Membrane Disorders Research Articles

Novel mechanism of hereditary pyropoikilocytosis phenotype due to co‐inheritance of β globin and α spectrin mutations

Novel mechanism of hereditary pyropoikilocytosis phenotype due to co‐inheritance of β globin and α spectrin mutations

Biophysical and rheological biomarkers of red blood cell physiology and pathophysiology.

This review summarizes the significant biophysical and rheological aspects of red blood cell physiology and pathophysiology in relation to recent advances in microfluidic biomarker assays and emerging targeted or curative intent therapies. Alterations in red cell biophysical properties and blood rheology have been associated with numerous hematologic and circulatory disorders. Recent advances in biomarker assays enable effective assessment of these biophysical and rheological properties in normoxia or physiological hypoxia in a clinically meaningful way. There are emerging targeted or curative therapies that aim to improve red cell pathophysiology, especially in the context of inherited hemoglobin disorders, such as sickle cell disease. Red cell pathophysiology can be therapeutically targeted and the improvements in membrane and cellular biophysics and blood rheology can now be feasibly assessed via new microfluidic biomarker assays. Recent advances provide a new hope and novel treatment options for major red cell ailments, including inherited hemoglobin disorders, membrane disorders, and other pathologies of the red cell, such as malaria.

Membrane-partitioned cell wall synthesis in mycobacteria.

Many antibiotics target the assembly of cell wall peptidoglycan, an essential, heteropolymeric mesh that encases most bacteria. In rod-shaped bacteria, cell wall elongation is spatially precise yet relies on limited pools of lipid-linked precursors that generate and are attracted to membrane disorder. By tracking enzymes, substrates, and products of peptidoglycan biosynthesis in Mycobacterium smegmatis, we show that precursors are made in plasma membrane domains that are laterally and biochemically distinct from sites of cell wall assembly. Membrane partitioning likely contributes to robust, orderly peptidoglycan synthesis, suggesting that these domains help template peptidoglycan synthesis. The cell wall-organizing protein DivIVA and the cell wall itself promote domain homeostasis. These data support a model in which the peptidoglycan polymer feeds back on its membrane template to maintain an environment conducive to directional synthesis. Our findings are applicable to rod-shaped bacteria that are phylogenetically distant from M. smegmatis, indicating that horizontal compartmentalization of precursors may be a general feature of bacillary cell wall biogenesis.

Change in Proteolytic Profile in Heifers After Oligofructose Overload.

Laminitis in cattle is an important underlying cause of lameness, which leads to a significant reduction in economic and animal welfare. Nevertheless, the disordered pathological processes of laminitis remain unclear. Several proteinases are probably involved in the disorder of basement membrane (BM) metabolism in laminitis, for instance, matrix metalloproteinases (MMPs), neutrophil elastase (NE), and myeloperoxidase (MPO). This study aimed to investigate the change in proteolytic profile in circulating and lamellar tissues using an oligofructose (OF) overload-induced laminitis model in heifers. Twelve clinically healthy and nonlame Chinese Holstein heifers were recruited and randomly divided into two groups: OF-induced and control (CON). The OF-induced heifers group (n = 6) was administered 17 g/kg of body weight (BW) of OF dissolved in 2 L/100 kg of BW of tap water via the oral-rumen tube. The CON group (n = 6) was given an equal volume of tap water. The plasma samples were collected 0, 6, 12, 18, 24, 36, 48, 60, and 72 h after administration, and the lamellar samples were collected 72 h after euthanasia. The plasma samples were analyzed by zymography and reverse zymography. Histological examination, zymography, reverse zymography, and Western blot of lamellar samples were conducted. In the plasma of the OF-induced group, the pro-MMP9 activity increased from 36 h (P < 0.001) to 60 h (P < 0.05). Moreover, the plasma tissue inhibitors of metalloproteinase 1 (TIMP1) activity decreased after 18 h (P < 0.05), while the ratio of pro-MMP9 to TIMP1 and TIMP2 increased after 18 h (P < 0.001) and 48 h (P < 0.05), respectively. The act-MMP2, pro-MMP9, and act-MMP9 activities increased in the lamellar tissue of the OF-induced group compared with the CON group (P < 0.05). In addition, the expression of lamellar NE protein was higher in the OF-induced group (P < 0.01), while no change was found in lamellar MPO protein compared with the CON group. In conclusion, increased pro-MMP9 combined with decreased TIMP1 activity in the circulation might have caused the activation of blood neutrophils, while the activation of proteolytic enzymes in lamellar tissue probably led to the dysfunction of BM in the OF-induced group.

