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The immunolocalization of protease-activated receptors (PARs) and related proteins in splenic sinus endothelial cells was examined using immunofluorescence and electron microscopy. Immunofluorescence microscopy showed that PAR1 colocalized with PAR2, PAR3, and PAR4. PAR4 colocalized with PAR3 and P2Y12. Myosin heavy chain IIA localized to the outer shape and at the base of cells, but did not colocalize with α-catenin. The localization of di-phosphorylated myosin regulatory light chains (ppMLC) was partially detected on the outer circumference and conspicuously at the base of cells. Macrophage migration inhibitory factor (MIF) also localized in cells. Immunogold electron microscopy revealed the localization of PAR1 on the caveolar membrane, plasma membrane, and junctional membrane of cells. PAR2 and PAR3 localized to the plasma membrane of cells. PAR4 localized to the plasma membrane, depressions in the plasma membrane, and cytoplasmic vesicles. PpMLC was detected in stress fibers, but rarely near the adherens junctions of neighboring cells. MIF localized in vesicles on the apical and basal sides of the Golgi apparatus. Electron microscopy of endothelial cells with saponin extraction showed the depression of many coated pits formed by clathrin from the plasma membrane. Stress fibers developed at the base of cells; however, few actin filaments were observed near adherens junctions. These results indicate that PARs play important roles in splenic sinus endothelial cells, such as in endothelial barrier protection and the maintenance of firm adhesion to ring fibers., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The flow patterns of red blood cells through the spleen are intimately linked to clearance of senescent RBCs, with clearance principally occurring within the open flow through the red pulp and slits of the venous sinus system that exists in humans, rats, and dogs. Passage through interendothelial slits (IESs) of the sinus has been shown by MacDonald et al. (Microvasc Res 33:118-134, 1987) to be mediated by the caliber, i.e., slit opening width, of these slits. IES caliber within a given slit of a given sinus section has been shown to operate in an asynchronous manner. Here, we describe a model and simulation results that demonstrate how the supporting forces exerted on the sinus by the reticular meshwork of the red pulp, combined with asymmetrical contractility of stress fibers within the endothelial cells comprising the sinus, describe this vital and intriguing behavior. These results shed light on the function of the sinus slits in species such as humans, rats, and dogs that possess sinusoidal sinuses. Instead of assuming a passive mechanical filtering mechanism of the IESs, our proposed model provides a mechanically consistent explanation for the dynamically modulated IES opening/filtering mechanism observed in vivo. The overall perspective provided is also consistent with the view that IES passage serves as a self-protective mechanism in RBC vesiculation and inclusion removal., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Beta-thalassaemia is an inherited blood disorder characterised by ineffective erythropoiesis and anaemia. Consequently, hepcidin expression is reduced resulting in increased iron absorption and primary iron overload. Hepcidin is under the negative control of transmembrane serine protease 6 (TMPRSS6) via cleavage of haemojuvelin (HJV), a co-receptor for the bone morphogenetic protein (BMP)-mothers against decapentaplegic homologue (SMAD) signalling pathway. Considering the central role of the TMPRSS6/HJV/hepcidin axis in iron homeostasis, the inhibition of TMPRSS6 expression represents a promising therapeutic strategy to increase hepcidin production and ameliorate anaemia and iron overload in β-thalassaemia. In the present study, we investigated a small interfering RNA (siRNA) conjugate optimised for hepatic targeting of Tmprss6 (SLN124) in β-thalassaemia mice (Hbb th3/+ ). Two subcutaneous injections of SLN124 (3 mg/kg) were sufficient to normalise hepcidin expression and reduce anaemia. We also observed a significant improvement in erythroid maturation, which was associated with a significant reduction in splenomegaly. Treatment with the iron chelator, deferiprone (DFP), did not impact any of the erythroid parameters. However, the combination of SLN124 with DFP was more effective in reducing hepatic iron overload than either treatment alone. Collectively, we show that the combination therapy can ameliorate several disease symptoms associated with chronic anaemia and iron overload, and therefore represents a promising pharmacological modality for the treatment of β-thalassaemia and related disorders., (© 2021 Silence Therapeutics. British Journal of Haematology published by British Society for Haematology and John Wiley & Sons Ltd.)

While lymphocytopenia is a common characteristic of coronavirus disease 2019 (COVID-19), the mechanisms responsible for this lymphocyte depletion are unclear. Here, we retrospectively reviewed the clinical and immunological data from 18 fatal COVID-19 cases, results showed that these patients had severe lymphocytopenia, together with high serum levels of inflammatory cytokines (IL-6, IL-8 and IL-10), and elevation of many other mediators in routine laboratory tests, including C-reactive protein, lactate dehydrogenase, α-hydroxybutyrate dehydrogenase and natriuretic peptide type B. The spleens and hilar lymph nodes (LNs) from six additional COVID-19 patients with post-mortem examinations were also collected, histopathologic detection showed that both organs manifested severe tissue damage and lymphocyte apoptosis in these six cases. In situ hybridization assays illustrated that SARS-CoV-2 viral RNA accumulates in these tissues, and transmission electronic microscopy confirmed that coronavirus-like particles were visible in the LNs. SARS-CoV-2 Spike and Nucleocapsid protein (NP) accumulated in the spleens and LNs, and the NP antigen restricted in angiotensin-converting enzyme 2 (ACE2) positive macrophages and dendritic cells (DCs). Furthermore, SARS-CoV-2 triggered the transcription of Il6 , Il8 and Il1b genes in infected primary macrophages and DCs in vitro , and SARS-CoV-2-NP + macrophages and DCs also manifested high levels of IL-6 and IL-1β, which might directly decimate human spleens and LNs and subsequently lead to lymphocytopenia in vivo . Collectively, these results demonstrated that SARS-CoV-2 induced lymphocytopenia by promoting systemic inflammation and direct neutralization in human spleen and LNs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Xiang, Feng, Diao, Tu, Qiao, Yang, Zhang, Wang, Wang, Wang, Liu, Wang, Liu, Chen, Wu and Chen.)

Vesiculation is a ubiquitous process undergone by most cell types and serves a variety of vital cell functions; vesiculation from erythrocytes, in particular, is a well-known example and constitutes a self-protection mechanism against premature clearance, inter alia. Herein, we explore a paradigm that red blood cell derived vesicles may form within the microvascular, in intense shear flow, where cells become adhered to either other cells or the extracellular matrix, by forming tethers or an evagination. Adherence may be enhanced, or caused, by diseased states or chemical anomalies as are discussed herein. The mechanisms for such processes are detailed via numerical simulations that are patterned directly from video-recorded cell microflow within the splenic venous sinus (MacDonald et al. 1987), as included, e.g., as Supplementary Material. The mechanisms uncovered highlight the necessity of accounting for remodeling of the erythrocyte's membrane skeleton and, specifically, for the time scales associated with that process that is an integral part of cell deformation. In this way, the analysis provides pointed, and vital, insights into the notion of what the, often used phrase, cell deformability actually entails in a more holistic manner. The analysis also details what data are required to make further quantitative descriptions possible and suggests experimental pathways for acquiring such.

