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CNS Neuroscience & Therapeutics Jul 2024Comorbid anxiodepressive-like symptoms (CADS) in chronic pain are closely related to the overactivation of the lateral habenula (LHb). Hyperpolarization-activated cyclic...
AIMS
Comorbid anxiodepressive-like symptoms (CADS) in chronic pain are closely related to the overactivation of the lateral habenula (LHb). Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels have been implicated to play a key role in regulating neuronal excitability. However, the role of HCN channels in the LHb during CADS has not yet been characterized. This study aimed to investigate the effect of HCN channels in the LHb on CADS during chronic pain.
METHODS
After chronic neuropathic pain induction by spared nerve injury (SNI), mice underwent a sucrose preference test, forced swimming test, tail suspension test, open-field test, and elevated plus maze test to evaluate their anxiodepressive-like behaviors. Electrophysiological recordings, immunohistochemistry, Western blotting, pharmacological experiments, and virus knockdown strategies were used to investigate the underlying mechanisms.
RESULTS
Evident anxiodepressive-like behaviors were observed 6w after the SNI surgery, accompanied by increased neuronal excitability, enhanced HCN channel function, and increased expression of HCN2 isoforms in the LHb. Either pharmacological inhibition or virus knockdown of HCN2 channels significantly reduced LHb neuronal excitability and ameliorated both pain and depressive-like behaviors.
CONCLUSION
Our results indicated that the LHb neurons were hyperactive under CADS in chronic pain, and this hyperactivation possibly resulted from the enhanced function of HCN channels and up-regulation of HCN2 isoforms.
Topics: Animals; Habenula; Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; Mice; Male; Depression; Neuralgia; Mice, Inbred C57BL; Chronic Pain; Potassium Channels
PubMed: 38961317
DOI: 10.1111/cns.14831 -
CNS Neuroscience & Therapeutics Jul 2024Paclitaxel (PTX) is extensively utilized in the management of diverse solid tumors, frequently resulting in paclitaxel-induced peripheral neuropathy (PIPN). The present...
AIMS
Paclitaxel (PTX) is extensively utilized in the management of diverse solid tumors, frequently resulting in paclitaxel-induced peripheral neuropathy (PIPN). The present study aimed to investigate sex differences in the behavioral manifestations and underlying pathogenesis of PIPN and search for clinically efficacious interventions.
METHODS
Male and female C57BL/6 mice (5-6 weeks and 12 months, weighing 18-30 g) were intraperitoneally (i.p.) administered paclitaxel diluted in saline (NaCl 0.9%) at a dose of 2 mg/kg every other day for a total of 4 injections. Von Frey and hot plate tests were performed before and after administration to confirm the successful establishment of the PIPN model and also to evaluate the pain of PIPN and the analgesic effect of PD-L1. On day 14 after PTX administration, PD-L1 protein (10 ng/pc) was injected into the PIPN via the intrathecal (i.t.) route. To knock down TRPV1 in the spinal cord, adeno-associated virus 9 (AAV9)-Trpv1-RNAi (5 μL, 1 × 10 vg/mL) was slowly injected via the i.t. route. Four weeks after AAV9 delivery, the downregulation of TRPV1 expression was verified by immunofluorescence staining and Western blotting. The levels of PD-L1, TRPV1 and CGRP were measured via Western blotting, RT-PCR, and immunofluorescence staining. The levels of TNF-α and IL-1β were measured via RT-PCR.
RESULTS
TRPV1 and CGRP protein and mRNA levels were higher in the spinal cords of control female mice than in those of control male mice. PTX-induced nociceptive behaviors in female PIPN mice were greater than those in male PIPN mice, as indicated by increased expression of TRPV1 and CGRP. The analgesic effects of PD-L1 on mechanical hyperalgesia and thermal sensitivity were significantly greater in female mice than in male mice, with calculated relative therapeutic levels increasing by approximately 2.717-fold and 2.303-fold, respectively. PD-L1 and CGRP were partly co-localized with TRPV1 in the dorsal horn of the mouse spinal cord. The analgesic effect of PD-L1 in PIPN mice was observed to be mediated through the downregulation of TRPV1 and CGRP expression following AAV9-mediated spinal cord specific decreased TRPV1 expression.
CONCLUSIONS
PTX-induced nociceptive behaviors and the analgesic effect of PD-L1 in PIPN mice were sexually dimorphic, highlighting the significance of incorporating sex as a crucial biological factor in forthcoming mechanistic studies of PIPN and providing insights for potential sex-specific therapeutic approaches.