Single Layer Centrifugation Improves the Quality of Fresh Donkey Semen and Modifies the Sperm Ability to Interact with Polymorphonuclear Neutrophils.

Simple SummaryDonkey Artificial Insemination (AI) with frozen/thawed semen results in poor fertility outcomes. Jennies show a significant post-AI endometrial reaction, with a large amount of defense cells—polymorphonuclear neutrophils (PMN)—migrating to the uterine lumen. Seminal plasma (SP) has a detrimental effect on sperm conservation and its removal is a necessary step in the semen freezing protocol. However, several SP proteins seem to control sperm-PMN binding. Single layer centrifugation (SLC) with colloids, which has been used to select spermatozoa and improve reproductive performance in different species, is known to remove SP proteins attached to the sperm membrane. In this study, two experiments were performed. The first one compared the quality of SLC-selected and non-selected fresh donkey spermatozoa. In the second experiment, PMN obtained from the peripheral blood were co-incubated with selected and unselected spermatozoa, and the interaction between PMN and spermatozoa was analyzed. In conclusion, SLC of fresh donkey semen increases the proportion of functionally intact spermatozoa and appears to remove the SP proteins that inhibit sperm-PMN binding, thus increasing sperm phagocytosis by PMN.This study sought to determine whether single layer centrifugation (SLC) of fresh donkey semen with Equicoll has any impact on sperm quality parameters and on the modulation of endometrial reaction following semen deposition using an in vitro model. Seventeen ejaculates from five jackasses were obtained using an artificial vagina and diluted in a skim-milk extender. Samples were either selected through SLC (Equicoll) or non-treated (control). Two experiments were performed. The first one consisted of incubating selected or non-selected spermatozoa at 38 °C for 180 min. Integrity and lipid disorder of sperm plasma membrane, mitochondrial membrane potential, and intracellular levels of calcium and reactive oxygen species were evaluated at 0, 60, 120, and 180 min. In the second experiment, polymorphonuclear neutrophils (PMN) isolated from jennies blood were mixed with selected and unselected spermatozoa. Interaction between spermatozoa and PMN was evaluated after 0, 60, 120, and 180 min of co-incubation at 38 °C. SLC-selection increased the proportions of spermatozoa with an intact plasma membrane and low lipid disorder, of spermatozoa with high mitochondrial membrane potential and with high calcium levels, and of progressively motile spermatozoa. In addition, selection through SLC augmented the proportion of phagocytosed spermatozoa, which supported the modulating role of seminal plasma proteins on sperm-PMN interaction. In conclusion, SLC of fresh donkey semen increases the proportions of functionally intact and motile spermatozoa, and appears to remove the seminal plasma proteins that inhibit sperm-PMN binding.

Molecular Sizes and Antibacterial Performance Relationships of Flexible Ionic Liquid Derivatives.