To evaluate how a cell responds to the external stimuli, treatment, or alteration of the microenvironment, the quantity and quality of mitochondria are commonly used as readouts. However, it is challenging to apply mitochondrial analysis to the samples that are composed of mixed cell populations originating from tissues or when multiple cell populations are of interest, using methods such as Western blot, electron microscopy, or extracellular flux analysis.Flow cytometry is a technique allowing the detection of individual cell status and its identity simultaneously when used in combination with surface markers. Here we describe how to combine mitochondria-specific dyes or the dyes targeting the superoxide produced by mitochondria with surface marker staining to measure the mitochondrial content and activity in live cells by flow cytometry. This method can be applied to all types of cells in suspension and is particularly useful for analysis of samples composed of heterogeneous cell populations.

The spleen is the largest peripheral lymphoid organ and an important site of immune response, in which the blood-spleen barrier (BSB) plays a significant role to resist various pathogens. The BSB structure of duck spleen is different from that of chicken and mammals. However, no information about the development of BSB after the postembryonic age has been reported in ducks. The current study observed the spleen of 1, 7, 14, 21, 35, and 60-day-old ducks by light and electron microscopy to analyze the cellular structural development. The results showed that the spleen index was continuously increased from 1 to 14-day-old ducks. During their early age, the spleen of ducks showed no definite zone of white and red pulp, but the area of the white pulp was large compared to that of the red pulp. The diameter of the ellipsoid was constantly increased in up to 35-day-old duck spleen, while the periellipsoidal lymphatic sheath (PELS) and periarterial lymphatic sheath continuously developed after 1 D. The reticular fibers developed with age; their branching reached the ellipsoidal wall to show a developed framework in the BSB of 14-day-old ducks. After 7 D, the endothelial cells of the sheathed capillary showed a typical cuboidal shape; between these cells, the gaps increased as age advanced, while the thickness of the basement membrane and collagen fibers increased in 35-day-old ducks. The mechanical filtration function of BSB by intravenous injection showed a 1-layer ring of carbon particles restricted in the white pulp in 1-day-old duck spleen; however, in 14 to 60 D, these particles were restricted in the ellipsoid and PELS, forming 2-layer rings of carbon particles. Collectively, the cellular features of the duck BSB developed up to 35 D of postembryonic age to perform their immune function., (Copyright © 2020. Published by Elsevier Inc.)

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the cause of an ongoing pandemic, with increasing deaths worldwide. To date, documentation of the histopathological features in fatal cases of the disease caused by SARS-CoV-2 (COVID-19) has been scarce due to sparse autopsy performance and incomplete organ sampling. We aimed to provide a clinicopathological report of severe COVID-19 cases by documenting histopathological changes and evidence of SARS-CoV-2 tissue tropism., Methods: In this case series, patients with a positive antemortem or post-mortem SARS-CoV-2 result were considered eligible for enrolment. Post-mortem examinations were done on 14 people who died with COVID-19 at the King County Medical Examiner's Office (Seattle, WA, USA) and Snohomish County Medical Examiner's Office (Everett, WA, USA) in negative-pressure isolation suites during February and March, 2020. Clinical and laboratory data were reviewed. Tissue examination was done by light microscopy, immunohistochemistry, electron microscopy, and quantitative RT-PCR., Findings: The median age of our cohort was 73·5 years (range 42-84; IQR 67·5-77·25). All patients had clinically significant comorbidities, the most common being hypertension, chronic kidney disease, obstructive sleep apnoea, and metabolic disease including diabetes and obesity. The major pulmonary finding was diffuse alveolar damage in the acute or organising phases, with five patients showing focal pulmonary microthrombi. Coronavirus-like particles were detected in the respiratory system, kidney, and gastrointestinal tract. Lymphocytic myocarditis was observed in one patient with viral RNA detected in the tissue., Interpretation: The primary pathology observed in our cohort was diffuse alveolar damage, with virus located in the pneumocytes and tracheal epithelium. Microthrombi, where observed, were scarce and endotheliitis was not identified. Although other non-pulmonary organs showed susceptibility to infection, their contribution to the pathogenesis of SARS-CoV-2 infection requires further examination., Funding: None., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Selenium (Se) is an essential trace element and its deficiency can lead to immune dysfunction. Many studies have investigated the immune damage caused by Se deficiency in chickens, but its mechanism still needs to be explored. In this study, we fed 1-day-old Hyline male chickens with Se deficient diets (the Se content was 0.008 mg kg -1 of diet) and a basal diet (the Se content was 0.15 mg kg -1 of diet). The spleen was collected at the sixth week and used for subsequent experiments. The pathological analysis showed that Se deficiency leads to the destruction of the normal nuclear structure of the spleen cell, and we can observe obvious chromatin condensation and nuclear debris. We constructed a transcriptome database and analyzed the abundance of various genes in the spleen by transcriptome sequence. The analysis of differentially expressed genes (DEGS) showed significant changes in 337 genes, including 210 up-regulations and 127 down-regulations after feeding Se deficient diets. Se deficiency can significantly change oxidative stress and inflammatory response genes in chicken spleen. This study confirmed that Se deficiency increased the IL-2 levels, whereas it down-regulated IL-17, IFN-γ and Foxp3, which indicates that the immune dysfunction of the spleen and Th1/Th2 is imbalanced. We also found that Se deficiency down-regulated some related genes for endoplasmic reticulum Ca 2+ transport, leading to endoplasmic reticulum stress (ERS). Moreover, we determined that Se deficiency triggered the low expression of DUSP1/NF-κB. In summary, our results indicate that Se deficiency can inhibit the spleen immune function of chickens by regulating the DUSP1/NF-κB pathway and ERS, leading to spleen damage in chickens. Based on transcriptomics research, our results will help further study the harmful effects of Se deficiency.

Presence of cell-free DNA (cfDNA) in sera of patients with inflammatory bowel diseases (IBD) is a long-known fact. The biological effect of cfDNA administration on cellular autophagy within normal and inflammatory circumstances remains unclear. In this study, the effects of intravenous cfDNA pretreatment on autophagy response were studied in dextran sulfate sodium (DSS)-induced acute experimental colitis. Selected proinflammatory cytokine and autophagy-related gene and protein expressions were compared with clinical and histological activity parameters, and with transmission electron microscopic evaluations. A single intravenous dose of cfDNA pretreatment with cfDNA from colitis exhibited beneficial response concerning the clinical and histological severity of DSS-colitis as compared with effects of normal cfDNA. Pretreatment with colitis-derived cfDNA substantially altered the gene and protein expression of several autophagy and inflammatory cytokine genes in a clinically favorable manner. Autophagy in splenocytes is also altered after colitis-derived cfDNA pretreatment. During the process of acute colitis, the subsequent inflammatory environment presumably results in changes of cfDNA with the potential to facilitate cell protective autophagy. Understanding the molecular mechanisms behind the impact of colitis-associated autophagy, and elucidating alterations of the interaction between autophagy and innate immunity caused by nucleic acids may provide further insight into the etiology of IBD. By targeting or modifying cfDNA, novel anti-inflammatory therapies may be developed., Competing Interests: Competing interests: None declared., (© American Federation for Medical Research 2020. No commercial re-use. See rights and permissions. Published by BMJ.)

