3-D neurohistology of transparent tongue in health and injury with optical clearing

Tongue receives extensive innervation to perform taste, sensory, and motor functions. Details of the tongue neuroanatomy and its plasticity in response to injury offer insights to investigate tongue neurophysiology and pathophysiology. However, due to the dispersed nature of the neural network, standard histology cannot provide a global view of the innervation. We prepared transparent mouse tongue by optical clearing to reveal the spatial features of the tongue innervation and its remodeling in injury. Immunostaining of neuronal markers, including PGP9.5 (pan-neuronal marker), calcitonin gene-related peptide (sensory nerves), tyrosine hydroxylase (sympathetic nerves), and vesicular acetylcholine transporter (cholinergic parasympathetic nerves and neuromuscular junctions), was combined with vessel painting and nuclear staining to label the tissue network and architecture. The tongue specimens were immersed in the optical-clearing solution to facilitate photon penetration for 3-dimensiontal (3-D) confocal microscopy. Taking advantage of the transparent tissue, we simultaneously revealed the tongue microstructure and innervation with subcellular-level resolution. 3-D projection of the papillary neurovascular complex and taste bud innervation was used to demonstrate the spatial features of tongue mucosa and the panoramic imaging approach. In the tongue injury induced by 4-nitroquinoline 1-oxide administration in the drinking water, we observed neural tissue remodeling in response to the changes of mucosal and muscular structures. Neural networks and the neuromuscular junctions were both found rearranged at the peri-lesional region, suggesting the nerve-lesion interactions in response to injury. Overall, this new tongue histological approach provides a useful tool for 3-D imaging of neural tissues to better characterize their roles with the mucosal and muscular components in health and disease

Evaluation of Antioxidant and Free Radical Scavenging Capacities of Polyphenolics from Pods of Caesalpinia pulcherrima

Thirteen polyphenolics were isolated from fresh pods of Caesalpinia pulcherrima using various methods of column chromatography. The structures of these polyphenolics were elucidated as gallic acid (1), methyl gallate (2), 6-O-galloyl-d-glucoside (3), methyl 6-O-galloyl-β-d-glucoside (4), methyl 3,6-di-O-galloyl-α-d-glucopyranoside (5), gentisic acid 5-O-α-d-(6′-O-galloyl)glucopyranoside (6), guaiacylglycerol 4-O-β-d-(6′-O-galloyl)glucopyranoside (7), 3-methoxy-4-hydroxyphenol 1-O-β-d-(6′-O-galloyl) glucopyranoside (8), (+)-gallocatechin (9), (+)-catechin (10), (+)-gallocatechin 3-O-gallate (11), myricetin 3-rhamnoside (12), and ampelopsin (13). All isolated compounds were tested for their antioxidant activities in the 1,1-diphenyl-2-picrylhydrazyl (DPPH), hydroxyl, and peroxynitrite radicals scavenging assays. Among those compounds, 11, 12, and 2 exhibited the best DPPH-, hydroxyl-, and peroxynitrite radical-scavenging activities, respectively. Compound 7 is a new compound, and possesses better scavenging activities towards DPPH but has equivalent hydroxyl radical scavenging activity when compared to BHT. The paper is the first report on free radical scavenging properties of components of the fresh pods of Caesalpinia pulcherrima. The results obtained from the current study indicate that the free radical scavenging property of fresh pods of Caesalpinia pulcherrima may be one of the mechanisms by which this herbal medicine is effective in several free radical mediated diseases

Effect of Aging on the Macrophage Response to Titanium Particles

Macrophage-mediated inflammatory reaction to implant wear particles drives bone loss around total joint replacements (TJR). Although most TJR recipients are elderly, studies linking wear particle-activated macrophages and peri-implant osteolysis have not taken into account the multiple effects that aging has on the innate immune system and, in particular, on macrophages. To address this, we compared the wear particle responses of bone marrow macrophages obtained from young (2-month) and aged (18-month) mice. Macrophages were polarized to M0, M1, or M2 phenotypes in vitro, challenged with titanium particles, and their inflammatory response was characterized at multiple time points by quantitative reverse-transcription polymerase chain reaction and enzyme-linked immunosorbent assay. In addition, age-dependent changes in activation of transcription factor nuclear factor-kappa B were analyzed by a lentiviral vector-based luciferase reporter system. The particle stimulation experiment was further repeated using human primary macrophages isolated from blood donors of different ages. We found that the pro-inflammatory responses were generally higher in macrophages obtained from young mice, but differences between the age groups remained small and of uncertain biological significance. Noteworthily, M2 polarization effectively suppressed the particle-induced inflammation in both young and aged macrophages. These results suggest that aging of the innate immune system per se plays no significant role in the response of macrophages to titanium particles, whereas induction of M2 polarization appears a promising strategy to limit macrophage-mediated inflammation regardless of age.Peer reviewe

