells.

Dynamic mapping of extracellular pH (pHe) at the single-cell level is critical for understanding the role of H+ in cellular and subcellular processes, with particular importance in cancer. While several pHe sensing techniques have been developed, accessing this information at the single-cell level requires improvement in sensitivity, spatial and temporal resolution. We report on a zwitterionic label-free pH nanoprobe that addresses these long-standing challenges. The probe has a sensitivity > 0.01 units, 2 ms response time, and 50 nm spatial resolution. The platform was integrated into a double-barrel nanoprobe combining pH sensing with feedback-controlled distance dependance via Scanning Ion Conductance Microscopy. This allows for the simultaneous 3D topographical imaging and pHe monitoring of living cancer cells. These classes of nanoprobes were used for real-time high spatiotemporal resolution pHe mapping at the subcellular level and revealed tumour heterogeneity of the peri-cellular environments of melanoma and breast cancer cells

High-resolution label-free 3D mapping of extracellular pH of single living cells

Abstract: Dynamic mapping of extracellular pH (pHe) at the single-cell level is critical for understanding the role of H+ in cellular and subcellular processes, with particular importance in cancer. While several pHe sensing techniques have been developed, accessing this information at the single-cell level requires improvement in sensitivity, spatial and temporal resolution. We report on a zwitterionic label-free pH nanoprobe that addresses these long-standing challenges. The probe has a sensitivity > 0.01 units, 2 ms response time, and 50 nm spatial resolution. The platform was integrated into a double-barrel nanoprobe combining pH sensing with feedback-controlled distance dependance via Scanning Ion Conductance Microscopy. This allows for the simultaneous 3D topographical imaging and pHe monitoring of living cancer cells. These classes of nanoprobes were used for real-time high spatiotemporal resolution pHe mapping at the subcellular level and revealed tumour heterogeneity of the peri-cellular environments of melanoma and breast cancer cells