Cationic agents, such as ionic liquids (ILs)-based species, have broad-spectrum antibacterial activities. However, the antibacterial mechanisms lack systematic and molecular-level research, especially for Gram-negative bacteria, which have highly organized membrane structures. Here, we designed a series of flexible fluorescent diketopyrrolopyrrole-based ionic liquid derivatives (ILDs) with various molecular sizes (1.95-4.2 nm). The structure-antibacterial activity relationships of the ILDs against Escherichia coli (E. coli) were systematically studied thorough antibacterial tests, fluorescent tracing, morphology analysis, molecular biology, and molecular dynamics (MD) simulations. ILD-6, with a relatively small molecular size, could penetrate through the bacterial membrane, leading to membrane thinning and intracellular activities. ILD-6 showed fast and efficient antimicrobial activity. With the increase of molecular sizes, the corresponding ILDs were proven to intercalate into the bacterial membrane, leading to the destabilization of the lipid bilayer and further contributing to the antimicrobial activities. Moreover, the antibacterial activity of ILD-8 was limited, where the size was not large enough to introduce significant membrane disorder. Relative antibacterial experiments using another common Gram-negative bacteria, Pseudomonas aeruginosa (PAO1), further confirmed the proposed structure-antibacterial activity relationships of ILDs. More impressively, both ILD-6 and ILD-12 displayed significant in vivo therapeutic effects on the PAO1-infected rat model, while ILD-8 performed poorly, which confirmed the antibacterial mechanism of ILDs and proved their potentials for future application. This work clarifies the interactions between molecular sizes of ionic liquid-based species and Gram-negative bacteria and will provide useful guidance for the rational design of high-performance antibacterial agents.

Hereditary Spherocytosis Due to a Novel Variant, P.Q1034X, in the Beta Subunit of the Spectrin Gene

Introduction: Hereditary Spherocytosis (HS) is the most common red cell membrane disorder. 25-30% of cases involve the SPTB gene which encodes for β-Spectrin, a protein that maintains red blood cell shape. Heterozygous variants in SPTB are associated with autosomal dominant HS and elliptocytosis. While genetic testing is not routinely done to confirm HS, it is useful in atypical presentations. Case Description: A 1 week old male presented to the pediatric hematology/oncology clinic for anemia. He was born late preterm and had a history of hyperbilirubinemia requiring phototherapy, failure to thrive, and developmental delay. On examination, he was noted to have hypotonia. There was no known family history of hematologic problems. Based on this constellation of signs and symptoms, he had a comprehensive hematologic and genetic workup. On lab evaluation, his peripheral blood smear showed normocytic normochromic red blood cells with some spherocytes, significant polychromasia, normal WBC and normal platelet morphology. His newborn screen was normal, direct coombs' negative, osmotic fragility test was positive, and protein band 3 reduction was abnormal. His abdominal ultrasound was normal. Whole exome sequencing with variant segregation analysis was significant for heterozygosity of the p.Q1034X variant of the SPTB gene. This variant in the SPTB gene has not been previously reported. Discussion: We found a novel, de novo variant in an infant with HS through whole exome sequencing. This variant is predicted to cause loss of normal protein function either through protein truncation or non-mediated mRNA decay resulting in fragile red blood cells. While neither parent was found to carry this mutation, germline mosaicism should not be excluded. Physicians should be aware that prenatal diagnosis is available to address the risk of recurrence in future pregnancies.

Dehydrated Hereditary Stomatocytosis in a Multi-Generational American Family with a KCNN4 Gardos Channel Mutation

