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The study aimed to evaluate how plasma total proteins, their fractions (i.e. albumin, α1-, β-, γ-globulins) and A/G ratio change in lambs and kids over the first month of life. Ten female Maltese kids and 10 female Comisana lambs were enrolled in the study. Starting from the first day after birth, and for a period of 4 weeks, blood samples were taken from each subject. Specifically, for each kid and lamb, blood sampling was performed within 30 min after birth (T0) (pre-colostral), after colostrum intake (T1), and, weekly, for the subsequent 28 days (T7, T14, T21 and T28). The application of two-way analysis of variance for repeated measures (ANOVA) showed the effect of time in the kids and lambs for the values of plasma total proteins, albumin, α1-, β-, γ-globulins and A/G ratio (p < 0.05). A significant species effect was recorded for α1-and β-globulins (p < 0.05). Specifically, kids showed higher α1-globulins levels at T0, T14, T21 and T28, and higher and β-globulins at T21 and T28 than lambs. The variations found in the levels of plasma total proteins and their fractions in kids and lambs could be related to the physiological changes likely to occur in a newborn animal following transition from the uterine environment to the free-living state. The variability on changes found in α1-and β-globulins measured in kid and lamb seems to suggest that the first one would tend to have a higher acute phase response than lamb stressing the difference between these two small ruminant species., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2024 The Authors. Published by Elsevier Ltd.)

Inner ear disorders, including sensorineural hearing loss, Meniere's disease, and vestibular neuritis, are prevalent conditions that significantly impact the quality of life. Despite their high incidence, the underlying pathophysiology of these disorders remains elusive, and current treatment options are often inadequate. Emerging evidence suggests that pericytes, a type of vascular mural cell specialized to maintain the integrity and function of the microvasculature, may play a crucial role in the development and progression of inner ear disorders. The pericytes are present in the microvasculature of both the cochlea and the vestibular system, where they regulate blood flow, maintain the blood-labyrinth barrier, facilitate angiogenesis, and provide trophic support to neurons. Understanding their role in inner ear disorders may provide valuable insights into the pathophysiology of these conditions and lead to the development of novel diagnostic and therapeutic strategies, improving the standard of living. This comprehensive review aims to provide a detailed overview of the role of pericytes in inner ear disorders, highlighting the anatomy and physiology in the microvasculature, and analyzing the mechanisms that contribute to the development of the disorders. Furthermore, we explore the potential pericyte-targeted therapies, including antioxidant, anti-inflammatory, and angiogenic approaches, as well as gene therapy strategies.

Background/Objectives : Rare diseases are a wide and heterogeneous group of multisystem life-threatening or chronically debilitating clinical conditions with reduced life expectancy and a relevant mortality rate in childhood. Some of these disorders have typical neurological symptoms, presenting from birth to adulthood. Dietary patterns and nutritional compounds play key roles in the onset and progression of neurological disorders, and the impact of alimentary needs must be enlightened especially in rare neurological diseases. This work aims to collect the in vitro , in vivo , and clinical evidence on the effects of diet and of nutrient intake on some rare neurological disorders, including some genetic diseases, and rare brain tumors. Herein, those aspects are critically linked to the genetic, biological, biochemical, and pathophysiological hallmarks typical of each disorder. Methods : By searching the major web-based databases (PubMed, Web of Science Core Collection, DynaMed, and Clinicaltrials.gov), we try to sum up and improve our understanding of the emerging role of nutrition as both first-line therapy and risk factors in rare neurological diseases. Results : In line with the increasing number of consensus opinions suggesting that nutrients should receive the same attention as pharmacological treatments, the results of this work pointed out that a standard dietary recommendation in a specific rare disease is often limited by the heterogeneity of occurrent genetic mutations and by the variability of pathophysiological manifestation. Conclusions : In conclusion, we hope that the knowledge gaps identified here may inspire further research for a better evaluation of molecular mechanisms and long-term effects.

