CREB Research Articles

Malignant epithelioid tumors with EWSR1::CREB fusion involving the kidney: a report of two cases.

Soft tissue tumors with EWSR1/FUS fusion to genes encoding the cyclic adenosine monophosphate response element-binding (CREB) transcription factor family (ATF1, CREB1, and CREM) are rare and heterogeneous aggressive tumors, often found in the peritoneal cavity. Here, we report two cases of malignant epithelioid tumors with EWSR1::CREB fusion involving the kidney in females in their 30s. Both tumors appeared as solitary masses, measuring 5.4cm and 4.0cm in diameter. Histologically, the tumors were similar, growing invasively with unclear boundaries and composed of epithelial cells with eosinophilic and clear cytoplasm arranged in sheets, nests, and trabeculae. Immunohistochemically, case 1 showed focal AE1/AE3 positivity, whereas case 2 was negative. Anaplastic lymphoma kinase was diffusely positive in case 1 and focally positive in case 2. Both cases were positive for epithelial membrane antigen, mucin-4, and synaptophysin. High-throughput sequencing identified EWSR1::CREM fusion in case 1, whereas fluorescence in situ hybridization detected EWSR1::CREB1 fusion in case 2. These cases expand the morphological and immunophenotypic characteristics of malignant epithelioid tumors with EWSR1::CREB fusion, highlighting the diagnostic challenges of immunohistochemistry and value of molecular testing for accurate diagnosis.

Reassessing the involvement of the CREB pathway in the circadian clock of Drosophila melanogaster

Circadian clocks are ubiquitous in almost all organisms on Earth and many key genes are highly conserved among species. In the mammalian suprachiasmatic nucleus, the cAMP response element binding protein (CREB) pathway is known to play a crucial role in conveying light-input to the transcription of clock genes. The fruit fly Drosophila melanogaster also expresses two Creb proteins, CrebA and CrebB, which have been associated with the circadian clock. For example, Drosophila Creb has been suggested to constitute a molecular link between neuronal excitability and clock gene transcription. In this study we subjected flies with clock cell specific CrebA or CrebB mutations to circadian behavioral and bioluminescence assays. Surprisingly, we found that neither loss of CrebA or CrebB did affect free-running locomotor behavior, rhythmic period oscillations in clock neurons, or light-dependent synchronization. In conclusion our findings question the conserved circadian role of the Creb pathway in Drosophila and encourage further studies to elucidate its potential function within insect circadian clocks.

GLP-1 Receptor Agonists Alleviate Diabetic Kidney Injury via β-Klotho-Mediated Ferroptosis Inhibition.

Semaglutide (Smg), a GLP-1 receptor agonist (GLP-1RA), shows renal protective effects in patients with diabetic kidney disease (DKD). However, the exact underlying mechanism remains elusive. This study employs transcriptome sequencing and identifies β-Klotho (KLB) as the critical target responsible for the role of Smg in kidney protection. Smg treatment alleviates diabetic kidney injury by inhibiting ferroptosis in patients, animal models, and HK-2 cells. Notably, Smg treatment significantly increases the mRNA expression of KLB through the activation of the cyclic adenosine monophosphate (cAMP) signaling pathway, specifically through the phosphorylation of protein kinase A (PKA) and cAMP-response element-binding protein (CREB). Subsequently, the adenosine monophosphate-activated protein kinase (AMPK) signaling pathway is activated, reprograming the key metabolic processes of ferroptosis such as iron metabolism, fatty acid synthesis, and the antioxidant response against lipid peroxidation. Suppression of ferroptosis by Smg further attenuates renal inflammation and fibrosis. This work highlights the potential of GLP-1RAs and KLB targeting as promising therapeutic approaches for DKD management.