INTRODUCTION: Dehydrated hereditary stomatocytosis (DHSt) is an autosomal dominant red blood cell membrane disorder characterized by hemolytic anemia and splenomegaly. DHSt has an estimated incidence of 1:50,000 births, and the degree of anemia varies within and between families. Although transfusion support during childhood is not uncommon, continued requirement into adulthood is rare. The most frequent cause of DHSt is a gain-of-function mutation of the PIEZO1 gene, leading to delayed channel inactivation that results in a monovalent cation leak and an increase in intracellular calcium (Ca2+). Many of these patients develop recurrent thromboses post splenectomy. Other DHSt patients have mutations in KCNN4, which encodes the Gardos channel, with mutations causing increased Ca2+ sensitivity and potassium efflux. To our knowledge, 42 patients from ten families have been described with four distinct KCNN4 mutations: Arg352His, Val282Met or Val282Glu, and a 28bp deletion encompassing the exon-intron 7 junction. We report herein the eighth family with the Arg352His locus mutation. CASE REPORT: Five subjects from a single family were enrolled in this study (affected proband, unaffected husband, two affected children, and an unaffected grandchild). The proband has had hemolytic anemia since childhood. She had undergone splenectomy at age three and cholecystectomy at eight. Her anemia persisted, with a mean hemoglobin (Hgb) of 10.5g/dL and a reticulocyte count (retic ct) of 12.6%, with no need for transfusion support or iron chelation therapy (Table 1). Extensive testing revealed slightly decreased osmotic fragility and mildly elevated intracellular sodium concentration, of 19.4 mEq/L. Her daughter has mild splenomegaly and anemia, with a mean Hgb of 11.1g/dL and retic ct of 6.7%, whereas her son has more severe disease, with a mean Hgb of 9.3g/dL and a retic ct of &amp;gt;22%. He underwent splenectomy as a teenager for immune thrombocytopenia, and has required chelation therapy with deferasirox since age 31, when his ferritin rose to &amp;gt;1000ng/mL. Similar to results reported by others, splenectomy did not alter the severity of hemolysis in either the proband or her son, and neither developed thrombotic complications 57 and 7 years post splenectomy, respectively. The proband's daughter has a child with a normal Hgb, and is presumed unaffected. Stomatocytes were rarely seen on the peripheral blood smears of the proband and her children. METHODS: A clinically available 39 gene hemolytic anemia panel on the proband failed to identify the mutation underlying this disorder. We therefore performed whole exome sequencing on all five family members. We prioritized the analysis of 23 additional genes that are included in hemolytic anemia panels from two other reference laboratories and are involved in disorders of red blood cell membrane or cytoskeletal proteins of potential clinical relevance to the study population. RESULTS: A single missense mutation, Arg352His within KCNN4, was identified in all three affected individuals. This heterozygous mutation was present in the proband and her affected two children, and absent in her unaffected grandchild and husband. CONCLUSIONS: To our knowledge, this Pennsylvania family is only the eleventh described to have DHSt secondary to a KCNN4 mutation. This disorder is likely much more prevalent than reported, due to the rarity of stomatocytes on peripheral blood smears, the omission of the KCNN4 gene from hemolytic panels offered by some reference laboratories, and variable clinical presentation. KCNN4 mutations should be investigated if other causes are not identified in patients with lifelong hemolytic anemia suspected of having a red cell membrane protein or cytoskeletal disorder. Appropriate diagnosis may allow severely affected patients to be considered for treatment with the experimental Gardos channel inhibitor senicapoc. Additionally, mutational diagnosis is especially important when considering the adverse outcomes post splenectomy in PIEZO1 as compared to KCNN4 mutations. Disclosures Eyster: SPARK:Research Funding;Sanofi:Research Funding;Novo Nordisk:Research Funding;Baxalta/Shire:Research Funding.

Oxidative and nitrosative stress in frozen-thawed pig spermatozoa. II: Effect of the addition of saccharides to freezing medium on sperm function

Saccharides have bioprotective properties, with a high capacity to preserve biological proteins and membranes during sperm cryopreservation. The aim of this study was to evaluate how replacing the lactose of cryopreservation media by sucrose (SUC) or trehalose (TRE) at concentrations of 0.2 M (SUC-1 and TRE-1) and 0.25 M (SUC-2 and TRE-2) affects frozen/thawed pig spermatozoa. The media used were composed of medium A (saccharide/egg yolk) and B (saccharide/egg yolk/glycerol), their osmolality being determined prior to freezing. Cell viability, membrane lipid disorder, acrosome integrity, mitochondrial membrane potential (MMP), lipid peroxidation, thiol group oxidation, total reactive oxygen species (ROS), peroxynitrite and superoxide anion (O2●-) were determined through flow cytometry; total motility (TM), progressive motility (PM) and kinetic parameters motility were determined immediately after thawing (T0) and again 30 (T30) and 60 (T60) minutes later. The SUC-2 and TRE-2 groups maintained viability significantly and presented fewer lipid membrane disorders, respectively, both with a significant increase in MMP. The production of O2●- and peroxynitrite was lower in the TRE-2 groups compared to the control (P < 0.05). Total motility at T0 was greater in the TRE-2 group (P < 0.05). Sperm kinetics was not affected by the treatment. The use of saccharides SUC and TRE at a concentration of 0.25 M improves sperm quality, so that both non-penetrating cryoprotectants can be utilized in pig sperm freezing media.

