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Experimental Neurology Jul 2024Chronic hypoxia in utero causes intrauterine growth restriction (IUGR) of the fetus. IUGR infants are known to be at higher risk for neurodevelopmental disorders, but...
Chronic hypoxia in utero causes intrauterine growth restriction (IUGR) of the fetus. IUGR infants are known to be at higher risk for neurodevelopmental disorders, but the mechanism is unclear. In this study, we analyzed the structure of the cerebral cortex using IUGR model rats generated through a reduced uterine perfusion pressure operation. IUGR rats exhibited thinner cerebral white matter and enlarged lateral ventricles compared with control rats. Expression of neuron cell markers, Satb2, microtubule-associated protein (MAP)-2, α-tubulin, and nestin was reduced in IUGR rats, indicating that neurons were diminished at various developmental stages in IUGR rats, from neural stem cells to mature neurons. However, there was no increase in apoptosis in IUGR rats. Cells positive for Ki67, a marker of cell proliferation, were reduced in neurons and all glial cells of IUGR rats. In primary neuron cultures, axonal elongation was impaired under hypoxic culture conditions mimicking the intrauterine environment of IUGR infants. Thus, in IUGR rats, chronic hypoxia in utero suppresses the proliferation of neurons and glial cells as well as axonal elongation, resulting in cortical thinning and enlarged lateral ventricles. Thrombopoietin (TPO), a platelet growth factor, inhibited the decrease in neuron number and promoted axon elongation in primary neurons under hypoxic conditions. Intraperitoneal administration of TPO to IUGR rats resulted in increases in the number of NeuN-positive cells and the area coverage of Satb2. In conclusion, suppression of neuronal proliferation and axonal outgrowth in IUGR rats resulted in cortical thinning and enlargement of lateral ventricles. TPO administration might be a novel therapeutic strategy for treating brain dysmaturation in IUGR infants.
Topics: Animals; Fetal Growth Retardation; Rats; Neurons; Female; Cell Proliferation; Pregnancy; Thrombopoietin; Neuronal Outgrowth; Neuroprotective Agents; Rats, Sprague-Dawley; Cells, Cultured; Animals, Newborn; Cerebral Cortex
PubMed: 38636773
DOI: 10.1016/j.expneurol.2024.114781 -
Neurochemical Research Mar 2024Therapeutic hypothermia (TH) provides neuroprotection. However, the cellular mechanisms underlying the neuroprotective effects of TH are not fully elucidated. Regulation...
Therapeutic hypothermia (TH) provides neuroprotection. However, the cellular mechanisms underlying the neuroprotective effects of TH are not fully elucidated. Regulation of microglial activation has the potential to treat a variety of nervous system diseases. Transient receptor potential vanilloid 4 (TRPV4), a nonselective cation channel, is activated by temperature stimulus at 27-35 °C. Although it is speculated that TRPV4 is associated with the neuroprotective mechanisms of TH, the role of TRPV4 in the neuroprotective effects of TH is not well understood. In the present study, we investigated whether hypothermia attenuates microglial activation via TRPV4 channels. Cultured microglia were incubated under normothermic (37 °C) or hypothermic (33.5 °C) conditions following lipopolysaccharide (LPS) stimulation. Hypothermic conditions suppressed the expression of pro-inflammatory cytokines, inducible nitric oxide synthase, and the number of phagocytic microglia. AMP-activated protein kinase (AMPK)-NF-κB signaling was inhibited under hypothermic conditions. Furthermore, hypothermia reduced neuronal damage induced by LPS-treated microglial cells. Treatment with TRPV4 antagonist in normothermic culture replicated the suppressive effects of hypothermia on microglial activation and microglia-induced neuronal damage. In contrast, treatment with a TRPV4 agonist in hypothermic culture reversed the suppressive effect of hypothermia. These findings suggest that TH suppresses microglial activation and microglia-induced neuronal damage via the TRPV4-AMPK-NF-κB pathway. Although more validation is needed to consider differences according to age, sex, and specific central nervous system regions, our findings may offer a novel therapeutic approach to complement TH.
Topics: Humans; NF-kappa B; Microglia; TRPV Cation Channels; Neuroprotective Agents; Hypothermia; Lipopolysaccharides; AMP-Activated Protein Kinases; Antineoplastic Agents; Nitric Oxide
PubMed: 38112974
DOI: 10.1007/s11064-023-04075-8 -
Psychopharmacology Jan 2024Clozapine N-oxide (CNO) has been developed as a ligand to selectively activate designer receptors exclusively activated by designer drugs (DREADDs). However, previous...