An outbreak of suspected iridovirus disease in cultured hybrid grouper (♀Tiger Grouper Epinephelus fuscoguttatus × ♂ Giant Grouper Epinephelus lanceolatus) occurred in the Guangxi Province in July, 2018. In this study, grouper iridovirus Guangxi (SGIV-Gx) was isolated from diseased hybrid grouper that were collected from Guangxi. Cytopathic effects were observed and identified in grouper spleen cells that were incubated with diseased tissue homogenates after 24 h, and the effects increased at 48 h postinfection. The transmission electron microscopy results showed that viral particles that were about 200 nm in diameter with hexagonal profiles were present in the cell cytoplasm of suspected virus-infected cells. The presence of SGIV-Gx (accession number: MK107821) was identified by polymerase chain reaction (PCR) and amplicon sequencing, which showed that this strain was most closely related to Singapore grouper iridovirus (AY521625.1). The detection of SGIV-Gx infection was further supported by novel aptamer (Q2c)-based detection technology. The effects of temperature and pH on viral infectivity were analyzed by using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and cell culture. The results indicated that SGIV-Gx was resistant to exposure to pH levels 5, 7, and 7.5 for 1 h, but its infectivity was remarkably lower at pH levels 3 and 10 after 1 h. The analyses showed that SGIV-Gx was stable for 1 h at 4°C and 25°C but was inactivated after 1 h at 40, 50, and 60°C., (© 2019 American Fisheries Society.)

Ferritin is a protein that regulates the iron ions in humans by storing them in the form of iron oxides. Despite extensive efforts to understand the ferritin iron oxide structures, it is still not clear how ferritin proteins with a distinct light (L) and heavy (H) chain subunit ratio impact the biomineralization process. In situ graphene liquid cell-transmission electron microscopy (GLC-TEM) provides an indispensable platform to study the atomic structure of ferritin mineral cores in their native liquid environment. In this study, we report differences in the iron oxide formation in human spleen ferritins (HSFs) and human heart ferritins (HHFs) using in situ GLC-TEM. Scanning transmission electron microscopy (STEM) along with selected area electron diffraction (SAED) of the mineral core and electron energy loss spectroscopy (EELS) analyses enabled the visualization of morphologies, crystal structures and the chemistry of iron oxide cores in HSFs and HHFs. Our study revealed the presence of metastable ferrihydrite (5Fe2O3·9H2O) as a dominant phase in hydrated HSFs and HHFs, while a stable hematite (α-Fe2O3) phase predominated in non-hydrated HSFs and HHFs. In addition, a higher Fe3+/Fe2+ ratio was found in HHFs in comparison with HSFs. This study provides new understanding on iron-oxide phases that exist in hydrated ferritin proteins from different human organs. Such new insights are needed to map ferritin biomineralization pathways and possible correlations with various iron-related disorders in humans.

Crystal-storing histiocytosis (CSH) is a rare disorder characterized by the accumulation of nonneoplastic histiocytes containing intracytoplasmic crystallized immunoglobulins. In most cases, there is an associated underlying lymphoplasmacytic neoplasm expressing Ig kappa light chain. About 131 cases of CSH have been identified. There is a localized and a generalized form of CSH and it can involve several sites including bone marrow, lungs, lymph nodes, liver, spleen, gastrointestinal tract, and kidney. Generalized CSH is less frequent and involves multiple organs and tends to have a worst prognosis than localized CSH. Around 20 cases of renal involvement in CSH have been reported so far. Paraprotein-induced crystalline nephropathy can be divided into two categories based on whether the crystals in the kidney are intracellular (including light chain proximal tubulopathy with crystals and CSH) or extracellular (including the crystalline variant of myeloma cast nephropathy and crystalglobulin-induced nephropathy). The former tends to present with slowly worsening kidney dysfunction and generally has a good prognosis, whereas the latter usually presents with rapidly progressive renal failure and is associated with poor renal outcome. We present a case of generalized CSH associated with extracellular crystalline nephropathy with a fulminant and fatal clinical course. Kappa light-chain crystals were found exclusively extracellularly within the tubular lumen, not within the tubular epithelial cells nor the histiocytes, and the massive presence of those precipitates led to the acute renal failure. Consequently, we review the renal involvement in CSH in the literature and discuss the unique mechanism of renal injury in this case.

Introduction : In human physiology, the spleen is generally neglected, and its role is considered anecdotal. In sickle cell disease, splenic dysfunction is the main cause of life-threatening complications, particularly in early childhood with the risk of pneumococcal overwhelming sepsis and acute splenic sequestration crisis, notably. During the course of the disease, the spleen functionally declines and anatomically disappears, albeit with great individual variability depending on modulating genetic and environmental factors. Areas covered : The present review aims to provide an overview of spleen structure and function in order to highlight its role in sickling disorders. The clinical features of spleen damage in sickle cell disease, as well as complications and short- and long-term consequences, are reviewed, along with the main therapeutic options. Expert opinion : Management of acute splenic sequestration recurrence and timing of splenectomy in children with sickling disorders are two main areas in which clinical studies are needed.

A quantitative assessment of the genotoxicity of silver nanoparticles (AgNPs) ascribed to its transplacental transfer and tissue distribution in pregnant rats was carried out in this study. A single intravenous (i.v.) injection of AgNPs with a size range from 4.0 to 17.0 nm was administered to pregnant rats at a dose of 2 mg/kg b.w. on the 19th day of gestation. Five groups beside control, each of the five rats were euthanized after 10 min, 1, 6, 12, or 24 h, respectively. The accumulation of nanoparticles (NPs) in mother and fetal tissues was quantified by inductively coupled plasma optical emission spectroscopy, where the highest accumulation level was recorded in maternal blood (0.523 µg/ml) after 24 h of administration. AgNPs induced accumulation in spleen tissue higher than placenta and fetal tissue homogenates. The data showed significantly detected levels of 8-hydroxydeoxyguanosine in all collected samples from administered animals compared with untreated individuals. Level of 8-OHdG in amniotic fluid exhibited the greatest values followed by maternal spleen, kidneys, and liver, respectively. Investigation by transmission electron microscope showed that the transfer of AgNPs through placental wall caused indentation of nuclei, clumped chromatin, pyknotic nuclei, and focal necrotic areas, while AgNPs appeared mainly accumulated in the macrophages of the spleen. Therefore, the data assume that the genotoxicity studies of AgNPs must be recommended during a comprehensive assessment of the safety of novel types of NPs and nanomaterials. Additionally, exposure to AgNPs must be prevented or minimized during pregnancy or prenatal periods.

Tetrodotoxin (TTX) is an extremely toxic marine compound produced by different genera of bacteria that can reach humans through ingestion mainly of pufferfish but also of other contaminated fish species, marine gastropods or bivalves. TTX blocks voltage-gated sodium channels inhibiting neurotransmission, which in severe cases triggers cardiorespiratory failure. Although TTX has been responsible for many human intoxications limited toxicological data are available. The recent expansion of TTX from Asian to European waters and diversification of TTX-bearing organisms entail an emerging risk of food poisoning. This study is focused on the acute toxicity assessment of TTX administered to mice by oral gavage following macroscopic and microscopic studies. Necropsy revealed that TTX induced stomach swelling 2 h after administration, even though no ultrastructural alterations were further detected. However, transmission electron microscopy images showed an increase of lipid droplets in hepatocytes, swollen mitochondria in spleens, and alterations of rough endoplasmic reticulum in intestines as hallmarks of the cellular damage. These findings suggested that gastrointestinal effects should be considered when evaluating human TTX poisoning.