Synovial chondromatosis: a rare cause of knee pain and swelling

&lt;p&gt;Synovial chondromatosis (SC) is a rare, benign, metaplastic, monoarticular disorder of synovial membrane and bursae of large joints. It commonly affects the large joints such as the knee, hips, wrist, ankle and shoulder. Here we report a case of SC of right knee in a 60-year-old female presenting with chief complaints of pain, swelling, restriction of movement in right knee for 1 year after a fall at home. X-ray of right knee revealed multiple calcified loose bodies. The final diagnosis was established by correlating radiologic findings with the histopathology of the excised specimen. Patient was successfully treated by open partial synovectomy of knee using anterior approaches in a single step procedure.&lt;/p&gt;

Utility of hyperchromic cell percentage, flags, and red cell cytograms generated by Advia-120 hematology analyzer as a potential screening tool in hereditary spherocytosis

Hereditary spherocytosis (HS) is the commonest inherited red cell membrane disorder seen in clinical practice. Modern hematology analyzers generate newer parameters that can help in screening of HS. The present study focuses on the utility of red blood cell data generated by Advia-120 hematology analyzer as a screening tool for patients with HS. Blood samples from 21 patients with HS were analyzed on Advia-120 (Technicon H*1 series) analyzer. The percentage of hyperchromic cells and hyperchromic and hemoglobin concentration variance flags along with the RBC cytogram were studied. A control group of 50 healthy individuals and 50 patients with other hematological disorders was also studied. Osmotic fragility test was done in all patients with suspected HS. All hematological parameters including percentage of hyperchromic red cells were compared between the study group, healthy controls, and other hematological disorders. Twenty of the 21 patients with HS were anemic. Peripheral blood smear in all patients showed variable numbers of spherocytes (5–50%). Osmotic fragility test was positive in all patients. The mean percentage of hyperchromic cells in patients with HS-18.2 ± 13.0% was significantly higher (p 6% showed a sensitivity of 100% and specificity of 99% for the diagnosis of HS. All patients showed flagging for hyperchromic cells. The RBC cytograms in all 21 patients showed significant number of RBCs plotted in the hyperchromic zone. The percentage of hyperchromic cells, hyperchromic cell flagging, and RBC cytogram pattern generated by Advia-120 hematology analyzer are useful tools in the screening of patients with suspected HS.

Telomere-Related Disorders in Fetal Membranes Associated With Birth and Adverse Pregnancy Outcomes.

Telomere disorders have been associated with aging-related diseases, including diabetes, vascular, and neurodegenerative diseases. The main consequence of altered telomere is the induction of the state of irreversible cell cycle arrest. Though several mechanisms responsible for the activation of senescence have been identified, it is still unclear how a cell is indeed induced to become irreversibly arrested. Most tissues in the body will experience senescence throughout its lifespan, but intrinsic and extrinsic stressors, such as chemicals, pollution, oxidative stress (OS), and inflammation accelerate the process. Pregnancy is a state of OS, as the higher metabolic demand of the growing fetus results in increased reactive oxygen species production. As a temporary organ in the mother, senescence in fetal membranes and placenta is expected and linked to term parturition (>37 weeks of gestation). However, a persistent, overwhelming, or premature OS affects placental antioxidant capacity, with consequent accumulation of OS causing damage to lipids, proteins, and DNA in the placental tissues. Therefore, senescence and its main inducer, telomere length (TL) reduction, have been associated with pregnancy complications, including stillbirth, preeclampsia, intrauterine growth restriction, and prematurity. Fetal membranes have a notable role in preterm births, which continue to be a major health issue associated with increased risk of neo and perinatal adverse outcomes and/or predisposition to disease in later life; however, the ability to mediate a delay in parturition during such cases is limited, because the pathophysiology of preterm births and physiological mechanisms of term births are not yet fully elucidated. Here, we review the current knowledge regarding the regulation of telomere-related senescence mechanisms in fetal membranes, highlighting the role of inflammation, methylation, and telomerase activity. Moreover, we present the evidences of TL reduction and senescence in gestational tissues by the time of term parturition. In conclusion, we verified that telomere regulation in fetal membranes requires a more complete understanding, in order to support the development of successful effective interventions of the molecular mechanisms that triggers parturition, including telomere signals, which may vary throughout placental tissues.