In recent years, the presence of pharmaceuticals and microplastics (MPs) in aquatic ecosystems has raised concerns about their environmental impact. This study explores the combined effects of caffeine, a common pharmaceutical pollutant, and MPs on the marine mussel Mytilus galloprovincialis. Caffeine, at concentrations of 20.0 μg L -1 , and MPs (1 mg L -1 , 35-50 μm size range), was used to mimic real-world exposure scenarios. Two hundred M. galloprovincialis specimens were divided into four groups: caffeine, MPs, Mix (caffeine + MPs), and Control. After a two-week acclimation period, the mollusks were subjected to these pollutants in oxygen-aerated aquariums under controlled conditions for 14 days. Histopathological assessments were performed to evaluate gill morphology. Cellular volume regulation and digestive gland cell viability were also analyzed. Exposure to caffeine and MPs induced significant morphological changes in M. galloprovincialis gills, including cilia loss, ciliary disk damage, and cellular alterations. The chitinous rod supporting filaments also suffered damage, potentially due to MP interactions, leading to hemocyte infiltration and filament integrity compromise. Hemocytic aggregation suggested an inflammatory response to caffeine. In addition, viability assessments of digestive gland cells revealed potential damage to cell membranes and function, with impaired cell volume regulation, particularly in the Mix group, raising concerns about nutrient metabolism disruption and organ function compromise. These findings underscore the vulnerability of M. galloprovincialis to environmental pollutants and emphasize the need for monitoring and mitigation efforts. RESEARCH HIGHLIGHTS: The synergy of caffeine and microplastics (MPs) in aquatic ecosystems warrants investigation. MPs and caffeine could affect gill morphology of Mytilus galloprovincialis. Caffeine-exposed cells had lower viability than the control group in the NR retention test. MPs and mix-exposed cells struggled to recover their volume., (© 2024 The Authors. Microscopy Research and Technique published by Wiley Periodicals LLC.)

Specific subpopulations of neurons in nerve and sensory systems must be developed and maintained, and this is accomplished in significant part by neurotrophins (NTs) and the signaling receptors on which they act, called tyrosine protein kinase receptors (Trks). The neurotrophins-tyrosine protein kinase receptors (NTs/Trks) system is involved in sensory organ regulation, including the visual system. An NTs/Trks system alteration is associated with neurodegeneration related to aging and diseases, including retinal pathologies. An emergent model in the field of translational medicine, for instance, in aging study, is the annual killifish belonging to the Nothobranchius genus, thanks to its short lifespan. Members of this genus, such as Nothobranchius guentheri , and humans share a similar retinal stratigraphy. Nevertheless, according to the authors' knowledge, the occurrence and distribution of the NTs/Trks system in the retina of N. guentheri has never been investigated before. Therefore, the present study aimed to localize neurotrophin BDNF, NGF, and NT-3 and TrkA, TrkB, and TrkC receptors in the N. guentheri retina using the immunofluorescence method. The present investigation demonstrates, for the first time, the occurrence of the NTs/Trks system in N. guentheri retina and, consequently, the potential key role of these proteins in the biology and survival of the retinal cells.

The gilthead seabream, one of the most important species in Mediterranean aquaculture, with an increasing status of exploitation in terms of production volume and aquafarming technologies, has become an important research topic over the years. The accumulation of knowledge from several studies conducted during recent decades on their functional and biological characteristics has significantly improved their aquacultural aspects, namely their reproductive success, survival, and growth. Despite the remarkable progress in the aquaculture industry, hatchery conditions are still far from ideal, resulting in frequent abnormalities at the beginning of intensive culture, entailing significant economic losses. Those deformities are induced during the embryonic and post-embryonic periods of life, and their development is still poorly understood. In the present review, we created a comprehensive synthesis that covers the various aspects of skeletal morphogenesis and anomalies in the gilthead seabream, highlighting the genetic, environmental, and nutritional factors contributing to bone deformities and emphasized the potential of the gilthead seabream as a model organism for understanding bone morphogenesis in both aquaculture and translational biological research. This review article addresses the existing lack in the literature regarding gilthead seabream bone deformities, as there are currently no comprehensive reviews on this subject.

The morphology of the oral cavity of fish is related to their feeding habits. In this context, taste buds are studied for their ability to catch chemical stimuli and their cell renewal capacity. Vimentin RV202 is a protein employed as a marker for mesenchymal cells that can differentiate along different lineages and to self-renew, while Calretinin N-18 is employed as a marker of sensory cells, and ubiquitin is a protein crucial for guiding the fate of stem cells throughout development. In this study, a surface morphology investigation and an immunohistochemical analysis have been conducted. The results of the present study reveal, for the first time, the presence of Vimentin RV202 in a taste bud cell population of zebrafish. Some taste bud cells are just Vimentin RV202-immunoreactive, while in other cells Vimentin RV202 and Calretinin N-18 colocalize. Some taste buds are just reactive to Calretinin N-18. Vimentin RV202-immunoreactive cells have been observed in the connective layer and in the basal portion of the taste buds. The immunoreactivity of ubiquitin was restricted to sensory cells. Further studies are needed to elucidate the role of Vimentin RV202 in the maturation of taste bud cells, its potential involvement in the regeneration of these chemosensory organs, and its eventual synergic work with ubiquitin.