Topics: Animals; Paclitaxel; Male; Female; Mice; Calcitonin Gene-Related Peptide; Mice, Inbred C57BL; TRPV Cation Channels; B7-H1 Antigen; Sex Characteristics; Peripheral Nervous System Diseases; Antineoplastic Agents, Phytogenic; Spinal Cord; Hyperalgesia
PubMed: 38961264
DOI: 10.1111/cns.14829 -
Communications Biology Jul 2024Precise epitope determination of therapeutic antibodies is of great value as it allows for further comprehension of mechanism of action, therapeutic responsiveness...
Precise epitope determination of therapeutic antibodies is of great value as it allows for further comprehension of mechanism of action, therapeutic responsiveness prediction, avoidance of unwanted cross reactivity, and vaccine design. The golden standard for discontinuous epitope determination is the laborious X-ray crystallography method. Here, we present a combinatorial method for rapid mapping of discontinuous epitopes by mammalian antigen display, eliminating the need for protein expression and purification. The method is facilitated by automated workflows and tailored software for antigen analysis and oligonucleotide design. These oligos are used in automated mutagenesis to generate an antigen receptor library displayed on mammalian cells for direct binding analysis by flow cytometry. Through automated analysis of 33930 primers an optimized single condition cloning reaction was defined allowing for mutation of all surface-exposed residues of the receptor binding domain of SARS-CoV-2. All variants were functionally expressed, and two reference binders validated the method. Furthermore, epitopes of three novel therapeutic antibodies were successfully determined followed by evaluation of binding also towards SARS-CoV-2 Omicron BA.2. We find the method to be highly relevant for rapid construction of antigen libraries and determination of antibody epitopes, especially for the development of therapeutic interventions against novel pathogens.
Topics: Humans; SARS-CoV-2; Epitope Mapping; Epitopes; Spike Glycoprotein, Coronavirus; COVID-19; Peptide Library; Antibodies, Viral; Animals; HEK293 Cells; Cell Surface Display Techniques; Gene Library
PubMed: 38961245
DOI: 10.1038/s42003-024-06508-8 -
Scientific Reports Jul 2024Blast wave exposure, a leading cause of hearing loss and balance dysfunction among military personnel, arises primarily from direct mechanical damage to the...
Blast wave exposure, a leading cause of hearing loss and balance dysfunction among military personnel, arises primarily from direct mechanical damage to the mechanosensory hair cells and supporting structures or indirectly through excessive oxidative stress. We previously reported that HK-2, an orally active, multifunctional redox modulator (MFRM), was highly effective in reducing both hearing loss and hair cells loss in rats exposed to a moderate intensity workday noise that likely damages the cochlea primarily from oxidative stress versus direct mechanical trauma. To determine if HK-2 could also protect cochlear and vestibular cells from damage caused primarily from direct blast-induced mechanical trauma versus oxidative stress, we exposed rats to six blasts of 186 dB peak SPL. The rats were divided into four groups: (B) blast alone, (BEP) blast plus earplugs, (BHK-2) blast plus HK-2 and (BEPHK-2) blast plus earplugs plus HK-2. HK-2 was orally administered at 50 mg/kg/d from 7-days before to 30-day after the blast exposure. Cochlear and vestibular tissues were harvested 60-d post-exposure and evaluated for loss of outer hair cells (OHC), inner hair cells (IHC), auditory nerve fibers (ANF), spiral ganglion neurons (SGN) and vestibular hair cells in the saccule, utricle and semicircular canals. In the untreated blast-exposed group (B), massive losses occurred to OHC, IHC, ANF, SGN and only the vestibular hair cells in the striola region of the saccule. In contrast, rats treated with HK-2 (BHK-2) sustained significantly less OHC (67%) and IHC (57%) loss compared to the B group. OHC and IHC losses were smallest in the BEPHK-2 group, but not significantly different from the BEP group indicating lack of protective synergy between EP and HK-2. There was no loss of ANF, SGN or saccular hair cells in the BHK-2, BEP and BEPHK-2 groups. Thus, HK-2 not only significantly reduced OHC and IHC damage, but completely prevented loss of ANF, SGN and saccule hair cells. The powerful protective effects of this oral MFRM make HK-2 an extremely promising candidate for human clinical trials.
Topics: Animals; Spiral Ganglion; Rats; Blast Injuries; Hair Cells, Vestibular; Male; Oxidation-Reduction; Rats, Sprague-Dawley; Cochlea; Hair Cells, Auditory; Oxidative Stress; Hearing Loss, Noise-Induced
PubMed: 38961203
DOI: 10.1038/s41598-024-66406-1 -
Scientific Reports Jul 2024Inflammation, corticosteroids, and loading all affect tendon healing, with an interaction between them. However, underlying mechanisms behind the effect of...