Recombinant Zebrafish ␥-Glutamyl Hydrolase Exhibits Properties and Catalytic Activities Comparable with Those of Mammalian Enzyme

ABSTRACT: A cDNA encoding for zebrafish ␥-glutamyl hydrolase (␥GH) was cloned and inserted into a pET43.1a vector via SmaI and EcoRI sites and expressed in Rosetta (DE3) cells as a Nus-His-tag fusion enzyme (NH-z␥GH). After induction with isopropyl thiogalactoside, the enzyme was purified with a Ni-Sepharose column, and approximately 8 mg of pure enzyme was obtained per liter of culture. The primary sequence of the recombinant z␥GH was similar to mammalian ␥GH. Folate is an essential B vitamin and participates in the biosynthesis and metabolism of nucleic acids, proteins, several amino acids, methyl groups, many neurotransmitters, and some vitamins. Mammalian cells are unable to synthesize folates de novo and therefore depend on their food for the supply of folates. Naturally occurring folates are synthesized as poly-␥-glutamate forms (folylpolyglutamate) but are absorbed and transported most efficiently as folylmonoglutamates. The conversion of folylpolyglutamates in dietary food to folylmonoglutamates is catalyzed by carboxypeptidase II (EC 3.4.22.12) in mammals. In a recent study, ␥-glutamyl hydrolase (␥GH, EC 3.4.19.9), a lysosomal cysteine peptidase, was reported to be the enzyme responsible for hydrolyzing dietary folate in rat small intestine Consistent with this notion, the activity of ␥GH to hydrolyze the ␥-glutamyl peptide bonds of folylpolyglutamates has rendered this enzyme a potential target of antifolate chemotherapy and, at the same time, a primary component in regulating the intracellular levels of some antifolate drugs. Antifolate drugs, such as methotrexate, owe much of their effectiveness to being substrates for both folylpoly-␥-glutamate synthetase and ␥GH. Removal of ␥-linked glutamate residues decreases the retention and activity of these drugs. A polymorphism resulting in reduced catalytic activity of ␥GH was observed to be associated with greater accumulation of long-chain methotrexate polyglutamate forms The determination of individual folate derivatives in serum of patients receiving antifolate chemotherapy and in foods is an important current protocol. The first step in these determinations is converting folylpolyglutamates to folylmonoglutamates by ␥GH. Cur- The amino acid numbering used for z␥GH in this study is numbered starting from the first methionine in the full-length peptide with the signal peptide. Article, publication date, and citation information can be found a

Nanomedicine Approaches for Autophagy Modulation in Cancer Therapy

Cancer is a daunting global health problem with a steadily rising incidence. Despite the wide arsenal of current anticancer therapies, challenges such as drug resistance, tumor heterogeneity, poor targeting, and severe side effects often lead to suboptimal efficacy and poor patient outcomes, highlighting the need for innovative therapies. Autophagy modulation has emerged as an attractive approach to complement existing therapies. The dual role of autophagy in cancer promotion and suppression has inspired the development of new drugs and therapeutic strategies focusing on both inhibition and induction. Despite the promising results of current autophagy modulators in preclinical studies, challenges such as the lack of selectivity and potency, toxicity, poor pharmacokinetics, and inadequate tumor targeting continue to limit their successful clinical translation. Many of these challenges could be overcome using nanomedicine. This review explores recent advancements in nanomedicine strategies for autophagy modulation. Successful combination strategies leveraging nanoparticles and autophagy modulators in synergy with chemotherapy, immunotherapy, phototherapy, gene therapy, and other modalities are presented. Additionally, nanomaterials with intrinsic autophagy-modulating capabilities, such as self-assembling autophagy inhibitors, are discussed. Finally, limitations of autophagy modulators currently in clinical trials are discussed, and future perspectives on designing nanomedicine for successful clinical implementation are explored