Duck Tembusu virus (DTMUV) is a novel member of flavivirus with the highest viral loads in the spleen. Six-month egg-laying shelducks were intramuscularly injected with DTMUV strain XZ-2012. Morphological analysis revealed the presence of vacuolar degeneration in the periellipsoidal lymphatic sheaths (PELS) of spleen white pulp following infection, especially from 12 hpi to 3 dpi. Ultrastructural images showed an obvious swelling of cells and their mitochondria and endoplasmic reticulum. Using RNA-seq analysis, the expression levels of RIG-I like receptors (RLRs), downstream IRF7 and proinflammatory cytokines IL-6 from RIG-I signaling pathway were non-apparently upregulated at 2 hpi and apparently at 3 dpi, while MHC-II expression was obviously downregulated at 2 hpi. The expression levels of downstream antiviral cytokines type-I IFNs, anti-inflammatory cytokines IL-10, cell adhesion molecules (CAMs), chemokines and their receptors associated with lymphocyte homing were significantly upregulated at 3 dpi. The population of lymphocyte was increased at 6 dpi. The immune function of spleen was recovered starting from 9 dpi. These findings of this study suggest that DTMUV invaded into the spleen via RIG-I signaling pathway and enhanced immune evasion by inhibiting MHC-II expression during the early stage of infection. Additionally, DTMUV induced PELS lesions through activating IL-6 expression. Furthermore, DTMUV increased the expression levels of RLRs, antiviral type-I IFNs, lymphocyte homing-related genes and proteins as well as the number of lymphocytes in the infected duck spleen. Taken altogether, this study provides new insights into the cellular and molecular mechanisms of DTMUV infection in duck spleen., (Copyright © 2018 Elsevier B.V. All rights reserved.)

This study was designed to investigate the effects of excessive fluoride on spleen toxicity. Twenty-four healthy female rats were randomly divided into two groups, each of 12 rats. Each group of female rats was given a control diet and either F - = 0 mg/L or an excessive F - = 150 mg/L in the drinking water for 120 days. The histomorphological and ultrastructural changes in their splenic tissues were observed under light and transmission electron microscopes. DNA damage and splenocyte apoptosis were examined using the micronucleus (MN) assay, single-cell gel electrophoresis (SCGE), and flow cytometry. The expression levels of cytokines, including interleukin (IL)-1β, IL-2, IL-6, and tumor necrosis factor (TNF)-α, were determined through immunohistochemistry and Western-blot analysis. Results demonstrated that the histomorphological characteristics and ultrastructure of the splenic tissues were affected by excessive fluoride. Nuclear dying, nuclear membrane dissolution, mitochondrial vacuolation, and endoplasmic reticulum dilation were observed. SCGE and MN assays showed that the nuclear DNA of splenocytes was damaged by fluoride treatment, and splenocyte apoptosis was exacerbated in the fluoride group. With damage to the splenocyte structure and DNA, the protein expression levels of IL-1β, IL-2, IL-6, and TNF-α were significantly downregulated by exposure to fluoride. Excessive fluoride ingestion caused splenic pathological damage and abnormal cytokine expression in female rats.

Magnetic iron oxide nanoparticles (IONs) display the ability to cross blood - brain barrier and are envisioned as diagnostic and therapeutic applications, but there are few studies on their potential embryonic toxicity in higher vertebrates. This study investigates interaction of IONs with egg albumen and its subsequent toxicity on chicken embryo. Physicochemical interactions of IONs with egg albumen revealed alterations in friccohesity and secondary structural changes due to weak Vander Waals forces. Toxicity assessment of IONs (10, 25, 50, 100, and 200 µg/ml doses) on chicken embryo accounted for 100% mortality at 200 µg/ml dose due to Fe 2+ ions overload. However, lower doses (50 and 100 µg/ml) recorded decrement in whole weights and crown-rump lengths of chicken embryo possibly due to ION-albumen interactions. Histology of brain tissue revealed degeneration of neurons (50-60%) at 10-100 µg/ml dose range of IONs. Toxicity studies of IONs with diverse animal models are needed to set a toxicity benchmark for preventing embryonic toxicity prior to its use in biomedical applications. This is the first study on toxicity assessment of IONs in chicken embryo.

The antigenic heterogeneity of the reticular framework of the white pulp and marginal zone is well documented in the human adult spleen. Immunostaining of α-smooth muscle actin characterizes the heterogeneity of the reticular framework of the white pulp and marginal zone. In the human spleen, the blood cells flow in an open circulation. T and B lymphocytes flow out from the arterial terminal, and migrate in the reticular framework. Homing of lymphocytes to lymphoid tissues is regulated by selective interactions between cell surface homing receptors and tissue vascular addressins at sites of lymphocyte recruitment from the blood. In the present study, mucosal addressin cell adhesion molecule-1 was selectively expressed on α-smooth muscle actin-positive reticular framework. The reticular framework may function in lymphocyte homing and segregation into the periarteriolar lymphoid sheath, lymph follicle and marginal zone.

Structured Illumination Microscopy (SIM) is a super-resolution microscopy method that has significantly advanced studies of cellular structures. It relies on projection of illumination patterns onto a fluorescently labelled biological sample. The information derived from the sample is then shifted to a detectable band, and in the process of image calculation in Fourier space the resolution is doubled. Refractive index homogeneity along the optical path is crucial to maintain a highly modulated illumination pattern necessary for high-quality SIM. This applies in particular to thick samples consisting of large cells and tissues. Surprisingly, sample mounting media for SIM have not undergone a significant evolution for almost a decade. Through identification and systematic evaluation of a number of non-hazardous, water-soluble chemical components of mounting media, we demonstrate an unprecedented improvement in SIM-image quality. Mounting solutions presented in this research are capable of reducing abundant light scattering which constitutes the limiting factor in 3D-SIM imaging of large Hodgkin's lymphoma and embryonic stem cells as well as 10 µm tissue sections. Moreover, we demonstrate usefulness of some of the media in single molecule localisation microscopy. The results presented here are of importance for standardisation of 3D-SIM data acquisition pipelines for an expanding community of users.

Fluoride is known to impair organism's development and function via adverse effects, and autophagy plays a regulation role in human or animal health and disease. At present, there are no reports focused on fluoride-induced autophagy in the animal and human spleen. The objective of this study was to investigate sodium fluoride (NaF)-induced splenocyte autophagy and the potential mechanism via regulation of p-mTOR in growing mice by using the methods of transmission electron microscopy (TEM), immunohistochemistry (IHC), quantitative real-time polymerase chain reaction (qRT-PCR) and western blot. A total of 240 ICR mice were equally allocated into four groups with intragastric administration of distilled water in the control group and 12, 24, 48 mg/kg NaF solution in the experimental groups for 42 days. Results revealed that NaF increased autophagosomes or autolysosomes in spleen. Simultaneously, the autophagy marker LC3 brown punctate staining was increased with NaF dosage increase. On the other hand, NaF caused inhibition of mTOR activity, which was characterized by down-regulation of PI3K, Akt and mTOR mRNA and protein expression levels. And the suppression of mTOR activity in turn resulted in the significantly increased of ULK1 and Atg13 expression levels. Concurrently, NaF increased the levels of mRNA and protein expression of autophagy markers LC3, Beclin1, Atg16L1, Atg12, Atg5 and decreased the mRNA and protein expression levels of p62. The above-mentioned findings verify that NaF induces autophagy via mTOR signaling pathway. The inhibition of mTOR activity and alteration of autophagy-related genes and proteins are the potential molecular mechanism of NaF-induced splenocyte autophagy.