RATIONALE
Clozapine N-oxide (CNO) has been developed as a ligand to selectively activate designer receptors exclusively activated by designer drugs (DREADDs). However, previous studies have revealed that peripherally injected CNO is reverse-metabolized into clozapine, which, in addition to activating DREADDs, acts as an antagonist at various neurotransmitter receptors, suggesting potential off-target effects of CNO on animal physiology and behaviors. Recently, second-generation DREADD agonists compound 21 (C21) and JHU37160 (J60) have been developed, but their off-target effects are not fully understood.
OBJECTIVES
The present studies assessed the effect of novel DREADD ligands on reward-seeking behavior.
METHODS
We first tested the possible effect of acute i.p. injection of low-to-moderate (0.1, 0.3, 1, 3 mg/kg) of CNO, C21, and J60 on motivated reward-seeking behavior in wild-type mice. We then examined whether a high dose (10 mg/kg) of these drugs might be able to alter responding.
RESULTS
Low-to-moderate doses of all drugs and a high dose of CNO or C21 did not alter operant lick responding for a reward under a progressive ratio schedule of reinforcement, in which the number of operant lick responses to obtain a reward increases after each reward collection. However, high-dose J60 resulted in a total lack of responding that was later observed in an open field arena to be due to a sedative effect.
CONCLUSIONS
This study provides definitive evidence that commonly used doses of CNO, C21, and J60 have negligible off-target effects on motivated reward-seeking but urges caution when using high doses of J60 due to sedative effects.
Topics: Mice; Animals; Clozapine; Reward; Designer Drugs
PubMed: 37792024
DOI: 10.1007/s00213-023-06465-w -
Experimental Cell Research Nov 2023Atherosclerosis is a persistent inflammatory state that contributes significantly to cardiovascular disease, a primary cause of mortality worldwide. Enhanced lipid...
Atherosclerosis is a persistent inflammatory state that contributes significantly to cardiovascular disease, a primary cause of mortality worldwide. Enhanced lipid uptake by macrophages and their transformation into foam cells play a key role in the development of atherosclerosis. Recent studies using in vivo mouse models indicated that activation of AMPK has anti-atherosclerotic effects by upregulating the expression of cholesterol efflux transporters in foam cells and promoting cholesterol efflux. However, the pathway downstream of AMPK that contributes to elevated expression of cholesterol efflux transporters remains unclear. In this study, we found that activation of AMPK by AICAR and metformin inhibits foam cell formation via suppression of mTOR in macrophages. Specifically, activation of AMPK indirectly reduced the phosphorylation level of mTOR at Ser2448 and promoted the expression of cholesterol efflux transporters and cholesterol efflux. These inhibitory effects on foam cell formation were counteracted by mTOR activators. Metformin, a more nonspecific AMPK activator than AICAR, appears to inhibit foam cell formation via anti-inflammatory effects in addition to suppression of the mTOR pathway. The results of this study suggest that the development of new drugs targeting AMPK activation and mTOR inhibition may lead to beneficial results in the prevention and treatment of atherosclerosis.
Topics: Animals; Mice; AMP-Activated Protein Kinases; Macrophages; Cholesterol; Foam Cells; TOR Serine-Threonine Kinases; Metformin; Atherosclerosis; ATP Binding Cassette Transporter 1
PubMed: 37730144
DOI: 10.1016/j.yexcr.2023.113784 -
Journal of Pharmacological Sciences Jan 2023Bone remodeling mediated by bone-forming osteoblasts (OBs) and bone-resorbing osteoclasts (OCs) maintains bone structure and function. Excessive OC activation leads to...
Bone remodeling mediated by bone-forming osteoblasts (OBs) and bone-resorbing osteoclasts (OCs) maintains bone structure and function. Excessive OC activation leads to bone-destroying diseases such as osteoporosis and bone erosion of rheumatoid arthritis (RA). Differentiation of OCs from bone marrow cells (BMCs) is regulated by the bone microenvironment. The proinflammatory cytokine interleukin (IL)-1β reportedly enhances osteoclastogenesis and plays important roles in RA-associated bone loss. The present study investigated the effect of IL-1β on OC formation via microenvironmental cells. Treating mouse BMCs with IL-1β in the presence of receptor activator of NF-κB ligand and macrophage colony-stimulating factor increased the number of OCs. Real-time RT-PCR revealed increased expression of the IL-1β, IL-1RI, and IL-1RII genes in non-OCs compared with OCs. Removing CD45 cells which cannot differentiate into OCs, from mouse BMCs reduced the IL-1β-mediated enhancement of osteoclastogenesis. IL-1β treatment upregulated the expression of inducible nitric oxide synthase, insulin-like growth factor 2 (IGF2), and the chemokines stromal cell derived factor 1, C-X3-C motif ligand 1 (CX3CL1), and CXCL7 in non-OCs. Neutralizing antibodies against these chemokines and IGF2 suppressed osteoclastogenesis in the presence of IL-1β. These results suggest that IL-1β enhances osteoclastogenesis by upregulating IGF2 and chemokine expression in non-OCs.