Calcium-binding proteins (CaBPs) are members of a heterogeneous family of proteins able to buffer intracellular Ca 2+ ion concentration. CaBPs are expressed in the central and peripheral nervous system, including a subpopulation of retinal neurons. Since neurons expressing different CaBPs show different susceptibility to degeneration, it could be hypothesized that they are not just markers of different neuronal subpopulations, but that they might be crucial in survival. CaBPs' ability to buffer Ca 2+ cytoplasmatic concentration makes them able to defend against a toxic increase in intracellular calcium that can lead to neurodegenerative processes, including those related to aging. An emergent model for aging studies is the annual killifish belonging to the Nothobranchius genus, thanks to its short lifespan. Members of this genus, such as Nothobranchius guentheri , show a retinal stratigraphy similar to that of other actinopterygian fishes and humans. However, according to our knowledge, CaBPs' occurrence and distribution in the retina of N. guentheri have never been investigated before. Therefore, the present study aimed to localize Calretinin N-18, Parvalbumin, and S100 protein (S100p) in the N. guentheri retina with immunohistochemistry methods. The results of the present investigation demonstrate for the first time the occurrence of Calretinin N-18, Parvalbumin, and S100p in N. guentheri retina and, consequently, the potential key role of these CaBPs in the biology of the retinal cells. Hence, the suitability of N. guentheri as a model to study the changes in CaBPs' expression patterns during neurodegenerative processes affecting the retina related both to disease and aging can be assumed.

Background: Anorexia of aging, defined as a decrease in appetite and a preponderant loss of body weight occurring in late life, is one of the most common diseases affecting older people. The peptide hormone cholecystokinin (Cck) is known to play a key role in regulating food intake and satiety in higher vertebrates. In humans as well as in rats, an increased concentration of Cck was described as the basis of appetite loss in elderly. However, the role of increased plasma Cck concentrations in mediating the age-related decrease in appetite remains to be established. Although in vitro studies are an excellent resource for investigating aging, the use of a model organism that shares and imitates the human physiological processes guarantees a better understanding of the in vivo mechanisms. African annual fishes from the genus Nothobranchius are emerging as a prominent model organism in biogerontology and developmental biology due to their short captive lifespan. Therefore, in the current study, we aimed to investigate the possibility of using the genus Nothobranchius to model the anorexia of aging and their potential contribution to better understanding the pathway by which Cck induce appetite loss in older people providing a comparative/evolutionary localization of the current study model among the aging canonicals models, the morphology of its gastrointestinal tract and its Cck expression pattern., Methods: The comparative/evolutionary investigation was conducted using the NCBI blastp (protein-protein BLAST) and NCBI Tree Viewer. The macroscopic morphology, histological features, ultrastructural organization of Nothobranchius rachovii gastrointestinal tract were investigated using stereomicroscope, Masson's trichrome and alcian blue-PAS staining, and transmission electron microscopy, respectively. The cck expression pattern was studied through immunofluorescence labeling, western blotting, and quantitative RT-PCR., Results: The intestine was folded into different segments divided into an anterior intestine made of a rostral intestinal bulb and an intestinal annex of lower diameter, mid and posterior intestine. The gradual transition from the rostral intestinal bulb to the posterior intestine sections's epithelium is characterized by a gradual reduction in the striated muscular bundles, villi height, and goblet mucous cells count. The lining epithelium of the intestinal villi was characterized by a typical brush border enterocytes full of mitochondria. Moreover, Cck expression was detected in scattered intraepithelial cells concentrated in the anterior tract of the intestine., Conclusions: Our study introduces Nothobranchius rachovii as a model for anorexia of aging, giving the first bases on the gastrointestinal tract morphology and cck expression pattern. Future studies on young and elderly Notobranchius can divulge the contribution of cck in the mechanisms of anorexia associated with aging., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier GmbH.. All rights reserved.)

