Light Microscopic Level Research Articles

BALB/c Tipi Fare Embriyolarında Gelişen Nefronda Juxtaglomerular Aparatus Hücrelerinin Farklılaşmaları

Objective: This study examined the development of nephrons and the differentiation of juxtaglomerular apparatus (JGA) cells in BALB/c type mouse embryos. Thick and thin epon sections were investigated by light microscopy (LM) and transmission electron microscopy (TEM) respectively.
 Materials and Methods: Nephron development is completed by passing through the nephrogenic vesicles in the respected stages of comma shape body, S-shape body, precapillary, immature glomerular, and mature glomerular stage. JGA is distinguished in the mature glomerular stage of nephron development at LM level. According to TEM observations, signs of differentiation of juxtaglomerular (JG) cells and macula densa (MD) cells forming JGA are seen earlier, in precapillary stage. In developed Golgi fields that characterize JG cells, large number of electron lucent small vesicles and small number of electron dense large renin granules are seen in the precapillary stage. In the mature glomerulus, the number of electron dense large granules increases in JG cells. The differentiation of MD cells is indicated by the formation of primary cilia on their apical faces in precapillary stage. Deep membrane folds are formed in the basal and lateral faces of MD cells in the mature glomerular stage. There are many numbers of large mitochondria in these regions.
 Results: JG and MD cells, which play an absolute role in the regulation of the renin-angiotensin system are differentiated in precapillary stage before completion of nephron development and formation of blood vessels.
 Conclusion: Such early differentiation of JGA cells suggests that the renin-angiotensin system is important both in the development of the kidney and in the total development of embryo.

Seeing Keratinocyte Proteins through the Looking Glass of Intrinsic Disorder

Epidermal keratinocyte proteins include many with an eccentric amino acid content (compositional bias), atypical ultrastructural fate (built-in protease sensitivity), or assembly visible at the light microscope level (cytoplasmic granules). However, when considered through the looking glass of intrinsic disorder (ID), these apparent oddities seem quite expected. Keratinocyte proteins with highly repetitive motifs are of low complexity but high adaptation, providing polymers (e.g., profilaggrin) for proteolysis into bioactive derivatives, or monomers (e.g., loricrin) repeatedly cross-linked to self and other proteins to shield underlying tissue. Keratohyalin granules developing from liquid–liquid phase separation (LLPS) show that unique biomolecular condensates (BMC) and proteinaceous membraneless organelles (PMLO) occur in these highly customized cells. We conducted bioinformatic and in silico assessments of representative keratinocyte differentiation-dependent proteins. This was conducted in the context of them having demonstrated potential ID with the prospect of that characteristic driving formation of distinctive keratinocyte structures. Intriguingly, while ID is characteristic of many of these proteins, it does not appear to guarantee LLPS, nor is it required for incorporation into certain keratinocyte protein condensates. Further examination of keratinocyte-specific proteins will provide variations in the theme of PMLO, possibly recognizing new BMC for advancements in understanding intrinsically disordered proteins as reflected by keratinocyte biology.

Linking Fibrotic Remodeling and Ultrastructural Alterations of Alveolar Epithelial Cells after Deletion of Nedd4-2.

Our previous study showed that in adult mice, conditional Nedd4-2-deficiency in club and alveolar epithelial type II (AE2) cells results in impaired mucociliary clearance, accumulation of Muc5b and progressive, terminal pulmonary fibrosis within 16 weeks. In the present study, we investigated ultrastructural alterations of the alveolar epithelium in relation to interstitial remodeling in alveolar septa as a function of disease progression. Two, eight and twelve weeks after induction of Nedd4-2 knockout, lungs were fixed and subjected to design-based stereological investigation at the light and electron microscopic level. Quantitative data did not show any abnormalities until 8 weeks compared to controls. At 12 weeks, however, volume of septal wall tissue increased while volume of acinar airspace and alveolar surface area significantly decreased. Volume and surface area of alveolar epithelial type I cells were reduced, which could not be compensated by a corresponding increase of AE2 cells. The volume of collagen fibrils in septal walls increased and was linked with an increase in blood–gas barrier thickness. A high correlation between parameters reflecting interstitial remodeling and abnormal AE2 cell ultrastructure could be established. Taken together, abnormal regeneration of the alveolar epithelium is correlated with interstitial septal wall remodeling.