Inflammation, corticosteroids, and loading all affect tendon healing, with an interaction between them. However, underlying mechanisms behind the effect of corticosteroids and the interaction with loading remain unclear. The aim of this study was to investigate the role of dexamethasone during tendon healing, including specific effects on tendon cells. Rats (n = 36) were randomized to heavy loading or mild loading, the Achilles tendon was transected, and animals were treated with dexamethasone or saline. Gene and protein analyses of the healing tendon were performed for extracellular matrix-, inflammation-, and tendon cell markers. We further tested specific effects of dexamethasone on tendon cells in vitro. Dexamethasone increased mRNA levels of S100A4 and decreased levels of ACTA2/α-SMA, irrespective of load level. Heavy loading + dexamethasone reduced mRNA levels of FN1 and TenC (p < 0.05), while resolution-related genes were unaltered (p > 0.05). In contrast, mild loading + dexamethasone increased mRNA levels of resolution-related genes ANXA1, MRC1, PDPN, and PTGES (p < 0.03). Altered protein levels were confirmed in tendons with mild loading. Dexamethasone treatment in vitro prevented tendon construct formation, increased mRNA levels of S100A4 and decreased levels of SCX and collagens. Dexamethasone during tendon healing appears to act through immunomodulation by promoting resolution, but also through an effect on tendon cells.
Topics: Dexamethasone; Animals; Rats; Wound Healing; Tendon Injuries; Achilles Tendon; S100 Calcium-Binding Protein A4; Male; Annexin A1; Actins; Collagen; Rats, Sprague-Dawley; Tendons; Extracellular Matrix; RNA, Messenger; Basic Helix-Loop-Helix Transcription Factors
PubMed: 38961188
DOI: 10.1038/s41598-024-66038-5 -
Nature Communications Jul 2024
PubMed: 38961169
DOI: 10.1038/s41467-024-49969-5 -
Scientific Reports Jul 2024Although stem cell-based regenerative medicine has been extensively studied, it remains difficult to reconstruct three dimensional tissues and organs in combination with...
Although stem cell-based regenerative medicine has been extensively studied, it remains difficult to reconstruct three dimensional tissues and organs in combination with vascular systems in vitro. One clinically successful therapy is transplantation of mesenchymal stem cells (MSC) into patients with graft versus host disease. However, transplanted cells are immediately damaged and destroyed because of innate immune reactions provoked by thrombogenic inflammation, and patients need to take immunosuppressive drugs for the immunological regulation of allogeneic cells. This reduces the benefits of stem cell transplantation. Therefore, alternative therapies are more realistic options for clinical use. In this study, we aimed to take advantage of the therapeutic efficacy of MSC and use multiple cytokines released from MSC, that is, stem cells from human exfoliated deciduous teeth (SHEDs). Here, we purified components from conditioned media of immortalized SHED (IM-SHED-CM) and evaluated the activities of intracellular dehydrogenase, cell migration, and antioxidative stress by studying the cells. The immortalization of SHED could make the stable supply of CM possible. We found that the fractionated component of 50-100 kD from IM-SHED-CM had higher efficacy than the original IM-SHED-CM in terms of intracellular dehydrogenase and cell migration in which intracellular signal transduction was activated via receptor tyrosine kinases, and the glutathione peroxidase and reductase system was highly active. Although antioxidative stress activities in the fractionated component of 50-100 kD had slightly lower than that of original IM-SHE-CM, the fraction still had the activity. Thus, the use of fractionated components of 50-100 kD from IM-SHED-CM could be an alternative choice for MSC transplantation because the purified components from CM could maintain the effect of cytokines from SHED.
Topics: Humans; Tooth, Deciduous; Cell Movement; Oxidative Stress; Mesenchymal Stem Cells; Culture Media, Conditioned; Cells, Cultured; Antioxidants; Stem Cells; Signal Transduction
PubMed: 38961142
DOI: 10.1038/s41598-024-66213-8 -
Scientific Reports Jul 2024Cadmium (Cd) pollution is a serious threat to food safety and human health. Minimizing Cd uptake and enhancing Cd tolerance in plants are vital to improve crop yield and...