Regulation of sleep amount by CRTC1 via transcription ofCrhin mice

The cAMP-response element binding protein (CREB) is required for regulation of daily sleep amount, whereas gain-of-function of CREB-regulated transcription coactivator 1 (CRTC1) causes severe insomnia in mice. However, the physiological functions of CRTCs and their downstream target genes in the regulation of sleep amount remain unclear. Here, we use adult brain chimeric (ABC)-expression/knockout platform for somatic genetics analysis of sleep in adult male mice. ABC-expression of constitutively active mutant CRTC1/2CAin the mouse brain neurons significantly reduces the amount of non-rapid eye movement sleep (NREMS) and/or REMS. Consistent with that SIK3 phosphorylates and inhibits CRTCs, ABC-expression of CRTC1/2/3CArescues the hypersomnia phenotype ofSleepy(Sik3Slp) mice. While ABC-Crtc2KOorCrtc3KOcauses no sleep phenotype, ABC-Crtc1KOor ABC-expression of dominant-negative CRTC (dnCRTC) results in modest reduction of NREMS amount accompanied with elevated NREMS delta power. Moreover, ABC-expression of CRTC1CAor dnCRTC in the excitatory neurons causes bidirectional changes of NREMS/REMS amount and/or NREMS delta power, whereas expression of CRTC1CAin the inhibitory neurons decreases REMS amount and increases NREMS delta power. The ability of CRTC1CAto regulate sleep requires its transactivation domain and CREB-binding domain and is dependent on CREB. Furthermore, we showed that inducible ABC-expression of corticotropin releasing hormone (Crh) and brain-derived neurotrophic factor (Bdnf)–two target genes of CRTCs–significantly reduces daily sleep amount. Notably, ABC-CrhKO, but notBdnfKO, rescues the insomnia phenotype of ABC-CRTC1CAmice. Taken together, these results indicate that CREB-CRTC1 complex regulates daily sleep amount by modulating the transcription ofCrhin the mouse brain neurons.Significance StatementCRTCs function as coactivators for CREB, a transcription factor required for the regulation of daily sleep amount in mice. Here, we found that CRTC1/2/3 function similarly to suppress NREMS and REMS in a CREB-dependent manner. Consistent with that CRTCs are substrates of SIK3 kinase, expression of constitutive active mutant CRTCsCArescue the hypersomnia phenotype ofSleepy(Sik3Slp) mice. Furthermore, we showed that CRTC1 reduces NREMS amount by upregulating the expression of neuropeptide CRH. These results elucidate the mechanism by which CRTCs regulates sleep amount and suggest a potential transcriptional mechanism in the stress-induced sleep/wake regulation.

CREB: A Promising Therapeutic Target for Treating Psychiatric Disorders.

Psychiatric disorders are complex, multifactorial illnesses. It is challenging for us to understand the underlying mechanism of psychiatric disorders. In recent years, the morbidity of psychiatric disorders has increased yearly, causing huge economic losses to the society. Although some progress, such as psychotherapy drugs and electroconvulsive therapy, has been made in the treatment of psychiatric disorders, including depression, anxiety, bipolar disorder, obsessive-compulsive and autism spectrum disorders, antidepressants and psychotropic drugs have the characteristics of negative effects and high rate of relapse. Therefore, researchers continue to seek suitable interventions. cAMP response element binding protein (CREB) belongs to a protein family and is widely distributed in the majority of brain cells that function as a transcription factor. It has been demonstrated that CREB plays an important role in neurogenesis, synaptic plasticity, and neuronal growth. This review provides a 10-year update of the 2013 systematic review on the multidimensional roles of CREB-mediated transcriptional signaling in psychiatric disorders. We also summarize the classification of psychiatric disorders and elucidate the involvement of CREB and related downstream signalling pathways in psychiatric disorders. Importantly, we analyse the CREB-related signal pathways involving antidepressants and antipsychotics to relieve the pathological process of psychiatric disorders. This review emphasizes that CREB signalling may have a vast potential to treat psychiatric disorders like depression. Furthermore, it would be helpful for the development of potential medicine to make up for the imperfection of current antidepressants and antipsychotics.