Establishment of Green Fluorescent Protein and Firefly Luciferase Expressing Mouse Primary Macrophages for In Vivo Bioluminescence Imaging

Macrophages play a key role in tissue homeostasis as well as in a range of pathological conditions including atherosclerosis, cancer, and autoimmunity. Many aspects of their in vivo behavior are, however, poorly understood. Bioluminescence imaging (BLI) with green fluorescent protein (GFP) and firefly luciferase (FLUC) labelled autologous reporter macrophages could potentially offer a powerful tool to study macrophage biology, but this approach has been hindered by the relative difficulty of efficient gene transfer into primary macrophages. Here we describe a straightforward method for producing large numbers of GFP/FLUC expressing mouse primary macrophages utilizing lentivirus vector, cyclosporine, and a double infection strategy. Using this method we achieved up to 60% of macrophages to express GFP with correspondingly high FLUC signal. When injected into the circulation using a mouse model of local biomaterial induced inflammation and osteolysis, macrophages were initially detectable within the lungs, followed by systemic homing to the local area of chronic inflammation in the distal femur. In addition, transduced macrophages maintained their ability to assume M1 and M2 phenotypes although the GFP/FLUC expression was altered by the polarizing signals. These reporter macrophages could prove to be valuable tools to study the role of macrophages in health and disease.Peer reviewe

Gamma Ray and Radon Anomalies in Northern Taiwan as a Possible Preearthquake Indicator around the Plate Boundary

Taiwan is tectonically situated in an oblique collision zone between the Philippine Sea Plate (PSP) and the Eurasian Plate (EP). Continuous observations of gamma rays at the Yangmingshan (YMSG) station and soil radon at the Tapingti (TPT) station were recorded in the volcanic area and around a major fault zone, respectively, in Taiwan for seismic studies. A number of anomalous high gamma ray counts and radon concentrations at certain times were found. It is noted that significant increases of soil radon concentrations were observed and followed by the increase in gamma rays a few days to a few weeks before earthquakes that occurred in northeastern Taiwan. Earthquakes such as these are usually related to the subduction of the PSP beneath the EP to the north along the subduction zone in northern Taiwan (e.g., ML = 6 4, April 20, 2015). It is suggested that the preseismic activity may be associated with slow geodynamic processes at the subduction interface, leading to the PSP movement triggering radon enhancements at the TPT station. Furthermore, the further movement of the PSP might be blocked by the EP, with the accumulated elastic stress resulting in the increase of gamma rays due to the increase in porosity and fractures below the YMSG station. The continuous monitoring of the multiple parameters can improve the understanding of the relationship between the observed radon and gamma ray variations and the regional crustal stress/strain in north and northeastern Taiwan

Precision Medicine for Apical Lesions and Peri-Endo Combined Lesions Based on Transfer Learning Using Periapical Radiographs

An apical lesion is caused by bacteria invading the tooth apex through caries. Periodontal disease is caused by plaque accumulation. Peri-endo combined lesions include both diseases and significantly affect dental prognosis. The lack of clear symptoms in the early stages of onset makes diagnosis challenging, and delayed treatment can lead to the spread of symptoms. Early infection detection is crucial for preventing complications. PAs used as the database were provided by Chang Gung Memorial Medical Center, Taoyuan, Taiwan, with permission from the Institutional Review Board (IRB): 02002030B0. The tooth apex image enhancement method is a new technology in PA detection. This image enhancement method is used with convolutional neural networks (CNN) to classify apical lesions, peri-endo combined lesions, and asymptomatic cases, and to compare with You Only Look Once-v8-Oriented Bounding Box (YOLOv8-OBB) disease detection results. The contributions lie in the utilization of database augmentation and adaptive histogram equalization on individual tooth images, achieving the highest comprehensive validation accuracy of 95.23% with the ConvNextv2 model. Furthermore, the CNN outperformed YOLOv8 in identifying apical lesions, achieving an F1-Score of 92.45%. For the classification of peri-endo combined lesions, CNN attained the highest F1-Score of 96.49%, whereas YOLOv8 scored 88.49%