Early Dysfunctions in Amyotrophic Lateral Sclerosis: Pathogenetic Mechanisms and the Role in Disease Initiation

Amyotrophic lateral sclerosis (ALS) is a progressive incurable neurodegenerative disease, which is characterized by selective death of the motoneurons of the spinal cord and brain. Currently, ALS is considered as a disease with a wide range of etiological causes, which trigger the cellular and molecular mechanisms leading to the development of similar clinical symptoms associated with degeneration of motoneurons. At the same time, there is no single hypothesis combining various pathogenetic mechanisms, which makes it difficult to develop an effective neuroprotective therapy. This review is devoted to the analysis of the early pathogenetic mechanisms of ALS and their possible role in the initiation of disease on the basis of the published data and our own research. The data on participation of various elements of the motoneuron–skeletal muscle system and membrane disorders in the initiation and development of ALS in humans and in animal models are presented. The analysis of scientific results of the last decades allows us to conclude that a key role in the initiation of pathogenetic mechanisms of ALS is played by dysfunction/destruction of neuromuscular synapses, distal axonopathy, and changes in skeletal muscle metabolism; the methods for correction of these disorders are promising for the development of therapeutic strategies in ALS.

Membrane interactions of non-membrane targeting antibiotics: The case of aminoglycosides, macrolides, and fluoroquinolones

Numerous antibiotics are known to target intracellular pathways, such as protein translation or DNA replication. Membrane transporters typically regulate drug uptake; however, little is known about direct interactions between these antibiotics and the cell membranes. Here, we studied the interactions between different aminoglycosides (kanamycin, gentamicin, streptomycin, neomycin), macrolides (azithromycin, clarithromycin, erythromycin), and fluoroquinolones (ciprofloxacin, levofloxacin) with bacterial membrane mimics to determine drug partitioning and potential drug-induced membrane disruption. The antibiotics' exact location in the bilayers and their effect on membrane thickness and fluidity were determined from high-resolution X-ray diffraction. While the antibiotics did not change membrane thickness at low (1:100 drug/lipid) or high (1:10 drug/lipid) concentrations, they were found to increase membrane disorder in a dose-dependent manner. However, no membrane damage, such as membrane disruption or pore formation, was observed for any of the antibiotics. To note, all antibiotics partitioned into the lipid head groups, while macrolides and fluoroquinolones also partitioned into the bilayer core. The results suggest that the bacterial membrane is relatively inert in the direct mechanisms of actions of these antibiotics.

Southeast Asian Ovalocytosis and Hemoglobinopathies in Newborns: Prevalence, Molecular, and Hematologic Analyses.