Nowadays scientific achievements in various areas of lives have caused the creation of more and more «foreign body substances» known as xenobiotics. As it is widely accepted that human health is a product of both genetics and the environment; and premise that also holds true for the immune system with unclear morphogenetic aspect, so we selected the purpose of our work as detection of ultrastructural changes in the spleen and thymus under the influence of tryglycidyl ether of polyoxypropylenetriol (TEPPT) and propylene glycol (PP). Subacute experiment has been performed on the matured male rat's with administration of 1/10 LD50 and 1/100 LD50 of TEPPT and PP during 7 days, 15 days, 30 days and 45 days. Obtained materials of spleen and thymus have been investigated with ultramicroscopic and histological examination. Detection of cellular density has been performed. On the base of obtained results we can conclude that structure of spleen and thymus is susceptible to influence of TEPPT and PP. Ultrastructural changes in those organs of the immune system are characterized by margination of chromatin in nuclei, appearance of pronounced invaginations of karyolemma till fragmentation of nuclei; condensed, wrinkled cytoplasm, dilatation of mitochondria, vacuolization of cytoplasm. Such changes are manifestation of hydropic dystrophy and apoptosis development with resulting in reducing of cellular density in 45 days more pronounced under TEPPT influence with 1/10 LD50 dose: in mantle zone of spleen follicle from 171.1±4.1to 123.7±10.8 cells/104 μm2, in marginal zone of spleen follicle from 104.6±3.8 to 79.4±9.7, in cortical zone of thymus from 180.1±3.9 to 128.3±9.1, in medullar zone of thymus from 137.4±3.7 to 98.6±8.3.

The immune system is very sensitive to radiation. This study revealed that adenosine 5′-monophosphate (5′-AMP) increased the DNA contents of the spleen and the spleen index of irradiated mice. Moreover, the exogenous 5′-AMP could significantly repair the ultra-structure of the damaged spleen through transmission electron microscopy. When indicators of the mouse immune system were assessed, the flow cytometry and enzyme-linked immunosorbent assay (ELISA) revealed that the administration of exogenous 5′-AMP could reduce the apoptosis in the splenic cells. It could also regulate the transition of cells towards S phase, increase the proportion of CD4⁺ and CD8⁺ cellular subsets, and enhance the secretion of interleukin-2 (IL-2), IL-4, IL-10, and interferon-γ (IFN-γ). These effects were associated with a decrease in oxidative stress, as evidenced by changes in superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), catalase (CAT), reduced glutathione (GSH), and malondialdehyde (MDA) levels of spleen tissues. These results suggested that exogenous 5′-AMP could repair the damaged spleen, increase the spleen index, and regulate the cell cycles and apoptosis. There was an increase in the production of various cytokines and play a protective role on the immune system of irradiated mice by dynamically adjusting the REDOX balance.

Background: Titanium dioxide (TiO 2 ) is produced at high volumes and applied in many consumer and food products. Recent toxicokinetic modelling indicated the potential of TiO 2 to accumulate in human liver and spleen upon daily oral exposure, which is not routinely investigated in chronic animal studies. A health risk from nanosized TiO 2 particle consumption could not be excluded then., Results: Here we show the first quantification of both total titanium (Ti) and TiO 2 particles in 15 post-mortem human livers and spleens. These low-level analyses were enabled by the use of fully validated (single particle) inductively coupled plasma high resolution mass spectrometry ((sp)ICP-HRMS) detection methods for total Ti and TiO 2 particles. The presence of TiO 2 in the particles in tissues was confirmed by Scanning Electron Microscopy with energy dispersive X-ray spectrometry., Conclusions: These results prove that TiO 2 particles are present in human liver and spleen, with ≥24% of nanosize (< 100 nm). The levels are below the doses regarded as safe in animals, but half are above the dose that is deemed safe for liver damage in humans when taking into account several commonly applied uncertainty factors. With these new and unique human data, we remain with the conclusion that health risks due to oral exposure to TiO 2 cannot be excluded.

The ultrastructure and morphometrics of pigmented macrophages (PMs) were assessed in the spleen of European sea bass experimentally dosed with Cd and Hg. PMs occurred either as solitary cells or as variably structured aggregations, defined as macrophage aggregates (MAs). Light microscopy revealed a high degree of morphological heterogeneity amongst MAs of all experimental groups. At the ultrastructural level, MAs showed a heterogeneous pigment content that was not influenced by the treatment. Cytoplasm rarefaction/vacuolation and euchromatic nuclei, were observed in PMs of dosed fish. Undosed and Cd-dosed samples differ significantly with regard to the following morphometric features: the Minor axis of the best fitting ellipse, Aspect Ratio, and Roundness. In Cd-dosed fish, MAs showed reduced size and complexity. Lacunarity showed significant differences between undosed and both Cd and Hg-dosed samples. These results suggest that heavy metals, and especially Cd, may influence the dynamics of PM aggregation/disaggregation. Variability in splenic MAs was observed both by light and electron microscopy. However, only the morphometric techniques adequately and objectively described the phenomenon, allowing a quantitative/statistical comparison of morphology among experimental groups. These morphometric analyses could be usefully applied in toxicological and ecotoxicological, as well as morpho-functional studies., (© 2018 Wiley Periodicals, Inc.)

Iron (Fe) is an essential metal involved in a wide spectrum of physiological functions. Sub-cellular characterization of the size, composition, and distribution of ferritin(iron) can provide valuable information on iron storage and transport in health and disease. In this study we employ magnetic force microscopy (MFM), transmission electron microscopy (TEM), and electron energy loss spectroscopy (EELS) to characterize differences in ferritin(iron) distribution and composition across injured and non-injured tissues by employing a rodent model of spinal cord injury (SCI). Our biophysical and ultrastructural analyses provide novel insights into iron distribution which are not obtained by routine biochemical stains. In particular, ferritin(iron) rich lysosomes revealed increased heterogeneity in MFM signal from tissues of SCI animals. Ultrastructural analysis using TEM elucidated that both cytosolic and lysosomal ferritin(iron) density was increased in the injured (spinal cord) and non-injured (spleen) tissues of SCI as compared to naïve animals. In-situ EELs analysis revealed that ferritin(iron) was primarily in Fe 3+ oxidation state in both naïve and SCI animal tissues. The insights provided by this study and the approaches utilized here can be applied broadly to other systemic problems involving iron regulation or to understand the fate of exogenously delivered iron-oxide nanoparticles.

Rapid technology growth and its implementation in all spheres of the people's lives dictates the necessity for thorough study of the influence of different chemicals on human's health. This study was undertaken to elucidate the structural changes that occur in the matured rats' spleen experimentally induced by selected xenobiotic, so, purpose of our work was detection of microscopic peculiarities of the spleen under the influence of laproxides. In subacute experiment were uncovered organometric alterations of the matured male rat's spleen after the administration of 1/10 LD50 of polyether-tryglycidyl ether of polyoxypropylene triol (TEPPT). The study was performed on 72 outbreed WAG male matured rats with the weight 200±10g. Histological slides were studied with performing morphometric and statistical methods. We revealed changes of morphologiс data in comparison to control data which shows reactivity of the spleen in response to the induced xenobiotic. The received and analyzed data demonstrate the morphological changes of the spleen, specifically changes of the linear dimensions and weight of the spleen due to the influence of the TEPPT. The spleen is very sensitive to the effects of xenobiotics, in particular, TEPPT that is even reflected in its grossly (weight and linear dimensions) and histological features (reliable changes of the of the white pulp area of the spleen from 17.87±1.04% to 27.37±1.71%, diameter of lymphatic follicles from 426.59±11.18 μm to 382.31±11.73 μm, width of the mantle zone from 45.73±1.08 μm to 37.18±2.29 μm, width of the marginal zone from 81.32±1.79 μm to 74.63±2.08 μm, width of the periarterial zone from 88.73±2.69 μm to 97.24±2.61 μm).