Upregulation of cAMP/PKA/CREB signaling mediated by GABABR might contribute to the amelioration effects of sea cucumber peptides (Acaudina leucoprocta) on cognitive dysfunction in mice

This study aimed to determine the effects and underlying mechanisms of sea cucumber peptides (SCP) obtained from Acaudina leucoprocta in mitigating induced by D-galactose cognitive dysfunction in mice. Additionally, it aimed to validate the cognitive-enhancing potential of peptides identified via in silico and peptidomics using SH-SY5Y cells. The results demonstrated significant improvement in behavioral performance with oral administration of SCP. Additionally, SCP normalized oxidative stress and neurotransmitter imbalances (especially gamma-aminobutyric acid, GABA). Also, SCP effectively rectified gut microbiota dysbiosis. Mechanistically, SCP activated the γ-aminobutyric acid type B receptors (GABABR)/ cyclic adenosine monophosphate (cAMP)/ cAMP-dependent protein kinase A (PKA)/ cAMP response element-binding protein (CREB) axis to promote GABA release and alleviate neuronal and oxidative stress damage, which subsequently ameliorated cognitive dysfunction. Furthermore, docking analysis predicted the binding of fourteen peptides with high abundance and PeptideRanker scores to GABABR. Notably, Lys-Gly-Phe-Pro (KGFP) had the strongest binding affinities and actively interacted with GABABR sites through hydrogen bonds, van der Waals, hydrophobic, and electrostatic interactions. The capacity of KGFP to improve cognitive function was further demonstrated when the SH-SY5Y cells were subjected to D-galactose. These results imply that the implementation of SCP intervention could be a useful tactic for improving cognitive function.

Biochemical investigation and in silico analysis of the therapeutic efficacy of Ipriflavone through Tet-1 Surface-Modified-PLGA nanoparticles in Streptozotocin-Induced Alzheimer’s like Disease: Reduced oxidative damage and etiological Descriptors

Ipriflavone (IPRI), an isoflavone derivative, is clinically used to prevent postmenopausal bone loss in addition to its antioxidant and cognitive benefits. However, its poor aqueous solubility retained its bioavailability. New strategies have been developed to improve the bioavailability and solubility of neurological medications to enhance their potency and limit adverse effects. This study aimed to prepare targeted IPRI-poly-lactic-co-glycolic acid (PLGA) nanoparticles coupled with Tet-1 peptide to increase the therapeutic potency of IPRI in a rat model of Alzheimer’s disease (AD). Streptozotocin (STZ) exacerbates Alzheimer-related alterations by promoting central insulin resistance resulted from defective signaling pathways related to neuroinflammation and neurotoxicity. Bilateral intracerebroventricular (icv) injection of STZ was used to introduce the AD model. Icv-STZ injection significantly affected brain insulin, oxidative stress, inflammatory, and apoptotic indicators and caused behavioral abnormalities. STZ promoted the formation of amyloid β42 (Aβ42) by increasing BACE1 and reducing ADAM10 and ADAM17 expression levels. STZ also triggered the accumulation of neurofibrillary tangles and synaptic dysfunction, which are crucial for neurological impairments. Icv-STZ injection showed evident degenerative changes in the pyramidal cell layer and significantly reduced the count of viable cells in both CA1 and prefrontal cortex, indicating increased neuronal cell death. IPRI successfully ameliorated cognitive dysfunction by improving the phosphorylated forms of cAMP-response element-binding protein (pCREB) and extracellular signal-regulated kinase 1/2 (pERK1/2) related to synaptic plasticity. Targeted IPRI nanoparticles exceeded free IPRI potential in reducing oxidative stress, acetylcholinesterase/monoamine oxidase activities, Tau phosphorylation, and Aβ42 levels revealing less degenerative changes and increased viable neuron counts. IPRI-targeted nanoparticles improved the neuroprotective potential of free IPRI, making this strategy applicable to treat many neurodegenerative diseases. Finally, the in silico study predicted its ability to cross the BBB and to bind various protein targets in the brain.

Exploring the Anti-melanogenic, Antioxidant, and Anti-inflammatory Activities of A Composition: Glabridin, Resveratrol and Ellagic Acid