The incidence rates of light-induced retinopathies have increased significantly in the last decades because of continuous exposure to light from different electronic devices. Recent studies showed that exposure to blue light had been related to the pathogenesis of light-induced retinopathies. However, the pathophysiological mechanisms underlying changes induced by light exposure are not fully known yet. In the present study, the effects of exposure to light at different wavelengths with emission peaks in the blue light range (400-500 nm) on the localization of Calretinin-N18 (CaR-N18) and Calbindin-D28K (CaB-D28K) in adult zebrafish retina are studied using double immunofluorescence with confocal laser microscopy. CaB-D28K and CaR-N18 are two homologous cytosolic calcium-binding proteins (CaBPs) implicated in essential process regulation in central and peripheral nervous systems. CaB-D28K and CaR-N18 distributions are investigated to elucidate their potential role in maintaining retinal homeostasis under distinct light conditions and darkness. The results showed that light influences CaB-D28K and CaR-N18 distribution in the retina of adult zebrafish, suggesting that these CaBPs could be involved in the pathophysiology of retinal damage induced by the short-wavelength visible light spectrum.

In the framework of investigations aimed to detect new available bioindicators in marine environment, haemolymph cells and ctenidia of the Mediterranean spiny oyster, Spondylus gaederopus, have been investigated. Haemocyte count and characterisation, phagocytosis and superoxide anion production and enzyme activity assays, have been carried out. TEM observations have been performed. After gross anatomy observations, cito-histological determinations have been carried out, especially focused on ctenidia structure and function. Main results concerned the relatively low number of circulating cells, and the rich in granules granulocytes, most of which were lysosomes. Release of lysosomal enzymes was confirmed a shared trait inside bivalves. Glycogen deposits as probable result of conversion of bacteria carbohydrates, have been detected, as well as the occurrence of both acidophilic and basophilic haemocytes. Phagocytosis, both in granulocytes and agranulocytes, has been recorded, together with the production of superoxide anion. Haemocytes were found positive to acid phosphatase, alkaline phosphatase, β-glucuronidase, chloroacetylesterase and arylsulphatase. Ctenidia showed a complex organization, including two demibranch to each ctenidium, two different kinds of lamellae filament and specialized structures as ciliated disks connecting filaments in "eutherorhabdic ctenidia". The occurrence of three different types of mucous cells in the same region of ordinary filaments has been underlined. Such features, suggesting high resistance to environmental stress and disease, allow to consider spiny oysters as promising bioindicators, although deserving of further investigations to evaluate the physiological responses to stress in controlled conditions. Present data, moreover, providing basic information on the biology of S. gaederopus, notably implement the present knowledge on the Mediterranean spiny oysters, whose under-evaluated ecological role should be carefully considered., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

A morphological study of the alimentary tract, from the oropharyngeal cavity to the rectum, including the attached glands, of African bony-tongue, Heterotis niloticus (Cuvier, 1829) was carried out by gross anatomy, and light microscope analysis. This study aimed to give a deeper knowledge of the alimentary tract morphological features of this species of commercial interest. H. niloticus is distinguished by individual morphological characteristics showing a digestive tract similar to that of reptiles and birds. Within the oropharyngeal cavity, two tubular structures with digitiform ends are arranged on both lateral sides of the triangular tongue. The oropharyngeal cavity connects the stomach by a short esophagus. This latter is adapted to mechanical trituration, and it is divided into a pars glandularis and a thick-walled pars muscularis. The gizzard flows into the anterior intestine and two blind pyloric appendages, which exhibit specific functions, including immune defense for the presence of secondary lymphoid organs. The anterior intestine continues with the middle and posterior tracts up into the rectum. According to the histological observations, all regions of the alimentary tract have common structural features, typical of hollow organs, with differences in the mucosa structure that reflects the different functions of the apparatus, from mouth to anus. Within this study, we provided the first basis for future studies on optimizing rearing conditions, feed conversion ratio, and the digestive capacity, improving the growth performance of this species, and ensuring its conservation.

Obesity is a pathological condition, defined as an excessive accumulation of fat, primarily caused by an energy imbalance. The storage of excess energy in the form of triglycerides within the adipocyte leads to lipotoxicity and promotes the phenotypic switch in the M1/M2 macrophage. These changes induce the development of a chronic state of low-grade inflammation, subsequently generating obesity-related complications, commonly known as metabolic syndromes. Over the past decade, obesity has been studied in many animal models. However, due to its competitive aspects and unique characteristics, the use of zebrafish has begun to gain traction in experimental obesity research. To counteract obesity and its related comorbidities, several natural substances have been studied. One of those natural substances reported to have substantial biological effects on obesity are flavonoids. This review summarizes the results of studies that examined the effects of flavonoids on obesity and related diseases and the emergence of zebrafish as a model of diet-induced obesity.