Southeast Asian ovalocytosis (SAO) is an inherited red blood cell (RBC) membrane disorder, whereas hemoglobinopathies are inherited globin gene disorders. In an area where both diseases are prevalent, the interaction between them resulting in variable hematologic parameters can be encountered. However, little is known about the genetic interaction of SAO and thalassemia. We investigated the prevalence of SAO and hemoglobinopathy genotypes among newborns in southern Thailand. This study was carried out on 297 newborns recruited consecutively at Naradhiwas Rajanagarindra Hospital in the south of Thailand. The SAO was identified on blood smear examination and polymerase chain reaction analysis. Thalassemia genotypes were defined. Hematologic parameters and hemoglobin (Hb) profiles were recorded and analyzed. Among 297 newborns, 15 (5.1%) carried SAO, whereas 70 (23.6%) had thalassemia with 15 different thalassemia genotypes. Abnormal Hb including Hb C, Hb Q-Thailand, and Hb D-Punjab were observed in 5 newborns. It was found in the nonthalassemic newborns that RBC count, Hb, and hematocrit of the nonthalassemic newborns with SAO were significantly lower than those without SAO. The same finding was also observed in the thalassemic newborns; RBC count, Hb, and hematocrit of the thalassemic newborns with SAO were significantly lower than those without SAO. However, the mean corpuscular volume, mean corpuscular Hb, and RBC distribution width of the SAO-newborns were significantly higher. Both SAO and hemoglobinopathy genotypes are common in southern Thailand. One should take this into consideration when evaluating neonatal anemia and other hematologic abnormalities. Identification of both genetic defects and long-term monitoring on the clinical outcome of this genetic interaction should be essential to understand the pathogenesis of these common genetic disorders in the region.

Relationship between red blood cell membrane disorders and Doppler ultrasound indices of renal blood flow in patients with chronic kidney disease

the study aimed to investigate the relationship between red blood cell membrane disorders and Doppler ultrasound indices of renal blood flow in patients with chronic kidney disease (CKD) stages1-2.&#x0D; Methods. The complex clinical, laboratory and instrumental (color duplex Doppler scanning of renal parenchyma vessels) examination was conducted in 131 CKD stage 1-2 patients. Among them, there were 47 patients with pyelonephritis, 48 patients with diabetic nephropathy, and 36 patients with primary glomerulonephritis. Each patients’ group was divided into 2 subgroups: with and without arterial hypertension. Twenty healthy volunteers were also included for the study control.&#x0D; Results. A significant correlation between indices of the erythrocytes morphology and basic Doppler ultrasonography parameters of the kidney vessels circulation, namely a.interlobaris was demonstrated in CKD patients with arterial hypertension. The most significant changes were observed in diabetic patients.&#x0D; Conclusion. The results of our study indicated a high informative value of renal Doppler ultrasonography as a non-invasive technique to diagnose the CKD progression in patients with comorbid hypertension

Recommendations for Pregnancy in Rare Inherited Anemias

Rare inherited anemias are a subset of anemias caused by a genetic defect along one of the several stages of erythropoiesis or in different cellular components that affect red blood cell integrity, and thus its lifespan. Due to their low prevalence, several complications on growth and development, and multi-organ system damage are not yet well defined. Moreover, during the last decade there has been a lack of proper understanding of the impact of rare anemias on maternal and fetal outcomes. In addition, there are no clear-cut guidelines outlining the pathophysiological trends and management options unique to this special population. Here, we present on behalf of the European Hematology Association, evidence- and consensus-based guidelines, established by an international group of experts in different fields, including hematologists, gynecologists, general practitioners, medical geneticists, and experts in rare inherited anemias from various European countries for standardized and appropriate choice of therapeutic interventions for the management of pregnancy in rare inherited anemias, including Diamond-Blackfan Anemia, Congenital Dyserythropoietic Anemias, Thalassemia, Sickle Cell Disease, Enzyme deficiency and Red cell membrane disorders.

Irradiating frozen-thawed stallion sperm with red-light increases their resilience to withstand post-thaw incubation at 38 °C