Objectives: Acquired aplastic anemia (AA) is a hematopoietic stem cell disease that leads to hematopoietic disorder and peripheral blood pancytopenia. We investigated whether nutritional support is helpful to AA recovery., Methods: We established a rat model with AA. A nutrient mixture was administered to rats with AA through different dose gavage once per day for 55 d. Animals in this study were assigned to one of five groups: normal control (NC; group includes normal rats); AA (rats with AA); high dose (AA + nutritional mixture, 2266.95 mg/kg/d); medium dose (1511.3 mg/kg/d); and low dose (1057.91 mg/kg/d). The effects of nutrition administration on general status and mitochondrial function of rats with AA were evaluated., Results: The nutrient mixture with which the rats were supplemented significantly improved weight, peripheral blood parameters, and histologic parameters of rats with AA in a dose-dependent manner. Furthermore, we observed that the number of mitochondria in the liver, spleen, kidney, and brain was increased after supplementation by transmission electron microscopy analysis. Nutrient administration also improved mitochondrial DNA content, adenosine triphosphate content, and membrane potential but inhibited oxidative stress, thus, repairing the mitochondrial dysfunction of the rats with AA., Conclusions: Taken together, nutrition supplements may contribute to the improvement of mitochondrial function and play an important role in the recuperation of rats with AA., (Copyright © 2017 Elsevier Inc. All rights reserved.)

Objective: Introduction: The spleen is a multifunctional odd organ, which is a biological filter, but its structure in experimental animals is not fully understood., Patients and Methods: Materials and methods: The ultramicroscopic structure of the cells and vessels of the spleen of amphibian animals (Pelophylax lessonae С. - the pool frog and Pelophylax ridibundus Р. - the marsh frog) are elucidated in the research., Results: Results: The cytopupulation of the spleen is formed by lymphocytes, neutrophils, erythrocytes, macrophages, plasma cells etcetera. A feature of the cellular composition of the parenchyma of the spleen of the frogs is a significant number of neutrophilic eosinophils. The organelles, which situated in the cell cytoplasm, are characteristic of somatic cells. The endothelial lining and the structure of the capillary basement membrane, which has the form of a plate, formed from 8-12 endothelial cells, grouped in complexes and adjoining contacts in adjacent zones, are experimentally substantiated by the somatic type of vascular endothelium of the microcirculatory channel. Such cells are contained on the basal membrane, surrounded by a dense interlacing of collagen, elastic fibers and the main substance, and the sites of endothelial cell adhesion have the uneven contours. The nuclei of the endothelial cells are surrounded by a cytoplasm on all sides, which reveals foamy cysts., Conclusion: Conclusions: The results obtained are of great interest for evolutionary immunomorphology, the extrapolation of which improves understanding of microcirculation, transport of substances in the spleen, and mechanisms for formation of the immune response.

This is the first utilization of advanced analytical electron microscopy methods, including high-resolution transmission electron microscopy, high-angle annular dark field scanning transmission electron microscopy, electron energy loss spectroscopy, and energy-dispersive X-ray spectroscopy mapping to characterize the organ-specific bioprocessing of a relatively inert nanomaterial (nanoceria). Liver and spleen samples from rats given a single intravenous infusion of nanoceria were obtained after prolonged (90 days) in vivo exposure. These advanced analytical electron microscopy methods were applied to elucidate the organ-specific cellular and subcellular fate of nanoceria after its uptake. Nanoceria is bioprocessed differently in the spleen than in the liver.

Background: Lead is well known environmental pollutant, which can cause toxic effects in multiple organ systems. However, the influence of lead oxide nanoparticles, frequently emitted to the environment by high temperature technological processes, is still concealed. Therefore, we investigate lead oxide nanoparticle distribution through the body upon their entry into lungs and determine the microscopic and ultramicroscopic changes caused by the nanoparticles in primary and secondary target organs., Methods: Adult female mice (ICR strain) were continuously exposed to lead oxide nanoparticles (PbO-NPs) with an average concentration approximately 10 6 particles/cm 3 for 6 weeks (24 h/day, 7 days/week). At the end of the exposure period, lung, brain, liver, kidney, spleen, and blood were collected for chemical, histological, immunohistochemical and electron microscopic analyses., Results: Lead content was found to be the highest in the kidney and lungs, followed by the liver and spleen; the smallest content of lead was found in brain. Nanoparticles were located in all analysed tissues and their highest number was found in the lung and liver. Kidney, spleen and brain contained lower number of nanoparticles, being about the same in all three organs. Lungs of animals exposed to lead oxide nanoparticles exhibited hyperaemia, small areas of atelectasis, alveolar emphysema, focal acute catarrhal bronchiolitis and also haemostasis with presence of siderophages in some animals. Nanoparticles were located in phagosomes or formed clusters within cytoplasmic vesicles. In the liver, lead oxide nanoparticle exposure caused hepatic remodeling with enlargement and hydropic degeneration of hepatocytes, centrilobular hypertrophy of hepatocytes with karyomegaly, areas of hepatic necrosis, occasional periportal inflammation, and extensive accumulation of lipid droplets. Nanoparticles were accumulated within mitochondria and peroxisomes forming aggregates enveloped by an electron-dense mitochondrial matrix. Only in some kidney samples, we observed areas of inflammatory infiltrates around renal corpuscles, tubules or vessels in the cortex. Lead oxide nanoparticles were dispersed in the cytoplasm, but not within cell organelles. There were no significant morphological changes in the spleen as a secondary target organ. Thus, pathological changes correlated with the amount of nanoparticles found in cells rather than with the concentration of lead in a given organ., Conclusions: Sub-chronic exposure to lead oxide nanoparticles has profound negative effects at both cellular and tissue levels. Notably, the fate and arrangement of lead oxide nanoparticles were dependent on the type of organs.

The α-lipoic acid is a soft material and useful for the potential biomedical applications. The study was aimed to assess the potency of α-lipoic acid (ALA) against the toxicity of dimethylnitrosamine (DMN) on spleen of mice by assessing the antioxidants, histopathological and ultrastructure changes. The experiment was achieved on six groups of male mice as following; groups 1, 2, 3, and 4 were served as a control, ALA groups, low dose of DMN (DMN-LD; 2mgkg -1 ) and high dose of DMN (DMN-HD; 4mgkg -1 ). Group 5 was received DMN-LD plus ALA and group 6 was given DMN-HD plus ALA. The results indicated that DMN elevated lipid peroxidation, xanthine oxidase, nitric oxide, and decline the antioxidant enzymes as well as raise the C-reactive protein and tumor necrosis factor. A critical obstruction was harmonized with a reduced in lymphocyte number in the white pulp were observed. All the lymphatic nodules appeared smaller in DMN-HD group. In spleen tissues, marked changes of rough endoplasmic reticulum and appearance of three large lymphocytes were noticed. ALA/DMN treatments were improved all the oxidative damage and the ultrastructure changes. The data evince that ALA was eliminated the adverse effects of DMN on spleen of mice., (Copyright © 2017 Elsevier Masson SAS. All rights reserved.)