Background: Plant extracts have wide applications in food, nutrition, and cosmetics, which results in a deeper investigation of natural ingredients. Numerous natural ingredients have been demonstrated to exhibit multiple activities, including antioxidation, anti-inflammation, and anti-melanogenesis. However, their combinations have not been well investigated, which could provide stronger performance with less toxicity and easier applications. Methods: We used B16F10 cells treated with alpha-melanocyte stimulating hormone (αMSH) for melanogenesis-related studies, including cellular melanin content, tyrosinase activity, and gene or protein expression. MTT assay was used to evaluate cell viability. DPPH scavenging activity was measured for antioxidation. Nitric oxide (NO) content was evaluated in lipopolysaccharide (LPS) treated RAW264.7 cells to indicate the performance on anti-inflammation. Results: In this study, six different compounds and their combinations were tested for melanogenesis. The results showed that the combination of glabridin, resveratrol, and ellagic acid (GRE) exhibited the highest efficiency, which was mainly manifested as inhibition of melanin production and tyrosinase activity, higher DPPH scavenging rate, and inhibition of nitric oxide (NO) production. Meanwhile, our results showed that GRE could significantly downregulate the expression of microphthalmia-associated transcription factor (MITF) related genes and proteins and could also inhibit the phosphorylation of cyclic AMP response element-binding protein (CREB), which was the upstream signal of MITF. Conclusion: The results suggest GRE exhibits high efficiency in inhibiting anti-melanogenesis, antioxidation, and anti-inflammation. Furthermore, GRE could downregulate the phosphorylation of the CREB and MITF signal pathway, which provides a theoretical basis for its application in pigmentation disorder disease and cosmetics.

Oxytocin Protects Against Corticosterone-Induced DA Dysfunction: An Involvement of the PKA/CREB Pathway

Chronic stress disrupts dopamine (DA) transmission, adversely affecting mood and contribution to neuropsychiatric disorders like ADHD, autism, schizophrenia, anxiety, depression, and drug addiction. The neuropeptide oxytocin (OXT) plays a key role in social cognition, bonding, attachment, and parenting behaviors. In addition, OXT can modulate the activity of the HPA axis, counteracting the effects of stress, and alleviating fear and anxiety. However, whether OXT can mitigate stress-induced DA dysfunction and the underlying mechanisms remains unclear. This study investigated the neuroprotective effects of OXT on corticosterone (CORT) induced DA dysfunction in the neuroblastoma cell line SH-SY5Y. The results revealed that CORT decreases the levels of intracellular signaling molecules associated with DA function, including phosphorylated tyrosine hydroxylase (pTH), phosphorylated cAMP response element-binding protein (pCREB), and protein kinase A (PKA). Interestingly, pretreatment with OXT mitigated CORT-induced DA dysfunction through its potent PKA activator properties. In addition, the neuroprotective effect of OXT was abolished by atosiban (an OXT receptor antagonist) or H89 (a PKA inhibitor). Our results suggest that OXT protects dopaminergic neuroblastoma cells from CORT-induced DA dysfunction, potentially through the involvement of oxytocin receptors and the PKA/CREB signaling pathway. These findings contribute to the understanding of the neurobiological mechanisms underlying stress resilience and highlight potential pathways for developing targeted treatments that leverage the neuroprotective properties of OXT to address disorders characterized by DA dysregulation and impaired stress responses.

4-Methoxycinnamic acid ameliorates post-traumatic stress disorder-like behavior in mice by antagonizing the CRF type 1 receptor

AimsPosttraumatic stress disorder (PTSD) is a debilitating neuropsychiatric illness caused by traumatic or life-threatening events and manifesting as various symptoms, including intrusive re-experiences of trauma, avoidance behaviors, hyperarousal, and negative changes in perception and mood. Main methodsCurrent monoamine-based medications commonly exhibit limited efficacy and significant side effects, which hamper their clinical utility. To address this unmet need, we explored 4-methoxycinnamic acid (4-MCA) as a potential novel treatment for PTSD in a single prolonged stress (SPS)-induced animal model. Key findingsAdministration of 4-MCA (3 and 10 mg/kg, p.o.) significantly mitigated anxiety-like behaviors, alleviated depression-like behaviors, and improved cognitive function in an SPS-treated PTSD mouse model. Further, 4-MCA treatment effectively rectified the fear extinction deficits in the fear conditioning test. Molecular analyses revealed that 4-MCA normalized the elevated corticotropin-releasing hormone (CRH) levels as well as the phosphorylation of protein kinase A (PKA) and cAMP response element-binding protein (CREB) in the amygdala, a pivotal region for fear memory formation. Co-administration of 4-MCA and the CRFR1 antagonist antalarmin at subeffective doses facilitated fear memory extinction. SignificanceThese findings suggest that 4-MCA alleviates SPS-induced PTSD-like behaviors by regulating the CRH-CRFR1-PKA-CREB signaling pathway in the amygdala, and that 4-MCA may be a potential candidate for future PTSD treatment.