The aim of this study was to evaluate whether red-light stimulation increases the longevity and resilience of cryopreserved stallion sperm to withstand post-thaw incubation for 120 min. Sixteen frozen straws of 0.5 mL from eight stallions were used. Samples were cryopreserved, thawed through incubation at 38 °C for 30 s and divided into the control and samples exposed to red-light using a triple LED photo-activation system (wavelength: 620–630 nm). Three irradiation protocols consisting of different light-dark-light intervals (1-1-1, 2-2-2 and 3-3-3 min) were tested. Sperm quality parameters were analyzed immediately after light-stimulation (0 min) and after 120 min of incubation at 38 °C. Sperm motility was evaluated using a Computerized Semen Analysis System (CASA), and flow cytometry and different fluorochromes were used to evaluate the integrity and lipid disorder of plasma membrane, mitochondrial membrane potential and intracellular levels of peroxides and superoxides. Irradiation significantly increased the percentages of spermatozoa with high mitochondrial membrane potential (1-1-1 pattern) and the intracellular levels of peroxides (2-2-2 pattern) at 0 min. In addition, sperm kinematic parameters (2-2-2 and 3-3-3 patterns) and percentages of viable spermatozoa with low membrane lipid disorder (3-3-3 pattern) were significantly higher in irradiated samples than in the control at 120 min. Our results indicate that red-light stimulation could help increase the resilience of frozen-thawed stallion sperm to withstand post-thaw incubation at 38 °C for 120 min and that these effects rely on the irradiation pattern. Further research should evaluate whether light-stimulation could also have a positive on fertility rates after artificial insemination.

Progress toward Alzheimer's disease treatment: Leveraging the Achilles' heel of Aβ oligomers?

After three decades of false hopes and failures, a pipeline of therapeutic drugs that target the actual root cause of Alzheimer's disease (AD) is now available. Challenging the old paradigm that focused on β-amyloid peptide (Aβ) aggregation in amyloid plaques, these compounds are designed to prevent the neurotoxicity of Aβ oligomers that form Ca2+ permeable pores in the membranes of brain cells. By triggering an intracellular Ca2+ overdose, Aβ oligomers induce a cascade of neurotoxic events including oxidative stress, tau hyperphosphorylation, and neuronal loss. Targeting any post-Ca2+ entry steps (e.g., tau) will not address the root cause of the disease. Thus, preventing Aβ oligomers formation and/or blocking their toxicity is by essence the best approach to stop any progression of AD. Three categories of anti-oligomer compounds are already available: antibodies, synthetic peptides, and small drugs. Independent in silico-based designs of a peptide (AmyP53) and a monoclonal antibody (PMN310) converged to identify a histidine motif (H13/H14) that is critical for oligomer neutralization. This "histidine trick" can be viewed as the Achilles' heel of Aβ in the fight against AD. Moreover, lipid rafts and especially gangliosides play a critical role in the formation and toxicity of Aβ oligomers. Recognizing AD as a membrane disorder and gangliosides as the key anti-oligomer targets will provide innovative opportunities to find an efficient cure. A "full efficient" solution would also need to be affordable to anyone, as the number of patients has been following an exponential increase, affecting every part of the globe.

Clinical Diagnosis of Red Cell Membrane Disorders: Comparison of Osmotic Gradient Ektacytometry and Eosin Maleimide (EMA) Fluorescence Test for Red Cell Band 3 (AE1, SLC4A1) Content for Clinical Diagnosis.

The measurement of band 3 (AE1, SLC4A1, CD233) content of red cells by eosin-5- maleimide (EMA) staining is swiftly replacing conventional osmotic fragility (OF) test as a tool for laboratory confirmation of hereditary spherocytosis across the globe. Our group has systematically evaluated the EMA test as a method to screen for a variety of anemias in the last 10 years, and compared these results to those obtained with the osmotic gradient ektacytometry (osmoscans) which we have used over three decades. Our overall experience allowed us to characterize the distinctive patterns with the two tests in several congenital erythrocyte membrane disorders, such as hereditary spherocytosis (HS), hereditary elliptocytosis (HE), Southeast Asian Ovalocytosis (SAO), hereditary pyropoikilocytosis (HPP) variants, erythrocyte volume disorders, various red cell enzymopathies, and hemoglobinopathies. A crucial difference between the two methodologies is that osmoscans measure red blood cell deformability of the entire sample of RBCs, while the EMA test examines the band 3 content of individual RBCs. EMA content is influenced by cell size as smaller red cells have lower amount of total membrane than larger cells. The SAO mutation alters the EMA binding site resulting in a lower EMA MCF even as the band 3 content itself is unchanged. Thus, EMA scan results should be interpreted with caution and both the histograms and dot plots should be analyzed in the context of the clinical picture and morphology.