The ZnGa 1.995 Cr 0.005 O 4 persistent luminescence nanoparticles offer the promise of revolutionary tools for biological imaging with applications such as cell tracking or tumor detection. They can be re-excited through living tissues by visible photons, allowing observations without any time constraints and avoiding the undesirable auto-fluorescence signals observed when fluorescent probes are used. Despite all these advantages, their uses demand extensive toxicological evaluation and control. With this purpose, mice were injected with a single intravenous administration of hydroxylated or PEGylated persistent luminescence nanoparticles at different concentrations and then a set of standard tests were carried out 1day, 1 month and 6 months after the administration. High concentrations of hydroxylated nanoparticles generate structural alterations at histology level, endoplasmic reticulum damage and oxidative stress in liver, as well as rising in white blood cells counts. A mechanism involving the endoplasmic reticulum damage could be the responsible of the observed injuries in case of ZGO-OH. On the contrary, no toxicological effects related to PEGylated nanoprobes treatment were noted during our in vivo experiments, denoting the protective effect of PEG-functionalization and thereby, their potential as biocompatible in vivo diagnostic probes., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Cyclophilins have diverse functions that may affect the course of central nervous system (CNS) inflammatory disorders. Anti-inflammatory and neuroprotective mechanisms may be targeted by inhibition of cyclophilin A-dependent and cyclophilin D-dependent functions, respectively. We tested the effect of cyclophilin inhibition on CNS inflammation by administering N-methyl-4-isoleucine-cyclosporin (NIM811) to mice undergoing experimental allergic encephalomyelitis (EAE). Treatment with NIM811 resulted in significant reduction of EAE clinical severity. Analysis of mitochondrial calcium retention capacity and the course of EAE in cyclophilin D knockout mice indicated that the effect of NIM811 on EAE was not entirely cyclophilin D-dependent. NIM811-treated EAE animals showed reduction in interleukin-2 expression and reduction in CNS inflammatory infiltrates. These results indicate that anti-inflammatory rather than neuroprotective mechanisms associated with cyclophilins are likely involved in the mechanism of NIM811 in EAE., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Red mark syndrome (RMS) and US strawberry disease (US SD) are skin disorders affecting rainbow trout farmed in Europe and USA. The disease etiology has not yet been established. In spite of specific investigations, identifying Rickettsia-like organism (RLO)- and Midichloria-like organism (MLO)-related DNA in affected individuals, these pathogens have never been observed. We performed histological, ultrastructural and biomolecular analysis on skin and spleen samples of trout with RMS. Examination by TEM revealed the presence of intracytoplasmic microorganisms resembling Rickettsiales within macrophages, fibroblasts and erythrocytes. The microorganisms were oval or short rod shaped (400-800 nm in length and 100-200 nm in width) and often showed a cell wall similar to Gram-negative bacteria. PCR analysis for Rickettsiales supported these findings: 53% of affected trout were positive by both PCR and TEM The primers RiFCfw-RiFCrev were used to anneal both the RLO 16S DNA sequence and the MLO 16S DNA sequence. For this reason, and in agreement with previous studies confirming the presence of Rickettsiales-related DNA in trout with RMS, we assume that TEM detected microorganisms morphologically consistent with bacteria belonging to Rickettsiales order and could be considered as possible causative agents of RMS., (© 2016 John Wiley & Sons Ltd.)

To investigate the effects of molybdenum (Mo) and/or cadmium (Cd) on antioxidant function and the apoptosis-related genes in duck spleens. Sixty healthy 11-day-old ducks were randomly divided into six groups of 10 ducks (control, low Mo group, high Mo, Cd, low Mo + Cd, and high Mo + Cd groups). All were fed a basal diet containing low or high dietary doses of Mo and/or Cd. Relative spleen weight, antioxidant indices, apoptosis-related gene mRNA expression levels, and ultrastructural changes were evaluated after 120 days. The results showed that the relative spleen weight decreased significantly in the high Mo + Cd treatment group which compared with control group. Malondialdehyde levels increased and xanthine oxidase and catalase activities decreased in the Mo and/or Cd groups compared with levels in the control group. Bak-1 and Caspase-3 expressions were upregulated in the high Mo + Cd group, while Bcl-2 was downregulated. In addition, mitochondrial crest fracture, swelling, vacuolation, deformed nuclei, and karyopyknosis in both Mo + Cd treated groups were more severe than in the other groups. The results suggest that Mo and/or Cd can induce oxidative stress and apoptosis of spleen via effects on the mitochondrial intrinsic pathway. Moreover, the results indicate the two elements have a possible synergistic relationship.

Niemann-Pick disease, type C1 (Npc1), is an atypical lysosomal storage disorder caused by autosomal recessive inheritance of mutations in Npc1 gene. In the Npc1 mutant mice (Npc1 -/- ), the initial manifestation is enlarged spleen, concomitant with free cholesterol accumulation. Telocytes (TCs), a novel type of interstitial cell, exist in a variety of tissues including spleen, presumably thought to be involved in many biological processes such as nursing stem cells and recruiting inflammatory cells. In this study, we found that the spleen is significantly enlarged in Npc1 -/- mice, and the results from transmission electron microscopy examination and immunostaining using three different TCs markers, c-Kit, CD34 and Vimentin revealed significantly increased splenic TCs in Npc1 -/- mice. Furthermore, hematopoietic stem cells and macrophages were also elevated in Npc1 -/- spleen. Taken together, our data indicate that splenic TCs might alleviate the progress of splenic malfunction via recruiting hematopoietic stem cells and macrophages., (© 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

The endocrine disruptor bisphenol A (BPA) and the pharmaceutical 17α-ethinyl estradiol (EE) are synthetic chemicals with estrogen-like activities. Despite ubiquitous human exposure to BPA, and the wide-spread clinical use of EE as oral contraceptive adjuvant, the impact of these estrogenic endocrine disrupting chemicals (EDCs) on the immune system is unclear. Here we report results of in vivo dose response studies that analyzed the histology and microstructural changes in the spleen of adult male and female CD-1 mice exposed to 4 to 40,000 μg/kg/day BPA or 0.02 to 2 μg/kg/day EE from conception until 12-14 weeks of age. Results of that analysis indicate that both BPA and EE have dose- and sex-specific impacts on the cellular and microanatomical structures of the spleens that reveal minor alterations in immunomodulatory and hematopoietic functions. These findings support previous studies demonstrating the murine immune system as a sensitive target for estrogens, and that oral exposures to BPA and EE can have estrogen-like immunomodulatory affects in both sexes.

