The trematode Acrolichanus auriculatus is a widely distributed intestine parasite of acipenserid fishes. For the first time the localization and distribution of the serotonergic nerve elements in A. ...auriculatus was studied using immunocytochemical method and confocal laser scanning microscopy. The study revealed the presence of biogenic amine, serotonin, in the central and peripheral nervous systems of A. auriculatus, that is in the neurons and neurites of the brain ganglia, brain commissure, the longitudinal nerve cords, and the connective nerve commissures. The innervation of the attachment organs, pharynx, oesophagus and distal regions of the reproductive system by the serotonergic nerve elements is observed. The distribution of serotonergic neurons in A. auriculatus is schematically marked. The comparative analysis of findings obtained in A. auriculatus with those recorded for other digeneans reveals the presence of both conservative and distinctive features in the organization of the serotonergic nervous system in various representatives of trematodes.
•Serotonin is found in central nervous system of A. auriculatus, a parasite of starlet.•Serotonergic innervation of the parasite attachment and internal organs is observed.•Number and size of serotonergic neurons varies among different species of trematodes.•Majority of serotonergic neurons is situated in brain and nerve cords of trematodes.
•Bio-actuated Ti(HPO4)2/Zn3(PO4)2(0.1).nH2O bioceramic was synthesized.•Bone-mimicking structure of phosphate-rich bioceramic improved bone cells proliferation.•Controlled released of Zn++ stimulates ...cells for up-regulation of osteogenic genes expression.•Osteoinduction of the bioceramic increased bone mineralization and new bone formation.
In this work, we describe the synthesis of an ideal bioceramic to which cells can easily respond that has geometry and composition that are similar to those of bone-like apatite. The successful preparation of bone-inspired titanium hydrate phosphate with an optimized weight percentage (∼1.1 wt%) of zinc in (Ti(HPO4)2/Zn3(PO4)2·nH2O) bioceramic could represent a potential regenerative framework for bone defect repair and bone void filler that would be useful for various clinical approaches. We have for the first time demonstrated the detailed physicochemical and biological characterizations of the bioceramic required to achieve a biocompatible and porous architecture that is similar to hydroxyapatite. In vitro studies have shown that the phosphate-rich titanium and zinc nanocomposite promote bone regeneration after stimulating MC3T3 − E1 and hBM-MSCs cells activity. The metallic ions and phosphate had the effects of enhancing the cell viability and osteogenic differentiation of the cells, as confirmed by the expression of bone-related markers that included ALP, Col1a1, RUNX2, OPN/Spp1, and OCN through qRT-PCR, Western blotting, and immunocytochemistry. Moreover, the bioceramic has been shown to significantly improves new bone growth in critical size calvarial defect in rat model. The findings of the present work are the first that show the bone regeneration effect of bioceramic, which could be crucial for subsidies of Ca–P-based artificial bone implant materials. Our results provide insight into the Ti(HPO4)2/Zn3(PO4)2(0.1)·nH2O bioceramic in osteogenesis which can help to establish a therapeutic strategy in bone tissue regenerative medicine.
CB
2
was first considered to be the ‘peripheral cannabinoid receptor’. This title was bestowed based on its abundant expression in the immune system and presumed absence from the central nervous ...system. However, multiple recent reports question the absence of CB
2
from the central nervous system. For example, it is now well accepted that CB
2
is expressed in brain microglia during neuroinflammation. However, the extent of CB
2
expression in neurons has remained controversial. There have been studies claiming either extreme‐its complete absence to its widespread expression‐as well as everything in between. This review will discuss the reported tissue distribution of CB
2
with a focus on CB
2
in neurons, particularly those in the central nervous system as well as the implications of that presence. As CB
2
is an attractive therapeutic target for pain management and immune system modulation without overt psychoactivity, defining the extent of its presence in neurons will have a significant impact on drug discovery. Our recommendation is to encourage cautious interpretation of data that have been presented for and against CB
2
's presence in neurons and to encourage continued rigorous study.