Prior to senescence, erythrocytes may experience injury, which compromises their integrity and thus triggers suicidal erythrocyte death or eryptosis. This mechanism is characterised by cell shrinkage, cell membrane blebbing, and cell membrane phospholipid scrambling after phosphatidylserine exposure on the cell surface that is identified by macrophages, which engulf and degrade the eryptotic cells. The term eryptosis also includes typical mechanisms, which contribute to the triggering of this process, such as oxidative stress, Ca2+ entry with an increase in cytosolic Ca2+ activity ([Ca ]i) and the activation of p38 kinase, which is a kinase expressed in human erythrocytes and activated after hyperosmotic shock. Enhanced eryptosis has been observed in several clinical conditions such as diabetes, renal insufficiency, haemolytic uremic syndrome, sepsis, mycoplasma infection, malaria, iron deficiency, sickle cell anaemia, beta-thalassemia, glucose-6-phosphate dehydrogenase-(G6PD) deficiency, hereditary spherocytosis, paroxysmal nocturnal haemoglobinuria, Wilson's disease, myelodysplastic syndrome, and phosphate depletion. Therefore, eryptosis may be considered as a useful mechanism of removal of defective erythrocytes to prevent haemolysis. Moreover, the clearance of infected erythrocytes in diseases such as malaria may counteract parasitemia. Indeed it is known that sickle-cell trait, beta-thalassemia trait, glucose-6-phosphate dehydrogenase (G6PD)- deficiency and iron deficiency confer some protection against a severe course of malaria. Importantly, strategies to control Plasmodium infection by inducing eryptosis are not expected to generate resistance of the pathogen, as the proteins involved in suicidal death of the host cell are not encoded by the pathogen and thus cannot be modified by mutations of its genes. However, excessive eryptosis could compromise microcirculation and lead to anemia., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Nutritional abnormalities and physical inactivity are risk factors of increased morbidity and mortality in patients with ESRD. Identify and define malnutrition, in particular protein-energy depletion (PEW), is an important task in the management of renal patients. The aim of this multicenter observational study was to implement the assessment of nutritional status and functional capacity in patients on peritoneal dialysis, including tests and validated methods which are relatively easy to apply in daily clinical practice. The study includes all the 133 prevalent patients (80 m, 53 f, age 65 14 years), in peritoneal dialysis treatment (vintage 26 19 months) in 9 centers in Tuscany. We performed anthropometry, bioimpedance (BIA), clinical biochemistry, evaluation of habitual physical activity (RAPA tests) and performance (Sit-To-Stand test), appetite-evaluation questionnaire, and indices including the Malnutrition Inflammation Score (MIS), Geriatric Nutrition Risk Index (GNRI), Charlson comorbidity index, Barthel and Karnowsky index. The latter showed a condition of dependence in 7.2% and 19.7% of cases, respectively. Poor appetite was recorded in 48.2%. The majority of patients fell within the overweight / obesity range (51%) with waist circumference values associated with increased cardiovascular risk in 51% of males and 60% of females. At the BIA analysis, a BCMI <8 kg/m2 was detected in 39% of patients; an estimated protein intake <1.0 g / kg/d was found in 59% of cases; 34% of patients had serum albumin <3.5 g / dl; control of acidosis was good (bicarbonate 25.4 3.8 mM) but hyperphosphatemia was present in 64.6% of patients. A condition of sedentary or light physical activity was reported by 65.1% of patients, vigorous activity only by 11.9%. The 86.5% of patients able to perform the Sit-to-stand test reported a lower than the reference values for age and sex. A diagnosis of PEW was possible in 8% of our series, while a MIS score> 11, indicative of PEW, took place in 12.7% of cases. The values of the MIS correlated directly with age and the degree of comorbidity and inversely with the sit-to-stand test, RAPA tests and appetite level. The data in this study show that single tests indicative of malnutrition disorders are frequent to be found in our series of peritoneal dialysis patients. However, a diagnosis of PEW is quite infrequent. A large percentage of patients are overweight with increased abdominal adiposity, and reduced cell mass and protein intake below recommended levels; the level of habitual physical activity is low, and the level of physical capability is scarce. Therefore it is conceivable a nutritional counseling intervention to increase the intake of proteins, limiting the phosphorus and (when indicated) energy intake and to stimulating spontaneous physical activity or arranging assisted programs for functional rehabilitation. Close monitoring of the nutritional status and implementation of programs of adapted physical activity should have a prominent role in the clinical management of patients on peritoneal dialysis.

