Local Membrane Deformations Activate Ca2+-Dependent K+ and Anionic Currents in Intact Human Red Blood Cells

BACKGROUND: The mechanical, rheological and shape properties of red blood cells are determined by their cortical cytoskeleton, evolutionarily optimized to provide the dynamic deformability required for flow through capillaries much narrower than the cell's diameter. The shear stress induced by such flow, as well as the local membrane deformations generated in certain pathological conditions, such as sickle cell anemia, have been shown to increase membrane permeability, based largely on experimentation with red cell suspensions. We attempted here the first measurements of membrane currents activated by a local and controlled membrane deformation in single red blood cells under on-cell patch clamp to define the nature of the stretch-activated currents. METHODOLOGY/PRINCIPAL FINDINGS: The cell-attached configuration of the patch-clamp technique was used to allow recordings of single channel activity in intact red blood cells. Gigaohm seal formation was obtained with and without membrane deformation. Deformation was induced by the application of a negative pressure pulse of 10 mmHg for less than 5 s. Currents were only detected when the membrane was seen domed under negative pressure within the patch-pipette. K(+) and Cl(-) currents were strictly dependent on the presence of Ca(2+). The Ca(2+)-dependent currents were transient, with typical decay half-times of about 5-10 min, suggesting the spontaneous inactivation of a stretch-activated Ca(2+) permeability (PCa). These results indicate that local membrane deformations can transiently activate a Ca(2+) permeability pathway leading to increased [Ca(2+)](i), secondary activation of Ca(2+)-sensitive K(+) channels (Gardos channel, IK1, KCa3.1), and hyperpolarization-induced anion currents. CONCLUSIONS/SIGNIFICANCE: The stretch-activated transient PCa observed here under local membrane deformation is a likely contributor to the Ca(2+)-mediated effects observed during the normal aging process of red blood cells, and to the increased Ca(2+) content of red cells in certain hereditary anemias such as thalassemia and sickle cell anemia

Etude du comportement dynamique des canaux ioniques de la membrane du globule rouge humain

Erythrocytes are one of the most frequently cellular models used so far to decipher the physiology of membrane transport. However, if pumps, antiporters and cotransporters are now well defined, the molecular identity and the physiological role of the conductive pathways present in the membrane of blood cell (RBC) is still elusive albeit the growing evidences of their role in physiological and pathological conditions. The present work, using the patch clamp technique and biophysical studies, show that:1) The changes in osmotic fragility observed after Gardos channel dependent dehydration is mainly not due to rehydration of the cell via membrane transporters but rather due to change in membrane properties elicited by the Ca2+ loading and must reflect a specific calcium-induced lytic vulnerability of the membrane causing rupture before the cells attain their maximal spherical volumes.2) The Gardos channel can be transiently activated when seal formation induces membrane deformation. This phenomenon can result only from activation of a permeability pathway with a finite Ca2+ conductance. This transient activity generates secondary transient anionic channel activity that has been studied further.3) The diversity of anionic channel activities recorded in normal human RBCs, as well as in P. falciparum-infected RBCs, corresponds to different kinetic modalities of a unique type of maxi anion channel with multiple conductance levels, gating properties and pharmacology.Finally, the present work contributes to the understanding of the role of ion channels in RBCs and opens questions on the contribution of ion channel in the rheological properties of RBCs.Les érythrocytes sont un des modèles pour l’étude des voies de transport membranaire. Si les pompes, les antiports et les cotransports sont bien définies, l’identité moléculaire et le rôle physiologique des voies de conductances du globule rouge restent méconnus. Le présent travail réalisé à l’aide des techniques de patch-clamp montre que :1) Les changements de fragilité osmotique observés lors d’une déhydratation des cellules après stimulation du canal Gardos, ne sont pas dus à une réhydration des cellules via l’activation de transporteurs, mais à des changements de propriétés de la membrane liées à l’augmentation du [Ca2+]i. Elle reflète une vulnérabilité spécifique de la membrane au Ca2+, induisant la rupture de la membrane avant que la cellule n’ait atteint son volume critique.2) Le canal Gardos peut-être activé transitoirement lors d’épisode de déformation de la membrane. Ce phénomène ne peut-être que le résultat de l’activation d’une voie de perméabilité au Ca2+ ayant une conductance déterminée.3) L’activité transitoire du canal Gardos active secondairement une voie de conductance anionique. La diversité des canaux anioniques précédemment décrites dans la membrane des érythrocytes humains correspond à l’activité d’un unique canal anionique de type maxi-chlorure ayant des modalités d’activation et de fonctionnement (état d’ouverture multiple, cinétique, pharmacologie…) différentes en fonction des conditions expérimentales.Ce travail contribue à une meilleure compréhension des canaux ioniques présents dans la membrane du globule rouge et permet d’envisager la participation de ces canaux dans la régulation des propriétés rhéologiques des globules rouges