Topics: Mice; Animals; Osteogenesis; Ligands; Cells, Cultured; Osteoclasts; Osteoblasts; Cell Differentiation; RANK Ligand
PubMed: 36522118
DOI: 10.1016/j.jphs.2022.10.007 -
Frontiers in Neuroscience 2022Importin α3 (Gene: , the ortholog of human Importin α4) is a member of the importin α family and participates in nucleocytoplasmic transport by forming trimeric...
Importin α3 (Gene: , the ortholog of human Importin α4) is a member of the importin α family and participates in nucleocytoplasmic transport by forming trimeric complexes between cargo proteins and importin β1. Evidence from human studies has indicated that single nucleotide polymorphisms (SNP) in the gene are associated with the occurrence of several psychiatric disorders accompanied by abnormal reward-related behavior, including schizophrenia, major depression, and substance addiction. However, the precise roles of importin α3 in controlling reward processing and motivation are still unclear. In this study, we evaluated the behavioral effects of knockout (KO) in mice on performance in touchscreen operant chamber-based tasks evaluating simple (fixed-ratio) and effortful (progressive-ratio) reward-seeking behaviors. While KO mice showed no significant differences in operant reward learning on a fixed-ratio schedule, they demonstrated significantly increased motivation (increased break point) to instrumentally respond for sucrose on a progressive-ratio schedule. We additionally measured the number of c-Fos-positive cells, a marker of neural activity, in 20 regions of the brain and identified a network of brain regions based on their interregional correlation coefficients. Network and graph-theoretic analyses suggested that deficiency enhanced overall interregional functional connectivity. These findings suggest the importance of in motivational control and indicate that KO mice may be an attractive line for modeling motivational abnormalities associated with several psychiatric disorders.
PubMed: 35844217
DOI: 10.3389/fnins.2022.905991 -
Journal of Pharmacological Sciences Jul 2022Osteoporosis is caused by an imbalance in bone remodeling due to abnormal osteoclast (OC) formation and activation. Hypoxia at the site of inflammation promotes OC...
Osteoporosis is caused by an imbalance in bone remodeling due to abnormal osteoclast (OC) formation and activation. Hypoxia at the site of inflammation promotes OC formation and activation in various species, including humans. We previously reported that insulin-like growth factor 2 (IGF2) plays an important role in osteoclastogenesis under hypoxia. In our present study, we focused on the mechanism of osteoclastogenesis in regard to IGF2 signaling under hypoxia. We confirmed that the addition of IGF2 promoted osteoclastogenesis under normoxic conditions. Conversely, IGF2-neutralizing antibodies inhibited osteoclastogenesis under both normoxic and hypoxic conditions. IGF2 addition increased levels of phosphorylated Akt (Thr308 and Ser473) and NF-κB (Ser536), indicating activation of the Akt-NF-κB pathway. IGF2 also increased the expression of inducible nitric oxide synthase, which promotes osteoclastogenesis via nitric oxide production. Expression levels of genes encoding inflammatory cytokines, such as tumor necrosis factor-α, interleukin (IL)-1β, and IL-6, were upregulated, indicating that IGF2 promotes osteoclastogenesis by increasing the expression of inflammatory cytokines via activation of the Akt-NF-κB pathway. These results suggest that IGF2 is a promising therapeutic target for osteoporosis and rheumatoid arthritis.
Topics: Cytokines; Humans; Hypoxia; Insulin-Like Growth Factor II; NF-kappa B; Osteogenesis; Osteoporosis; Proto-Oncogene Proteins c-akt
PubMed: 35641033
DOI: 10.1016/j.jphs.2022.03.007 -
Medical Oncology (Northwood, London,... Apr 2022Photodynamic therapy (PDT) damages cancer cells via photosensitization using harmless laser irradiation. We synthesized a new photosensitizer, mannose-conjugated-chlorin...