Background: The microtubule assembly inhibitor nocodazole has been shown to trigger caspase-independent mitotic death and caspase dependent apoptosis. Similar to apoptosis of nucleated cells, erythrocytes may undergo eryptosis, the suicidal erythrocyte death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Stimulators of eryptosis include increase of cytosolic Ca2+ activity ([Ca2+]i), oxidative stress and ceramide. The present study explored, whether and how nocodazole induces eryptosis., Methods: Flow cytometry was employed to determine phosphatidylserine exposure at the cell surface from annexin-V-binding, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, the abundance of reactive oxygen species (ROS) from 2',7'-dichlorodihydrofluorescein (DCF) diacetate dependent fluorescence as well as ceramide surface abundance utilizing specific antibodies. Tubulin abundance was quantified by TubulinTracker™ Green reagent and visualized by confocal microscopy., Results: A 48 hours exposure of human erythrocytes to nocodazole (≥ 30 µg/ml) significantly increased the percentage of annexin-V-binding cells without significantly modifying average forward scatter. Nocodazole significantly increased Fluo3-fluorescence, significantly increased DCF fluorescence and significantly increased ceramide surface abundance. The effect of nocodazole on annexin-V-binding was significantly blunted, but not abolished by removal of extracellular Ca2+ and was not modified in the presence of Caspase 3 inhibitor zVAD (1 µM). Nocodazole treatment reduced the content of total tubulin., Conclusions: Nocodazole triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part due to stimulation of Ca2+ entry, oxidative stress and ceramide., (© 2016 The Author(s) Published by S. Karger AG, Basel.)

The natural phosphoprotein phosphatase inhibitor cantharidin, primarily used for topical treatment of warts, has later been shown to trigger tumor cell apoptosis and is thus considered for the treatment of malignancy. Similar to apoptosis of tumor cells, erythrocytes may undergo eryptosis, a suicidal cell death characterized by cell shrinkage and translocation of cell membrane phosphatidylserine to the erythrocyte surface. Signaling of eryptosis includes increase of cytosolic Ca2+-activity ([Ca2+]i), ceramide, oxidative stress and dysregulation of several kinases. Phosphatidylserine abundance at the erythrocyte surface was quantified utilizing annexin-V-binding, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, ceramide from antibody binding, and reactive oxidant species (ROS) from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence. A 48 h treatment of human erythrocytes with cantharidin significantly increased the percentage of annexin-V-binding cells (≥10 mg/mL), significantly decreased forward scatter (≥25 mg/mL), significantly increased [Ca2+]i (≥25 mg/mL), but did not significantly modify ceramide abundance or ROS. The up-regulation of annexin-V-binding following cantharidin treatment was not significantly blunted by removal of extracellular Ca2+ but was abolished by kinase inhibitor staurosporine (1 mM) and slightly decreased by p38 inhibitor skepinone (2 mM). Exposure of erythrocytes to cantharidin triggers suicidal erythrocyte death with erythrocyte shrinkage and erythrocyte membrane scrambling, an effect sensitive to kinase inhibitors staurosporine and skepinone.

Background/aims: Fucoxanthin, a carotenoid isolated from brown seaweeds, induces suicidal death or apoptosis of tumor cells and is thus considered for the treatment or prevention of malignancy. In analogy to apoptosis of nucleated cell, erythrocytes may enter eryptosis, the suicidal death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i), oxidative stress and activation of p38 kinase or protein kinase C. The present study explored, whether and how fucoxanthin induces eryptosis., Methods: Phosphatidylserine exposure at the cell surface was estimated from annexin-V-binding, cell volume from forward scatter, hemolysis from hemoglobin release, [Ca2+]i from Fluo3-fluorescence, and abundance of reactive oxygen species (ROS) from DCFDA dependent fluorescence and lipid peroxidation using BODIPY fluoresence., Results: A 48 hours exposure of human erythrocytes to fucoxanthin significantly increased the percentage of annexin-V-binding cells (≥ 50 µM), significantly decreased average forward scatter (≥ 25 µM), significantly increased hemolysis (≥ 25 µM), significantly increased Fluo3-fluorescence (≥ 50 µM), significantly increased lipid peroxidation, but did not significantly modify DCFDA fluorescence. The effect of fucoxanthin on annexin-V-binding was significantly blunted, but not abolished by removal of extracellular Ca2+, and was insensitive to p38 kinase inhibitor skepinone (2 µM) and to protein kinase C inhibitor calphostin (100 nM)., Conclusion: Fucoxanthin triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part due to stimulation of Ca2+ entry., (© 2015 The Author(s) Published by S. Karger AG, Basel.)