Étude du comportement dynamique des canaux ioniques de la membrane du globule rouge humain

Erythrocytes are one of the cellular models frequently used to decipher the physiology of membrane transport. However, if pumps, antiporters and cotransporters are now well defined, the molecular identity and the physiological role of the conductive pathways present in the membrane of red blood cell (RBC) is still elusive albeit the growing evidences of their role in physiological and pathological conditions. The present work, using the patch clamp technique and biophysical studies, shows that: 1) The changes in osmotic fragility observed after Gardos channel dependent dehydration is mainly not due to rehydration of the cell via membrane transporters but rather due to change in membrane properties elicited by the Ca2+ loading. It reflects a specific calcium-induced lytic vulnerability of the membrane leading to rupture before the cells attain their maximal spherical volumes. 2) The Gardos channel can be transiently activated when seal formation induces membrane deformation. This phenomenon can result only from activation of a permeability pathway with a finite Ca2+ conductance. This transient activity generates secondary transient anionic channel activity that has been studied further. 3) The diversity of anionic channel activities recorded in normal human RBCs, as well as in P. falciparum-infected RBCs, corresponds to different kinetic modalities of a unique type of maxi anion channel with multiple conductance levels, gating properties and pharmacology. Finally, the present work contributes to the understanding of the role of ion channels in RBCs and opens questions on the contribution of ion channels in the rheological properties of RBCs. Keywords: erythrocyte, ionic channels, osmotic fragility, patch clamp technique.Les érythrocytes sont un des modèles pour l étude des voies de transport membranaire. Si les pompes, les antiports et les cotransports sont bien définis, l identité moléculaire et le rôle physiologique des voies de conductances du globule rouge restent méconnus. Le présent travail réalisé à l aide des techniques de patch-clamp montre que : 1) Les changements de fragilité osmotique observés lors d une déhydratation des cellules après stimulation du canal Gardos, ne sont pas dus à une réhydration des cellules via l activation de transporteurs, mais à des changements de propriétés de la membrane liées à l augmentation du [Ca2+]i. Elle reflète une vulnérabilité spécifique de la membrane au Ca2+, induisant la rupture de la membrane avant que la cellule n ait atteint son volume critique. 2) Le canal Gardos peut-être activé transitoirement lors d épisode de déformation de la membrane. Ce phénomène ne peut être que le résultat de l activation d une voie de perméabilité au Ca2+ ayant une conductance déterminée. 3) L activité transitoire du canal Gardos active secondairement une voie de conductance anionique. La diversité des canaux anioniques précédemment décrits dans la membrane des érythrocytes humains correspond à l activité d un unique canal anionique de type maxi-chlorure ayant des modalités d activation et de fonctionnement (état d ouverture multiple, cinétique, pharmacologie ) différentes en fonction des conditions expérimentales. Ce travail contribue à une meilleure compréhension des canaux ioniques présents dans la membrane du globule rouge et permet d envisager la participation de ces canaux dans la régulation des propriétés rhéologiques des globules rouges. Mots clés: érythrocyte, canaux ioniques, fragilité osmotique, patch clamp.RENNES1-BU Sciences Philo (352382102) / SudocROSCOFF-Observ.Océanol. (292393008) / SudocSudocFranceF

Mother and neonate suffering from COVID-19 infection. Is there any risk of vertical transmission? A case report

Objectives: This study was carried out to visualize possible infection of the fetus during pregnancy through verticaltransmission.Material and methods: A descriptive observational study was conducted on a 39th week pregnant woman who wassuspected COVID-19. Pregnancy was ended with a caesarean section due to epidemiological reasons. The mother andnewborn were examined for COVID-19 infection using PCR swabs.Results: At first there were discrepancies between SARS-CoV-2 test results from the nasopharyngeal swabs of the motherand the neonate. The mother tested positive, whereas the newborn tested negative. However, neonate’s control swab wasrepeated within the following 48 hours and revealed a positive test result.Conclusions: The risk of vertical transmission with SARS-COV2 is possible, therefore it is very important to isolate pregnantwomen from infected people to protect mothers and newborns