Photodynamic therapy (PDT) damages cancer cells via photosensitization using harmless laser irradiation. We synthesized a new photosensitizer, mannose-conjugated-chlorin e6 (M-chlorin e6), which targets mannose receptors that are highly expressed on M2-like tumor-associated macrophages (M2-TAMs) and cancer cells. In our previous study, we demonstrated that M-chlorin e6 PDT reduces tumor volume and decreases the proportion of M2-TAMs. Whether M-chlorin e6 PDT-treated cancer cells activate tumor immunity remains unclear, although the decrease in M2-TAMs is thought to be a direct injurious effect of M-chlorin e6 PDT. Calreticulin (CRT) is exposed at the surface of the membrane of cancer cells in response to treatment with chemotherapeutic agents such as anthracycline and oxaliplatin. Surface-exposed CRT induces phagocytosis of CRT receptor-positive cells, including macrophages, inducing anticancer immune responses. In the present study, we found that M-chlorin e6 PDT increases CRT on the surface of cancer cells, leading to macrophage phagocytosis of cancer cells. Furthermore, M-chlorin e6 PDT increases CD80CD86 macrophages. These results suggest that M-chlorin e6 PDT exerts anti-tumor effects by both enhancing the phagocytosis of cancer cells and strengthening the anti-tumor phenotype of macrophages.
Topics: Calreticulin; Chlorophyllides; Humans; Macrophages; Mannose; Neoplasms; Phagocytosis; Photochemotherapy
PubMed: 35478050
DOI: 10.1007/s12032-022-01674-3 -
The American Journal of Pathology Dec 2021Bone homeostasis depends on the balance between bone resorption by osteoclasts (OCs) and bone formation by osteoblasts. Bone resorption can become excessive under...
Bone homeostasis depends on the balance between bone resorption by osteoclasts (OCs) and bone formation by osteoblasts. Bone resorption can become excessive under various pathologic conditions, including rheumatoid arthritis. Previous studies have shown that OC formation is promoted under hypoxia. However, the precise mechanisms behind OC formation under hypoxia have not been elucidated. The present study investigated the role of inducible nitric oxide synthase (iNOS) in OC differentiation under hypoxia. Primary bone marrow cells obtained from mice were stimulated with receptor activator of NF-κB ligand and macrophage colony-stimulating factor to induce OC differentiation. The number of OCs increased in culture under hypoxia (oxygen concentration, 5%) compared with that under normoxia (oxygen concentration, 20%). iNOS gene and protein expression increased in culture under hypoxia. Addition of an iNOS inhibitor under hypoxic conditions suppressed osteoclastogenesis. Addition of a nitric oxide donor to the normoxic culture promoted osteoclastogenesis. Furthermore, insulin-like growth factor 2 expression was significantly altered in both iNOS inhibition experiments and nitric oxide donor experiments. These data might provide clues to therapies for excessive osteoclastogenesis under several hypoxic pathologic conditions, including rheumatoid arthritis.
Topics: Animals; Bone Resorption; Cell Differentiation; Cell Hypoxia; Cells, Cultured; Enzyme Induction; Hypoxia; Male; Mice; Mice, Inbred C57BL; Nitric Oxide Synthase Type II; Osteoclasts; Osteogenesis; Oxygen; Signal Transduction; omega-N-Methylarginine
PubMed: 34560064
DOI: 10.1016/j.ajpath.2021.08.014 -
Translational Oncology Feb 2021M2-like tumor-associated macrophages (M2-TAMs) in cancer tissues are intimately involved in cancer immunosuppression in addition to growth, invasion, angiogenesis, and...
M2-like tumor-associated macrophages (M2-TAMs) in cancer tissues are intimately involved in cancer immunosuppression in addition to growth, invasion, angiogenesis, and metastasis. Hence, considerable attention has been focused on cancer immunotherapies targeting M2-TAMs. However, systemic therapies inhibit TAMs as well as other macrophages important for normal immune responses throughout the body. To stimulate tumor immunity with fewer side effects, we targeted M2-TAMs using photodynamic therapy (PDT), which damages cells via a nontoxic photosensitizer with harmless laser irradiation. We synthesized a light-sensitive compound, mannose-conjugated chlorin e6 (M-chlorin e6), which targets mannose receptors highly expressed on M2-TAMs. M-chlorin e6 accumulated more in tumor tissue than normal skin tissue of syngeneic model mice and was more rapidly excreted than the second-generation photosensitizer talaporfin sodium. Furthermore, M-chlorin e6 PDT significantly reduced the volume and weight of tumor tissue. Flow cytometric analysis revealed that M-chlorin e6 PDT decreased the proportion of M2-TAMs and increased that of anti-tumor macrophages, M1-like TAMs. M-chlorin e6 PDT also directly damaged and killed cancer cells in vitro. Our data indicate that M-chlorin e6 is a promising new therapeutic agent for cancer PDT.
PubMed: 33401079
DOI: 10.1016/j.tranon.2020.101005