Anatomy of the Liver of Mizoram Local Pig (Zovawk)

Background: The present study was aimed for the promotion and advancement of the anatomical knowledge at the gross, light microscopic and electron microscopic level in Zovawk (mizo local pig). Methods: The current investigation was done at the Instructional livestock farm, Department of Veterinary Anatomy and Histology, College of Veterinary Sciences and Animal Husbandry, Central Agricultural University, Selesih, Aizawl, Mizoram and Sophisticated Analytical Instrument Facility (SAIF), North Eastern Hill University (NEHU), Shillong, Meghalaya. Six liver samples were collected from six apparently healthy Zovawk animal of either sex and gross morphological observations were done directly after collection. Thereafter tissue samples were collected as such and were preserved in neutral buffer formalin (NBF) and in Karnovsky’s fixative for routine histology and transmission electron microscopic examination, respectively.Result: Zovawk liver shows four distinct borders, i.e. medial, lateral, dorsal and ventral border, two surfaces, i.e. parietal surface or diaphragmatic surface and caudal or visceral surface and six distinct lobes. The average weight of Zovawk liver was 1.402 kg. Weight of the liver was highly correlated with body weight of animal. Histologically, Zovawk liver was characterized with thick Glisson’s capsule and thick connective tissue septa emerging from it, which gives the hepatic lobules its hexagonal shape. Sinusoids of adjacent hepatocytes were lined by stellate shaped Kupffer cells. The ultra-structural examination of liver shows that, the hepatocytes were rich in mitochondria, endoplasmic reticulum and glycogen granules. Free ribosome and well developed rER and dense lysosomal granules were common in those hepatocyte.

Effect of sterilization methods on the mechanical stability and extracellular matrix constituents of decellularized brain tissues

The cell and matrix content in the central nervous system (CNS) regulates a number of fundamental neural processes and decellularized brain matrices have drawn attention for biomedical applications. However, the effect of sterilization methods on the structural/mechanical stability and cell/matrix contents have yet to be addressed. In this study, supercritical carbon dioxide (SC-CO2), ultraviolet (UV) irradiation and ethylene oxide (EtO) sterilizations were applied to SDS-decellularized sheep brain cortical slices, then subjected to mechanical test, quantification of DNA and extracellular matrix (ECM) protein contents. The cell and matrix morphology were quantitatively analyzed at light and scanning electron microscopical levels. The optimum parameters for SC-CO2 sterilization were 150 bar, 37 °C and 60 min, which ensured complete sterility along with EtO sterilization unlike UV treatment. Although histomorphometry revealed no significant alterations on cellular nuclear content of decellularized matrices, SC-CO2 entrained with 6% ethanol yielded better results with respect to preservation of ECM proteins.

3D Analysis of the Synaptic Organization in the Entorhinal Cortex in Alzheimer's Disease.

The entorhinal cortex (EC) is especially vulnerable in the early stages of Alzheimer’s disease (AD). In particular, cognitive deficits have been linked to alterations in the upper layers of EC. In the present report, we examined Layers II and III from eight human brain autopsies (four subjects with no recorded neurologic alterations and four AD cases). We used stereological methods to assess cortical atrophy of the EC and possible changes in the volume occupied by different cortical elements (neuronal and glial cell bodies; blood vessels; and neuropil). We performed 3D ultrastructural analyses of synapses using focused ion beam/scanning electron microscopy (FIB/SEM) to examine possible alterations related to AD. At the light microscope level, we found a significantly lower volume fraction occupied by neuronal bodies in Layer III and a higher volume fraction occupied by glial cell bodies in Layer II in AD cases. At the ultrastructural level, we observed that (1) there was a significantly lower synaptic density in both layers in AD cases; (2) synapses were larger and more complex in Layer II in AD cases; and (3) there was a greater proportion of small and simple synapses in Layer III in AD cases than in control individuals. These structural differences may play a role in the anatomic basis for the impairment of cognitive functions in AD.

Ultrastructure of rhizoids in the marine bryozoan Dendrobeania fruticosa (Gymnolaemata: Cheilostomata).