Background/aims: The alkylating drug oxaliplatin is widely used for chemotherapy of malignancy. Oxaliplatin is effective by inducing both, necrosis and apoptosis. Similar to necrosis or apoptosis of nucleated cells, erythrocytes may enter hemolysis, which is apparent from hemoglobin release or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include oxidative stress and/or Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i). The present study explored, whether and how oxaliplatin induces eryptosis., Methods: Phosphatidylserine exposure at the cell surface was quantified utilizing annexin-V-binding, cell volume estimated from forward scatter, hemolysis deduced from hemoglobin release, [Ca2+]i determined utilizing Fluo-3 fluorescence, and reactive oxygen species (ROS) abundance visualized using 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) dependent fluorescence., Results: A 48 hours exposure of human erythrocytes to oxaliplatin (10 µg/ml) significantly increased the percentage of annexin-V-binding cells, significantly decreased forward scatter, significantly increased Fluo-3 fluorescence, and significantly increased DCFDA fluorescence. The effect of oxaliplatin on annexin-V-binding and forward scatter was rather augmented by removal of extracellular Ca2+, but was significantly blunted in the presence of the antioxidant N-acetyl-cysteine (1 mM)., Conclusions: Oxaliplatin triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect partially dependent on ROS formation., (© 2015 The Author(s) Published by S. Karger AG, Basel.)

Background/aims: The benzophenone garcinol from dried fruit rind of Garcinia indica counteracts malignancy, an effect at least in part due to stimulation of apoptosis. The proapototic effect of garcinol is attributed in part to inhibition of histone acetyltransferases and thus modification of gene expression. Moreover, garcinol triggers mitochondrial depolarisation. Erythrocytes lack gene expression and mitochondria but are nevertheless able to enter apoptosis-like suicidal death or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Stimulators of eryptosis include oxidative stress, energy depletion and Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i). The present study explored, whether and how garcinol induces eryptosis., Methods: To this end, phosphatidylserine exposure at the cell surface was estimated from annexin-V-binding, cell volume from forward scatter, hemolysis from hemoglobin release, [Ca2+]i from Fluo3-fluorescence, ROS formation from DCFDA dependent fluorescence and cytosolic ATP levels utilizing a luciferin-luciferase-based assay., Results: A 24 hours exposure of human erythrocytes to garcinol (2.5 or 5 µM) significantly increased the percentage of annexin-V-binding cells. Garcinol decreased (at 1 µM and 2.5 µM) or increased (at 5 µM) forward scatter. Garcinol (5 µM) further increased Fluo3-fluorescence, increased DCFDA fluorescence, and decreased cytosolic ATP levels. The effect of garcinol on annexin-V-binding was significantly blunted, but not abolished by removal of extracellular Ca2+., Conclusions: Garcinol triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part due to stimulation of ROS formation, energy depletion and Ca2+ entry., (© 2015 The Author(s) Published by S. Karger AG, Basel.)

Background: The P-glycoprotein inhibitor zosuquidar (LY335979) is clinically used to augment the effect of cytostatic drugs on suicidal tumor cell death or apoptosis. The present study explored whether the substance is cytotoxic to erythrocytes. Upon injury, erythrocytes may undergo suicidal cell death or eryptosis, which is characterized by cell shrinkage and translocation of cell membrane phosphatidylserine to the erythrocyte surface. Signaling of eryptosis include increase of cytosolic Ca2+-activity ([Ca2+]i), oxidative stress and activation of several kinases, such as p38 kinase and protein kinase C., Methods: Phosphatidylserine abundance at the erythrocyte surface was quantified from binding of FITC-labelled annexin-V, cell volume from forward scatter, [Ca2+]i from Fluo3-fluorescence, and reactive oxygen species (ROS) from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence., Results: A 48 h treatment of human erythrocytes with zosuquidar significantly increased the percentage of annexin-V-binding cells (2 and 4 µg/ml), significantly decreased forward scatter (4 µg/ml), significantly increased [Ca2+]i (4 µg/ml), but did not significantly modify ROS. The up-regulation of annexin-V-binding following zosuquidar (4 µg/ml) treatment was significantly blunted by removal of extracellular Ca2+, by presence of p38 kinase inhibitor SB203580 (2 µM) and by presence of protein kinase C inhibitor calphostin (100 nM)., Conclusions: Exposure of erythrocytes to zosuquidar triggers suicidal erythrocyte death with erythrocyte shrinkage and erythrocyte membrane scrambling, an effect involving Ca2+ entry and requiring activity of SB203580 and calphostin sensitive kinases., (© 2015 The Author(s) Published by S. Karger AG, Basel.)