Cadmium (Cd) pollution is a serious threat to food safety and human health. Minimizing Cd uptake and enhancing Cd tolerance in plants are vital to improve crop yield and reduce hazardous effects to humans. In this study, we designed three Cd concentration stress treatments (Cd1: 0.20 mg·kg, Cd2: 0.60 mg·kg, and Cd3: 1.60 mg·kg) and two foliar silicon (Si) treatments (CK: no spraying of any material, and Si: foliar Si spraying) to conduct pot experiments on soil Cd stress. The results showed that spraying Si on the leaves reduced the Cd content in brown rice by 4.79-42.14%. Si application increased net photosynthetic rate (Pn) by 1.77-4.08%, stomatal conductance (Gs) by 5.27-23.43%, transpiration rate (Tr) by 2.99-20.50% and intercellular carbon dioxide (CO) concentration (Ci) by 6.55-8.84%. Foliar spraying of Si significantly increased the activities of superoxide dismutase (SOD) and peroxidase (POD) in rice leaves by 9.84-14.09% and 4.69-53.09%, respectively, and reduced the content of malondialdehyde (MDA) by 7.83-48.72%. In summary, foliar Si spraying protects the photosynthesis and antioxidant system of rice canopy leaves, and is an effective method to reduce the Cd content in brown rice.
Topics: Oryza; Cadmium; Photosynthesis; Silicon; Antioxidants; Plant Leaves; Malondialdehyde; Superoxide Dismutase; Soil Pollutants; Peroxidase
PubMed: 38961133
DOI: 10.1038/s41598-024-66204-9 -
Scientific Reports Jul 2024A multicenter study of nonmetastatic castration-resistant prostate cancer (nmCRPC) was conducted to identify the optimal cut-off value of prostate-specific antigen (PSA)...
A multicenter study of nonmetastatic castration-resistant prostate cancer (nmCRPC) was conducted to identify the optimal cut-off value of prostate-specific antigen (PSA) doubling time (PSADT) that correlated with the prognosis in Japanese nmCRPC. Of the 515 patients diagnosed and treated for nmCRPC at 25 participating Japanese Urological Oncology Group centers, 450 patients with complete clinical information were included. The prognostic values of clinical factors were evaluated with respect to prostate specific antigen progression-free (PFS), cancer-specific survival (CSS), and overall survival (OS). The optimal cutoff value of PSADT was identified using survival tree analysis by Python. The Median PSA and PSADT at diagnosis of nmCRPC were 3.3 ng/ml, and 5.2 months, respectively. Patients treated with novel hormonal therapy (NHT) showed significantly longer PFS (HR: hazard ratio 0.38, p < 0.0001) and PFS2 (HR 0.45, p < 0.0001) than those treated with vintage nonsteroidal antiandrogen agent (Vintage). The survival tree identified 4.65 months as the most prognostic PSADT cutoff point. Among the clinical and pathological factors PSADT of < 4.65 months remained an independent prognostic factor for OS (HR 2.96, p = 0.0003) and CSS (HR 3.66, p < 0.0001). Current data represented optimal cut-off of PSADT 4.65 months for a Japanese nmCRPC.
Topics: Humans; Male; Prostate-Specific Antigen; Prostatic Neoplasms, Castration-Resistant; Aged; Middle Aged; Japan; Prognosis; Aged, 80 and over; East Asian People
PubMed: 38961131
DOI: 10.1038/s41598-024-65969-3 -
Communications Biology Jul 2024Oral administration of harmless antigens can induce suppression of reactive immune responses, a process that capitalises on the ability of the gastrointestinal tract to...
Oral administration of harmless antigens can induce suppression of reactive immune responses, a process that capitalises on the ability of the gastrointestinal tract to tolerate exposure to food and commensal microbiome without triggering inflammatory responses. Repeating exposure to type II collagen induces oral tolerance and inhibits induction of arthritis, a chronic inflammatory joint condition. Although some mechanisms underlying oral tolerance are described, how dysregulation of gut immune networks impacts on inflammation of distant tissues like the joints is unclear. We used undenatured type II collagen in a prophylactic regime -7.33 mg/kg three times/week- to describe the mechanisms associated with protective oral immune-therapy (OIT) in gut and joint during experimental Collagen-Induced Arthritis (CIA). OIT reduced disease incidence to 50%, with reduced expression of IL-17 and IL-22 in the joints of asymptomatic mice. Moreover, whilst the gut tissue of arthritic mice shows substantial damage and activation of tissue-specific immune networks, oral administration of undenatured type II collagen protects against gut pathology in all mice, symptomatic and asymptomatic, rewiring IL-17/IL-22 networks. Furthermore, gut fucosylation and microbiome composition were also modulated. These results corroborate the relevance of the gut-joint axis in arthritis, showing novel regulatory mechanisms linked to therapeutic OIT in joint disease.
Topics: Animals; Arthritis, Experimental; Collagen Type II; Mice; Gastrointestinal Microbiome; Homeostasis; Male; Joints; Mice, Inbred DBA; Interleukin-17; Interleukin-22; Administration, Oral
PubMed: 38961129
DOI: 10.1038/s42003-024-06476-z