Bryozoans form colonies of iterated modules, termed zooids, and display varying degrees of polymorphism. Polymorphic colonies comprise autozooids (or feeding zooids) and heteromorphic zooids, among which the most common types are avicularia and kenozooids. Kenozooids differ in shape, size, and presumed function. Among this diversity, there are rhizoids, which serve to attach colonies to the substrate or to lift them above it. To date, only general data on anatomy of kenozooids at light microscopy level are available. Here, we present the first description of the ultrastructure of the holdfast-like rhizoids of the cheilostome bryozoan Dendrobeania fruticosa. The rhizoid wall is composed of a single-layered epidermis, which produces the ectocyst. The voluminous cavity is acoelomate: it has no special cellular lining, nor any signs of an extracellular matrix toward the epidermis. It is traversed by delicate branching funicular strands that originate from the pore plate. The only cells in contact with the epidermis are the cells of the funicular system and the storage cells. The pore plate between the rhizoid and autozooid includes a variable number of communication pores. Each pore is plugged with a rosette complex, which includes a cincture cell and four special cells extending through the pore. The limiting cells are absent, and the special cells are in direct contact with the funicular strands. Cell contacts between special cells are absent; moreover, there are spaces between their proximal lobes filled with a heterogeneous matrix similar to that in the lumen of the funicular strands. Such matrix is also found outside of the extracellular matrix surrounding the special cells. These findings allow us to suggest that nutrient transport most likely occurs between, rather than through, the special cells. However, further studies are needed to understand how the rosette complex functions.

Improved Alveolar Dynamics and Structure After Alveolar Epithelial Type II Cell Transplantation in Bleomycin Induced Lung Fibrosis.

Idiopathic pulmonary fibrosis (IPF) is a progressively and ultimately fatal lung disease. Previously it has been shown that intratracheal administration of alveolar epithelial type II cells (AE2C) in the animal model of bleomycin-induced pulmonary fibrosis is able to reverse fibrosis and restore surfactant protein levels. However, to date, it has not been evaluated whether these changes involve any improvement in alveolar dynamics. Consequently, the aim of the present work was to study lung physiology after AE2C transplantation at different time points during the development of injury and fibrosis. Lung fibrosis was induced by intratracheal instillation of bleomycin (4U/kg) in rat lungs. The animals were transplanted with AE2C (2.5 × 106 cells/animal) 3 or 7 days after bleomycin instillation. Assessments were done at day 7 and 14 after the induction of fibrosis to plot time dependent changes in lung physiology and mechanics. To assess the pressures and rates at which closed alveoli reopens invasive pulmonary tests using a small-animal mechanical ventilator (Flexivent®, Scireq, Canada) including de-recruitability tests and forced oscillation technique as well as quasi-static pressure volume loops were performed. Afterwards lungs were fixed by vascular perfusion and subjected to design-based stereological evaluation at light and electron microscopy level. AE2C delivered during the lung injury phase (3 days) of the disease are only able to slightly recover the volume of AE2C and volume fraction of LB in AE2C. However, it did not show either positive effects regarding ventilated alveolar surface nor any increase of lung compliance. On the other hand, when AE2C are delivered at the beginning of the fibrotic phase (7 days after bleomycin instillation), an increased ventilated alveolar surface to control levels and reduced septal wall thickness can be observed. Moreover, transplanted animals showed better lung performance, with increased inspiratory capacity and compliance. In addition, a detailed analysis of surfactant active forms [mainly tubular myelin, lamellar body (LB)-like structures and multilamellar vesicles (MLV)], showed an effective recovery during the pro-fibrotic phase due to the healthy AE2C transplantation. In conclusion, AE2C transplantation during fibrogenic phases of the disease improves lung performance, structure and surfactant ultrastructure in bleomycin-induced lung fibrosis.

N-cadherin localization in taste buds of mouse circumvallate papillae.

Taste buds, the receptor organs for taste, contain 50-100 taste bud cells. Although these cells undergo continuous turnover, the structural and functional integrity of taste buds is maintained. The molecular mechanisms by which synaptic connectivity between taste buds and afferent fibers is formed and maintained remain ambiguous. In the present study, we examined the localization of N-cadherin in the taste buds of the mouse circumvallate papillae because N-cadherin, one of the classical cadherins, is important for the formation and maintenance of synapses. At the light microscopic level, N-cadherin was predominantly detected in type II cells and nerve fibers in the connective tissues in and around the vallate papillae. At the ultrastructural level, N-cadherin immunoreactivity appears along the cell membrane and in the intracellular vesicles of type II cells. N-cadherin immunoreactivity also is evident in the membranes of afferent terminals at the contact sites to N-cadherin-positive type II cells. At channel type synapses between type II cells and nerve fibers, N-cadherin is present surrounding, but not within, the presumed neurotransmitter release zone, identified by large mitochondria apposed to the taste cells. The present results suggest that N-cadherin is important for the formation or maintenance of type II cell afferent synapses in taste buds.