This article is part of a themed issue on Cannabinoids. To view the editorial for this themed issue visit
http://dx.doi.org/10.1111/j.1476‐5381.2010.00831.x
The serotonin (5-HT) 5-HT.sub.2A receptor (5-HT.sub.2A R) and 5-HT.sub.2C receptor (5-HT.sub.2C R) in the central nervous system are implicated in a range of normal behaviors (e.g., appetite, sleep) ...and physiological functions (e.g., endocrine secretion) while dysfunctional 5-HT.sub.2A R and/or 5-HT.sub.2C R are implicated in neuropsychiatric disorders (e.g., addiction, obesity, schizophrenia). Preclinical studies suggest that the 5-HT.sub.2A R and 5-HT.sub.2C R may act in concert to regulate the neural bases for behavior. Here, we utilize three distinct biophysical and immunocytochemistry-based approaches to identify and study this receptor complex in cultured cells. Employing a split luciferase complementation assay (LCA), we demonstrated that formation of the 5-HT.sub.2A R:5-HT.sub.2C R complex exists within 50 nm, increases proportionally to the 5-HT.sub.2C R:5-HT.sub.2A R protein expression ratio, and is specific to the receptor interaction and not due to random complementation of the luciferase fragments. Using a proximity ligation assay (PLA), we found that cells stably expressing both the 5-HT.sub.2A R and 5-HT.sub.2C R exhibit 5-HT.sub.2A R:5-HT.sub.2C R heteroreceptor complexes within 40 nm of each other. Lastly, bioluminescence resonance energy transfer (BRET) analyses indicates the formation of a specific and saturable 5-HT.sub.2A R:5-HT.sub.2C R interaction, suggesting that the 5-HT.sub.2A R and 5-HT.sub.2C R form a close interaction within 10 nm of each other in intact live cells. The bioengineered receptors generated for the LCA and the BRET exhibit 5-HT-mediated intracellular calcium signaling as seen for the native receptors. Taken together, this study validates a very close 5-HT.sub.2A R:5-HT.sub.2C R interaction in cultured cells.
Steroid-resistant nephrotic syndrome is a malfunction of the kidney glomerular filter that leads to proteinuria, hypoalbuminemia, edema, and renal failure. Recently, we identified recessive mutations ...in the phospholipase C epsilon 1 gene ( PLCE1) as a new cause of early-onset nephrotic syndrome and demonstrated interaction of PLCε1 with IQGAP1. To further elucidate the mechanism by which PLCE1 mutations cause nephrotic syndrome, we sought to identify new protein interaction partners of PLCε1. We utilized information from the genetic interaction network of C. elegans. It relates the PLCE1 ortholog ( plc-1) to the C. elegans ortholog ( lin-45) of human BRAF ( v-raf murine sarcoma viral oncogene homolog B1). We hypothesized that this may indicate a functional protein-protein interaction. Using GST pull down of HEK293T cell lysates in vitro and coimmunoprecipation of mouse kidney lysates in vivo, we show that BRAF interacts with PLCε1. By immunohistochemistry in rat kidney, we demonstrate that both proteins are coexpressed and colocalize in developing and mature glomerular podocytes, reporting for the first time the expression of BRAF in the glomerular podocyte.
CD20 and CD3 are considered reliable markers for B and T cells, respectively. This study aimed to develop a rapid multiple immunofluorescence (RMIF) method for the detection of CD20 and CD3 on a ...single cytology slide. Air-dried smears were prepared using samples collected from dogs (n=26) and cats (n=6). Immunosignal detection using the newly developed method required 60 min. Clear immunosignals for CD20 and CD3 were detected in 24 of 26 samples in dogs and in all 6 cats. As the RMIF (CD20/CD3) method can detect markers of both B and T cells simultaneously on a single cytology smear, it would be an efficient tool for the immunophenotyping of canine and feline lymphoma samples.
Immunofluorescence (IF) is an important immunochemical technique that allows for detection and localization of a wide variety of antigens in different types of tissues of various cell preparations. ...IF allows for excellent sensitivity and amplification of signal in comparison to immunohistochemistry, employing various microscopy techniques. There are two methods available, depending on the scope of the experiment or the specific antibodies in use: direct (primary) or indirect (secondary). Here, we describe preparation of specimens preserved in different types of media and step-by-step methods for both direct and indirect immunofluorescence staining.
The correct spatial distribution of proteins is vital for their function and often mis‐localization or ectopic expression leads to diseases. For more than a decade, the Human Protein Atlas (HPA) has ...constituted a valuable tool for researchers studying protein localization and expression in human tissues and cells. The centerpiece of the HPA is its unique antibody collection for mapping the entire human proteome by immunohistochemistry and immunocytochemistry. By these approaches, more than 10 million images showing protein expression patterns at a single‐cell level were generated and are publicly available at www.proteinatlas.org. The antibody‐based approach is combined with transcriptomics data for an overview of global expression profiles. The present article comprehensively describes the HPA database functions and how users can utilize it for their own research as well as discusses the future path of spatial proteomics.
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