Surgical Management for Refractory Bleb Dysesthesia

Purpose. To present long-term results of modified bleb-limiting conjunctivoplasty as a successful treatment for intractable bleb dysesthesia and to review the literature on the surgical management of dysesthetic bleb. Methods. Consecutive case series and literature review. We present four cases of surgically reduced painful blebs. Our technique consisted of the following steps: (1) conjunctival, radial incision to the bare sclera in the desired limit of the bleb; (2) suturing with buried, interrupted sutures at the nearest edge of the filtering bleb; (3) lower limbal peritomy including unwanted area of the extended bleb; (4) dissection and removal of the underlying fibrous tissue when present; (5) conjunctival and resorbable sutures. In addition, a systematic literature review was performed. Only reports presenting outcomes of surgical treatment of bleb dysesthesia after filtering procedure were included in review. Results. Four eyes were included consecutively in the study in a period of 4 years. On average, they developed circumferential bleb dysesthesia 9.3 ± 4.7 months after uneventful combined phacotrabeculectomy with Mitomycin C as primary procedure. Surgical reduction was decided after failure of lubricants in controlling ocular discomfort. Two cases showed a dense fibrous tissue beneath the conjunctiva that was excised to ensure filtration. In all cases, a rapid disappearance of symptoms with very good aesthetic and functional outcome was observed. After 12-month follow-up, patients remained asymptomatic and maintained intraocular pressure of 10.7 ± 1.2 mmHg without treatment. A systematic review of the literature obtained 15 eligible case series (n = 123) with rates of success within 46–100%, favoring less aggressive approaches to reduce bleb size. Conclusion. Bleb dysesthesia is a rare complication of filtering glaucoma surgery. This modified bleb-limiting conjunctivoplasty technique (with removal of subjacent fibrous tissue if present) is able to target the underlying etiology providing ocular discomfort relief while maintaining bleb function and may be considered as first-choice surgical treatment

Anion conductance of the human red cell is carried by a maxi-anion channel

International audienceHistorically, the anion transport through the human red cell membrane has been perceived to be mediated by Band 3, in the two-component concept with the large electroneutral anion exchange accompanied by the conductance proper, which dominated the total membrane conductance. The status of anion channels proper has never been clarified, and the informations obtained by different groups of electrophysiologists are rather badly matched. This study, using the cell-attached configuration of the patch-clamp technique, rationalizes and explains earlier confusing results by demonstrating that the diversity of anionic channel activities recorded in human erythrocytes corresponds to different kinetic modalities of a unique type of maxi-anion channel with multiple conductance levels and probably multiple gating properties and pharmacology, depending on conditions. It demonstrates the role of activator played by serum in the recruitment of multiple new conductance levels showing very complex kinetics and gating properties upon serum addition. These channels, which seem to be dormant under normal physiological conditions, are potentially activable and could confer a far higher anion conductance to the red cell than the ground leak mediated by Band 3

Clinical results of diffractive, refractive, hybrid multifocal, and monofocal intraocular lenses

PURPOSE: To present the outcomes of hybrid multifocal and monofocal intraocular lenses (IOLs) and to compare with refractive and diffractive multifocal IOLs (MFIOLs). METHODS: Three hundred twenty eyes (160 patients) underwent cataract surgery with randomized IOLs bilateral implantation. Changes in uncorrected and distance-corrected logMAR distance, intermediate and near (UNVA and DCNVA) visual acuity (VA), contrast sensitivity (CS), presence of dysphotopsia, spectacle independence, and patient satisfaction were analyzed. RESULTS: Postoperative VA in the hybrid (OptiVis) group was improved in all distances (p < 0.001). OptiVis acted superiorly to monofocal IOLs in UNVA and DCNVA (p < 0.001 for both) and to refractive ones in DCNVA (p < 0.005). Distance, mesopic, without glare CS in OptiVis was lower than in the monofocal group and similar to other MFIOLs. No differences in dysphotopsia pre- and postoperatively and spectacle independence in near for OptiVis and refractive MFIOLs were detected. OptiVis patients were more satisfied than those with monofocal IOLs (p=0.015). CONCLUSIONS: After cataract surgery, patients with OptiVis improved VA in all distances. Near and intermediate VA was better than monofocal, and DCNVA was better than the refractive group. CS was lower in OptiVis than in the monofocal group, but there was no difference between MFIOLs. Patient satisfaction was higher in OptiVis than in the monofocal group. This trial is registered with NCT03512626