Distribution and ultrastructural localization of the glucagon-like peptide-1 receptor (GLP-1R) in the rat brain

Glucagon-like peptide-1 (GLP-1) inhibits food intake and regulates glucose homeostasis. These actions are at least partly mediated by central GLP-1 receptor (GLP-1R). Little information is available, however, about the subcellular localization and the distribution of the GLP-1R protein in the rat brain. To determine the localization of GLP-1R protein in the rat brain, immunocytochemistry was performed at light and electron microscopic levels. The highest density of GLP-1R-immunoreactivity was observed in the circumventricular organs and regions in the vicinity of these areas like in the arcuate nucleus (ARC) and in the nucleus tractus solitarii (NTS). In addition, GLP-1R-immunreactive (IR) neuronal profiles were also observed in a number of telencephalic, diencephalic and brainstem areas and also in the cerebellum. Ultrastructural examination of GLP-1R-immunoreactivity in energy homeostasis related regions showed that GLP-1R immunoreactivity is associated with the membrane of perikarya and dendrites but GLP-1R can also be observed inside and on the surface of axon varicosities and axon terminals. In conclusion, in this study we provide a detailed map of the GLP-1R-IR structures in the CNS. Furthermore, we demonstrate that in addition to the perikaryonal and dendritic distribution, GLP-1R is also present in axonal profiles suggesting a presynaptic action of GLP-1. The very high concentration of GLP-1R-profiles in the circumventricular organs and in the ARC and NTS suggests that peripheral GLP-1 may influence brain functions via these brain areas.

Aspartam ve asesülfam K kullanımının testis yapısına etkilerinin ince yapı düzeyinde incelenmesi

Amaç: Bu çalışma, ülkemizde ve dünyada yaygın olarak kullanılan aspartam ve asesülfam K’nın, sıçanlarda testis dokuları üzerindeki etkilerinin ışık ve elektron mikroskobik düzeyde araştırılması amacıyla yapılmıştır.Gereç ve Yöntem: Çalışmada 60 adet erkek sıçan kullanıldı ve bu denekler rastgele olarak 6 eşit gruba ayrıldı. 1. gruptaki hayvanlar kontrol grubu olarak değerlendirildi; 2.gruptaki hayvanlara 200 mg/kg/gün aspartam; 3.gruptaki hayvanlara 300 mg/kg/gün aspartam; 4. gruptaki hayvanlara 300 mg/kg/gün asesülfam K; 5. gruptaki hayvanlara 600 mg/kg/gün asesülfam K ve son gruba da 300 mg/kg/gün aspartam+300 mg/kg/gün asesülfam K birlikte 8 hafta süreyle verildi. 8. haftanın sonunda deneklerden testis dokuları alındı ve dokular ışık ve elektron mikroskobik olarak değerlendirildi.Bulgular: Aspartam ve asesülfam K kullanımının testis dokusunda dejenerasyona yol açtığı ve spermatogenez sürecinde sağlıklı spermatid oluşumunu engellediği izlendi.Sonuç: Aspartam ve asesülfam K tüketiminin erkek sıçanlarda testis dejenerasyonuna neden olduğu görüldü. Bununla birlikte, erkek üreme sistemindeki aspartam ve asesülfam K toksisitesinin doğrulanması için daha kapsamlı çalışmaların yapılması gerektiği kanaatine varıldı.

Histomorphometrical study of the testis in the desert Rodent, Gerbillus tarabuli, sexually immature and adult during the annual reproductive cycle.