The incidence and global distribution of chloroquine resistant (CR) Plasmodium vivax infection has increased since emerging in 1989. The mechanism of resistance in CR P. vivax has not been defined. The resistance likely relates to the formation and disposition of hemozoin as chloroquine's primary mechanism of action involves disruption of hemozoin formation. CR P. berghei strains, like CR P. vivax strains, are confined to reticulocyte host cells and reportedly they do not accumulate appreciable intraerythrocytic hemozoin. Reports comparing hemozoin production between P. vivax strains and CR to chloroquine sensitive (CS) P. berghei are absent. Here we compare in vivo patterns of hemozoin formation and distribution in blood, spleen and liver tissue of male Swiss mice infected with CS or CR P. berghei not treated with chloroquine and CR P. berghei also treated with chloroquine. Light microscopy, laser desorption mass spectrometry and a colorimetric hemozoin assay detect trace hemozoin in the blood of CR P. berghei infected mice but significant hemozoin accumulation in liver and spleen tissue. Field emission in lens scanning electron microscopy reveals CR P. berghei hemozoin crystals are morphologically smaller but similar to those formed by CS parasites. CR P. berghei produces approximately five-fold less total hemozoin than CS strain. Lipid analysis of CS and CR P. berghei sucrose gradient purified bloodstage hemozoin indicates a similar lipid environment around the isolated hemozoin, predominately monopalmitic glycerol and monostearic glycerol. In contrast to CR and CS P. berghei, colorimetric hemozoin analysis of P. vivax strains indicates similar amounts of hemozoin are produced despite differing chloroquine sensitivities. These results suggest CR P. berghei forms significant hemozoin which accumulates in liver and spleen tissues and that the P. vivax chloroquine resistance mechanism differs from P. berghei., (Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Evaluation of iron distribution and density in biological tissues is important to understand the pathogenesis of a variety of diseases and the fate of exogenously administered iron-based carriers and contrast agents. Iron distribution in tissues is typically characterized via histochemical (Perl's) stains or immunohistochemistry for ferritin, the major iron storage protein. A more accurate mapping of iron can be achieved via ultrastructural transmission electron microscopy (TEM) based techniques, which involve stringent sample preparation conditions. In this study, we elucidate the capability of magnetic force microscopy (MFM) as a label-free technique to map iron at the nanoscale level in rodent spleen tissue. We complemented and compared our MFM results with those obtained using Perl's staining and TEM. Our results show how MFM mapping corresponded to sizes of iron-rich lysosomes at a resolution comparable to that of TEM. In addition MFM is compatible with tissue sections commonly prepared for routine histology., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Subcutaneous daily injection (with neglect of aseptics) of 0.5 ml solution of soybean cream substitute (10% volume in distilled water) during 30 days caused systemic amyloidosis in 30-day-old mice. All the known methods for induction of systemic amyloidosis are based on the use of old animals, as senile tissue bradytrophy allows effective simulation of amyloidosis.

The immune function of the chicken spleen depends on its different compartments of red and white pulps, but little is known about the mechanism underlying lymphocyte homing towards the different compartments. In the present study, the role of lymphocyte homing in the chicken spleen was investigated during lipopolysaccharide (LPS) stimulation. Morphological analysis demonstrated the cuboidal endothelial cells of the splenic sheathed capillary facilitated the passage of lymphocyte homing to the chicken spleen. The tissue-specific adhesion molecules- vascular cell adhesion molecule-1 (VCAM-1) and mucosal addressin cell adhesion molecule-1 (MADCAM-1) expressed on the sheathed capillary, which suggested the high endothelial venule (HEV)-like vessels of the chicken spleen. Electron microscope analysis showed LPS activated the endothelium of the sheathed capillary and recruited lymphocytes to the chicken spleen. Transferring of 5, 6- carboxyfluorescein diacetate, succinimidyl ester (CFSE) labeled lymphocytes depicted the rout of lymphocyte homing to the compartments of the chicken spleen was from the white pulp to the red pulp. Furthermore, the mRNA and protein levels of adhesion molecular integrin β1 and VCAM-1 increased after LPS stimulation. The mechanism underlying the integrin β1 and VCAM-1 during LPS stimulation might be associated with the integrin linked kinase (ILK)- dependent regulation of protein kinase B (PKB/AKT). This study firstly shows lymphocyte homing in the chicken spleen after LPS-induced inflammation. These results contribute to our knowledge of comparative immunology and provide a better means for investigating the pharmacological strategies concerning the possible role of lymphocyte homing in inflammation and immunological reactions in infectious disease.

Microvascular anatomy and histomorphology of larval and adult spleens of the Clawed Toad, Xenopus laevis were studied by light microscopy of paraplast embedded serial tissue sections and scanning electron microscopy (SEM) of vascular corrosion casts (VCCs). Histology showed i) that white and red pulp are present at the onset of metamorphic climax (stage 57) and ii) that splenic vessels penetrated deeply into the splenic parenchyma at the height of metamorphic climax (stage 64). Scanning electron microscopy of VCCs demonstrated gross arterial supply and venous drainage, splenic microvascular patterns as well as the structure of the interstitial (extravasal) spaces representing the "open circulation routes." These spaces identified themselves as interconnected resin masses of two distinct forms, namely "broccoli-shaped" forms and highly interconnected small resin structures. Arterial and venous trees were clearly identified, as were transitions from capillaries to interstitial spaces and from interstitial spaces to pulp venules. Venous sinuses were not diagnosed (nonsinusal spleen). The splenic circulation in Xenopus laevis is "open." It is hypothesized that red blood cells circulate via splenic artery, central arteries, penicillar arteries, and red pulp capillaries primarily via "broccoli-shaped" interstitial spaces, pulp venules and veins into subcapsular veins to splenic veins while lymphocytes circulate also via the interstitial spaces represented by the highly interconnected small resin structures in vascular corrosion casts. In physiological terms, the former most likely represent the fast route for blood circulation, while the latter represent the slow route. J. Morphol. 277:1559-1569, 2016. © 2016 Wiley Periodicals, Inc., (© 2016 Wiley Periodicals, Inc.)

The distribution of iron-carbon nanoparticles in FeC-DSPE-PEG-2000 modification (micellar particles with structure (Fe) core-carbon shell; PEG-based coating) is studied. The greater part of the nanoparticles accumulated in the spleen and liver, a small amount in the lungs, and the minimum amount in the thymus. The structural changes in the lymphoid organs were minor and involved only the microcirculatory bed. Analysis of the peripheral blood showed manifest anemia, thrombocytopenia, and leukocytosis.

The outbreak of Chinese Giant Salamander (Andrias davidianus, CGS) Iridovirus (CGSIV) caused massive death of CGSs. However, some CGSs with low level of CGSIV usually could survive. In our study, major capsid protein (MCP) DNA replicates of CGSIV in shedding skin were employed to assess the relative content of CGSIV in the living CGSs by qPCR. Furthermore, the examinations of autophagy and apoptosis in CGSs in vivo and in the primary renal cells in vitro were performed, respectively. The results showed that the relative contents of CGSIV in the shedding skin could reflect those in liver, spleen, and kidney of the CGSs. In these tissues of the CGSs with low-level replicates of CGSIV, there were not obviously macroscopic lesions. But the irregularly-shaped vesicles perhaps involving in autophagosome were observed by transmission electron microscopy (TEM). The LC3B protein displayed uneven distribution by Immunohistochemistry and the level mRNA of Atg5 was higher in these tissues than that in the tissues of healthy CGSs using qRT-PCR. Meanwhile, the apoptosis also appeared in these tissues by TUNEL staining and higher level mRNA of type I IFN were detected in these tissues using qRT-PCR. Further, both the expression level of LC3B II protein and Atg5 mRNA increased significantly at 2h after the virus infected the primary renal cells from the health CGSs in vitro. In addition, apoptosis and type I IFN mRNA began to increase significantly at 4h after the virus infected the renal cells. It was suggested that autophagy may be a pivotal role for survival of CGSIV in the CGSs during early infection and the rapid proliferation of CGSIV could be inhibited by innate immune response and apoptosis., (Copyright © 2016 Elsevier B.V. All rights reserved.)