Background/aims: The JAK1/JAK2 tyrosine kinase inhibitor ruxolitinib is widely used for the treatment of myeloproliferative neoplasm-associated myelofibrosis and other malignancies. Most important side effects include anemia. A common cause of anemia is accelerated suicidal death of erythrocytes or eryptosis, which is characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Mechanisms contributing to the triggering of eryptosis include oxidative stress, Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i), and activation of distinct kinases, such as p38 mitogen activated protein (MAP) kinase. The present study explored whether and how ruxolitinib induces eryptosis., Methods: Phosphatidylserine exposure at the cell surface was estimated from annexin-V-binding, cell volume from forward scatter, hemolysis from hemoglobin release, [Ca2+]i from Fluo3-fluorescence, and ROS formation from DCFDA dependent fluorescence., Results: A 48 hours exposure of human erythrocytes to ruxolitinib (25 µM) significantly increased the percentage of annexin-V-binding cells and significantly decreased forward scatter. Ruxolitinib did not significantly modify Fluo3-fluorescence and DCFDA fluorescence and the effect of ruxolitinib on annexin-V-binding was not significantly modified by removal of extracellular Ca2+. The effect of ruxolitinib on annexin-V-binding was, however, significantly blunted by the p38 MAP kinase inhibitor SB203580 and virtually abolished by the p38 MAP kinase inhibitor skepinone., Conclusion: Ruxolitinib triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part requiring p38 MAP kinase activity., (© 2015 The Author(s) Published by S. Karger AG, Basel.)

Background/aims: The anti-inflammatory, anti-autoimmune, antiparasitic, and anti-viral ether phospholipid edelfosine (1-O-octadecyl-2-O-methylglycero-3-phosphocholine) stimulates apoptosis of tumor cells and is thus considered for the treatment of malignancy. Similar to apoptosis of nucleated cells, erythrocytes may enter eryptosis, the suicidal erythrocyte death characterized by cell shrinkage and phospholipid scrambling of the cell membrane with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include Ca2+ entry with increase of cytosolic Ca2+ activity ([Ca2+]i) and oxidative stress. The present study explored, whether and how edelfosine induces eryptosis., Methods: Flow cytometry and photometry, respectively, were employed to estimate phosphatidylserine exposure at the cell surface from annexin-V-binding, cell volume from forward scatter, hemolysis from hemoglobin release, [Ca2+]i from Fluo3-fluorescence, and abundance of reactive oxygen species (ROS) from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence., Results: A 6 hours exposure of human erythrocytes to edelfosine (5 µM) significantly increased the percentage of annexin-V-binding cells, significantly decreased forward scatter, and significantly increased Fluo3-fluorescence, but did not significantly modify DCFDA fluorescence. The effect of edelfosine on annexin-V-binding was significantly blunted, but not abolished by removal of extracellular Ca2+., Conclusions: Edelfosine triggers cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect in part due to stimulation of Ca2+ entry., (© 2015 The Author(s) Published by S. Karger AG, Basel.)

Rhodococcus chlorophenolicus PCP-1, a mineralizer of polychlorinated phenols, was inoculated into natural sandy loam and peaty soils with pentachlorophenol (PCP) at concentrations usually found at lightly and heavily polluted industrial sites (30 to 600 mg PCP/kg). A single inoculum of 10(5) to 10(8) cells per g of peat soil and as little as 500 cells/g sandy soil initiated mineralization of(14)C-PCP. The mineralization rates of PCP were 130 to 250 mg mineralized per kg soil in 4 months in the heavily (600 mg/kg) polluted soils and 13 to 18 mg/kg in the lightly (30 mg/kg) polluted soils. There were no detectable PCP mineralizing organisms in the soils prior to inoculation, and also there was no significant adaptation of the indigenous microbial population to degrade PCP during 4 months observation in the uninoculated soils. The inoculum-induced mineralization continued for longer than 4 months after a single inoculation. Uninoculated, lightly polluted soils (30 mg PCP/kg) also showed loss of PCP, but some of this reappeared as pentachloroanisol and other organic chlorine compounds (EOX). Such products did not accumulate in theR. chlorophenolicus-inoculated soils, where instead EOX was mineralized 90 to 98%.R. chlorophenolicus mineralized PCP unhindered by the substrate competition offered by the PCP-methylating bacteria indigenously occurring in the soils or by simultaneously inoculated O-methylatingR. rhodochrous.