Our interest in the testicular study of the desert rodent, Gerbillus tarabuli, would provide a better comprehension of the seasonality of spermatogenesis in rodents. We thus performed a biometric, histomorphometric and statistical analyses in Gerbillus tarabuli sexually immature and adult during the annual reproductive cycle (active and regressed states of gonadal activity). For that, fixed testes were embedded in paraffin. Sections were stained using the modified Heidenhain Azan and then were subjected to morphometric measurements at the light microscopic level. The biometric study revealed a positive correlation between the body weight and the maturity state only, with an average of 20.8±2.92g in sexually immature Gerbils, 37.40±1.86g in the sexually quiescent adults (p<.01) and 40.40±2.98g in the sexually active adults (p>.05), while the testes weight (sexually immatures=0.02±0.003g, sexually quiescent adults=0.13±0.01g; p<.001 and sexually active adults=0.27±0.01g; p<.001) and the gonadosomatic index (sexually immatures=0.09±0.02%, sexually quiescent adults=0.36±0.04%; p<.01 and sexually active adults=0.68±0.07%; p<.01) varied according to the maturity state and as well as to the season in the adults. Through the histomorphometric study, the tubular diameter showed a continuous increase from sexually immature Gerbils (72.83±7.57μm) to sexually quiescent adults (99.45±4.34μm) p<.05 then to sexually active adults (182.58±5.22μm); p<.001, this is explained by the appearance of a large lumen after puberty in the quiescent (26.32±1.28μm); p<.001 and the enlargement of the seminiferous epithelium in actives (75.11±1.81μm); p<.001. Besides, the nucleo-cytoplasmic ratio of Leydig cells (sexually immatures=0.5±0.02) diminishes significantly after puberty in the rest period (0.42±0.03); p<.05 and then during the active period (0.24±0.01); p<.001. These striking biometric and morphometric differences in Gerbillus tarabuli testis between sexually immature Gerbils, sexually quiescent and active adults, follow the photoperiod-induced changes in endocrine and exocrine functions and prove that Gerbillus tarabuli is a useful model to study histo-physiological relationships in the testis in order to develop a more complete understanding of spermatogenesis phenomenon.

Comparative analyses of transgene expression patterns after intra-striatal injections of rAAV2-retro in rats and rhesus monkeys: A light and electron microscopic study.

Retrogradely-transducing viral vectors are versatile tools for anatomical and functional interrogations of neural circuits. These vectors can be applied in nonhuman primates (NHPs), powerful model species for neuroscientific studies with limited genetic tractability, but limited data are available regarding the tropism and transgene expression patterns of such viruses after injections in NHP brains. Consequently, NHP researchers must often rely on related data available from other species for experimental planning. To evaluate the suitability of rAAV2-retro in the NHP basal ganglia, we studied the transgene expression patterns at the light and electron microscope level after injections of rAAV2-retro vector encoding the opsin Jaws conjugated to a green fluorescent protein (GFP) in the putamen of rhesus macaques. For inter-species comparison, we injected the same vector in the rat dorsal striatum. In both species, GFP expression was observed in numerous cortical and subcortical regions with known striatal projections. However, important inter-species differences in pathway transduction were seen, including labeling of the intralaminar thalamostriatal projection in rats, but not monkeys. Electron microscopic ultrastructural observations within the basal ganglia revealed GFP labeling in both postsynaptic dendrites and presynaptic axonal terminals; the latter likely derived from anterograde transgene transport in neurons that project to the striatum, and from collaterals of these neurons. Our results suggest that certain neural pathways may be refractory to transduction by retrograde vectors in a species-specific manner, highlighting the need for caution when determining the suitability of a retrograde vector for NHP studies based solely on rodent data.

Unique blood vasculature and innervation in the cavernous tissue of murine vomeronasal organs.

The vomeronasal organ (VNO) is an accessory olfactory device related to reproductive behavior. The soft tissue of the tubular organ is composed of sensory/non-sensory epithelia and a highly developed vasculature, which in the latter the dilation and contraction of blood vessels are thought to contribute to pumping in and out luminal fluid or air, like penile erectile tissue. The present histological observation of the murine VNO revealed a more complicated vasculature than previously evaluated ones with large differences along the rostro-caudal axis. An immunohistochemical study for vasoactive substances displayed extremely dense innervation by cholinergic nerves containing nitric oxide synthase and VIP/PHI in the thick smooth muscle layer surrounding venous sinuses at light and electron microscopic levels. Furthermore, the differential distribution of cholinergic nerves and adrenergic nerves may provide a novel insight into the pumping mechanism of VNO.

Functional organisation of the endomembrane network in the digestive gland of the Venus flytrap: revisiting an old story with a new microscopy toolbox.

Up-to-date imaging approaches were used to address the spatiotemporal organisation of the endomembrane system in secretory cells of Dionaea muscipula. Different 'slice and view' methodologies were performed on resin-embedded samples to finally achieve a 3D reconstruction of the cell architecture, using ultrastructural tomography, array tomography, serial block face-scanning electron microscopy (SBF-SEM), correlation, and volume rendering at the light microscopy level. Observations of cryo-fixed samples by high-pressure freezing revealed changes of the endomembrane system that occur after trap activation and prey digestion. They provide evidence for an original strategy that adapts the secretory machinery to a specific and unique case of stimulated exocytosis in plant cells. A first secretion peak is part of a rapid response to deliver digestive fluids to the cell surface, which delivers the needed stock of digestive materials 'on site'. The second peak of activity could then be associated with the reconstruction of the Golgi apparatus (GA), endoplasmic reticulum (ER) and vacuolar machinery, in order to prepare for a subsequent round of prey capture. Tubular continuum between ER and Golgi stacks observed on ZIO-impregnated tissues may correspond to an efficient transfer mechanism for lipids and/or proteins, especially for use in rapidly resetting the molecular GA machinery. The occurrence of one vacuolar continuum may permit continuous adjustment of cell homeostasy. The subcellular features of the secretory cells of Dionaea muscipula outline key innovations in the organisation of plant cell compartmentalisation that are used to cope with specific cell needs such as the full use of the GA as a protein factory, and the ability to create protein reservoirs in the periplasmic space. Shape-derived forces of the pleiomorphic vacuole may act as signals to accompany the sorting and entering flows of the cell.

Clonable EM labeling

Cell Biology Widespread use of the green fluorescent protein for protein localization at the light microscopy level has revolutionized cell biology. The search for a similar genetically encoded label for electron microscopy (EM) that could raise the resolution to the single-molecule level has not yet been successful. Jiang et al. developed a method for synthesizing 2- to 6-nm-diameter gold nanoparticles directly and specifically on individual cysteine-rich tags. This enables the unambiguous visualization of genetically tagged proteins in bacterial, yeast, and mammalian cells with single-molecule resolution. This approach holds promise in circumventing the difficulties of conventional EM immunolabeling and enables biologists to probe the ultrastructure of cells with greater ease. Nat. Methods 17 , 937 (2020).

Cellular and Subcellular Localisation of Kv4-Associated KChIP Proteins in the Rat Cerebellum.

The K+ channel interacting proteins (KChIPs) are a family of cytosolic proteins that interact with Kv4 channels, leading to higher current density, modulation of channel inactivation and faster recovery from inactivation. Using immunohistochemical techniques at the light and electron microscopic level combined with quantitative analysis, we investigated the cellular and subcellular localisation of KChIP3 and KChIP4 to compare their distribution patterns with those for Kv4.2 and Kv4.3 in the cerebellar cortex. Immunohistochemistry at the light microscopic level demonstrated that KChIP3, KChIP4, Kv4.2 and Kv4.3 proteins were widely expressed in the cerebellum, with mostly overlapping patterns. Immunoelectron microscopic techniques showed that KChIP3, KChIP4, Kv4.2 and Kv4.3 shared virtually the same somato-dendritic domains of Purkinje cells and granule cells. Application of quantitative approaches showed that KChIP3 and KChIP4 were mainly membrane-associated, but also present at cytoplasmic sites close to the plasma membrane, in dendritic spines and shafts of Purkinje cells (PCs) and dendrites of granule cells (GCs). Similarly, immunoparticles for Kv4.2 and Kv4.3 were observed along the plasma membrane and at intracellular sites in the same neuron populations. In addition to the preferential postsynaptic distribution, KChIPs and Kv4 were also distributed presynaptically in parallel fibres and mossy fibres. Immunoparticles for KChIP3, KChIP4 and Kv4.3 were detected in parallel fibres, and KChIP3, KChIP4, Kv4.2 and Kv4.3 were found in parallel fibres, indicating that composition of KChIP and Kv4 seems to be input-dependent. Together, our findings unravelled previously uncharacterised KChIP and Kv4 subcellular localisation patterns in neurons, revealed that KChIP have additional Kv4-unrelated functions in the cerebellum and support the formation of macromolecular complexes between KChIP3 and KChIP4 with heterotetrameric Kv4.2/Kv4.3 channels.

Chemical and Structural Changes in Corn Stover After Ensiling: Influence on Bioconversion.

Production of biofuels, bioproducts, and bioenergy requires a well-characterized, stable, and reasonably uniform biomass supply and well-established supply chains for shipping biomass from farm fields to biorefineries, while achieving year-round production targets. Preserving and stabilizing biomass feedstock during storage is a necessity for cost-effective and sustainable biofuel production. Ensiling is a common storage method used to preserve and even improve forage quality; however, the impact of ensiling on biomass physical and chemical properties that influence bioconversion processes has been variable. Our objective in this work was to determine the effects of ensiling on lignocellulosic feedstock physicochemical properties and how that influences bioconversion requirements. We observed statistically significant decreases (p < 0.05) in the content of two major structural carbohydrates (glucan and xylan) of 5 and 8%, respectively, between the ensiled and non-ensiled materials. We were unable to detect differences in sugar yields from structural carbohydrates after pretreatment and enzymatic hydrolysis of the ensiled materials compared to non-ensiled controls. Based on this work, we conclude that ensiling the corn stover did not change the bioconversion requirements compared to the control samples and incurred losses of structural carbohydrates. At the light microscopy level, ensiled corn stover exhibited little structural change or relocation of cell wall components as detected by immunocytochemistry. However, more subtle structural changes were revealed by electron microscopy, as ensiled cell walls exhibit ultrastructural characteristics such as wall delimitation intermediate between non-ensiled and dilute-acid-pretreated cell walls. These findings suggest that alternative methods of conversion, such as deacetylation and mechanical refining, could take advantage of lamellar defects and may be more effective than dilute acid or hot water pretreatment for biomass conversion of ensiled materials.

Light and transmission electron microscopy of the haptoral sclerites of the monogenean gill parasite Macrogyrodactylus clarii.

The present study represents the first detailed description of the haptoral sclerites of Macrogyrodactylus clarii Gussev 1961. Light microscopy reveals outgrowths of the hamuli roots; two lateral spine-like extensions of the hood-like accessory sclerites; a ridged, fan-shaped distal end of an accessory sclerite; and two thread-like accessory sclerites with biforked ends associated with the pointed hooked region of each hamulus. Transmission electron microscopy (TEM) reveals the presence of parallel tubules in the root of the hamuli and structural differences along the length of the hamulus including the root, shaft, and pointed hooked region. The root consists of two layers, the shaft four layers and pointed hooked region only one layer with dense outer serrations. Characteristic features of the hamulus root are the presence of longitudinally orientated parallel tubules in its central core and parallel electron-dense ridges in the outer layer of its middle region; features not observed in either the shaft or the pointed hooked region. Each hamulus blade of M. clarii is associated with haptoral gland cells producing electron-dense secretory bodies. The 16 marginal hooklets each consist of a blade (sickle) articulating with a handle at the guard region and a domus. TEM revealed structural differences between the handle, the blade at the articulation region, and the distal hooked region. The domus, a filamentous thread-like sclerite at the light microscope level, consists of two electron-dense, fibrous thickenings connected to each other by a cytoplasmic process. Each marginal hooklet is associated with a small cavity and a large reservoir of homogeneous particles and secretory bodies. The possible functions of these structures are discussed in relation to equivalent features in other monogeneans.

Analyzing microglial-associated A\u03b2 in Alzheimer\u2019s disease transgenic mice with a novel mid-domain A\u03b2-antibody

The mechanisms of amyloid-β (Aβ)-degradation and clearance in Alzheimer’s disease (AD) pathogenesis have been relatively little studied. Short Aβ-fragments form by enzymatic cleavage and alternate amyloid-beta precursor protein (APP)-processing. Here we characterized a novel polyclonal Aβ-antibody raised against an Aβ mid-domain and used it to investigate microglial Aβ-uptake in situ by microscopy at the light- and ultrastructural levels. The rabbit Aβ-mid-domain antibody (ab338), raised against the mid-domain amino acids 21–34 (Aβ21–34), was characterized with biochemical and histological techniques. To identify the epitope in Aβ recognized by ab338, solid phase and solution binding data were compared with peptide folding scores as calculated with the Tango software. The ab338 antibody displayed high average affinity (KD: 6.2 × 10−10 M) and showed preference for C-terminal truncated Aβ-peptides ending at amino acid 34 and Aβ-mid domain peptides with high scores of β-turn structure. In transgenic APP-mouse brain, ab338 labelled amyloid plaques and detected Aβ-fragments in microglia at the ultra- and light microscopic levels. This reinforces a role of microglia/macrophages in Aβ-clearance in vivo. The ab338 antibody might be a valuable tool to study Aβ-clearance by microglial uptake and Aβ-mid-domain peptides generated by enzymatic degradation and alternate production.