IL-17A is a key regulator of neuroinflammation and neurodevelopment in cognitive impairment induced by sevoflurane

Increasing numbers of animal studies have shown that repeat sevoflurane exposure during developmental stage may lead to long-term cognitive impairment. Nevertheless, the exact pathogenesis remains unclear. Interleukin 17A (IL-17A) has been associated with cognitive decline in various neurological disorders. Here we found that the expression of IL-17A was up-regulated in hippocampus of sevoflurane exposed neonatal mice. Genetic deletion of IL-17A or inhibition of IL-17A improved behavioral function and down-regulated neuroinflammation related genes, interleukin 1β (IL-1β), interleukin 6 (IL-6), Nicotinamide adenine dinucleotide phosphate(NADPH) oxidase 2 (NOX2) and NADPH oxidase 4 (NOX4) in hippocampus of sevoflurane exposed neonatal mice. Moreover, negative regulation of IL-17A/Interleukin 17A receptor(IL-17RA) promoted the extracellular signal-regulated protein kinase (ERK) signaling pathway and nucleation of cyclic adenosine monophosphate (cAMP) response element-binding (CREB) in neurons of cognitive impaired mice. Knockdown of IL-17A in vivo identified neurons-localized IL-17A as a major factor in neuroinflammation and neurodevelopment. Collectively, our results suggested that IL-17A was required for the pathogenesis of neuroinflammatory response and identify IL-17A as a potential therapeutic target for cognitive impairment exposed by general anesthetics during infancy.

Exposure to Radiofrequency Electromagnetic Fields Enhances Melanin Synthesis by Activating the P53 Signaling Pathway in Mel-Ab Melanocytes.

The skin is the largest body organ that can be physiologically affected by exposure to radiofrequency electromagnetic fields (RF-EMFs). We investigated the effect of RF-EMFs on melanogenesis; Mel-Ab melanocytes were exposed to 1760 MHz radiation with a specific absorption rate of 4.0 W/kg for 4 h/day over 4 days. Exposure to the RF-EMF led to skin pigmentation, with a significant increase in melanin production in Mel-Ab melanocytes. The phosphorylation level of cAMP response element binding protein (CREB) and the expression of microphthalmia-associated transcription factor (MITF), which regulate the expression of tyrosinase, were significantly increased in Mel-Ab after RF-EMF exposure. Interestingly, the expression of tyrosinase was significantly increased, but tyrosinase activity was unchanged in the RF-EMF-exposed Mel-Ab cells. Additionally, the expression of p53 and melanocortin 1 receptor (MC1R), which regulate MITF expression, was significantly increased. These results suggest that the RF-EMF induces melanogenesis by increasing phospho-CREB and MITF activity. Importantly, when Mel-Ab cells were incubated at 38 °C, the melanin production and the levels of tyrosinase significantly decreased, indicating that the increase in melanin synthesis by RF-EMF exposure is not due to a thermal effect. In conclusion, RF-EMF exposure induces melanogenesis in Mel-Ab cells through the increased expression of tyrosinase via the activation of MITF or the phosphorylation of CREB, which are initiated by the activation of p53 and MC1R.

Identification of chikusetsusaponin IVa as a novel lysine-specific demethylase 1 inhibitor that ameliorates high fat diet-induced MASLD in mice.

Diet-induced metabolic dysfunction steatotic liver disease (MASLD) is also called as non-alcoholic fatty liver disease (NAFLD) with limited effective strategies available. We previously have shown that chikusetsusaponin IVa (CHS), a dietary saponin from herbs in South American known for their metabolic benefits, mitigates diet-induced diabetes. In this study we investigated the beneficial effects of CHS on MASLD and the underlying mechanisms. MAFLD mouse model was established by the high-fat diet (HFD) for 6 weeks and then were treated with CHS (50 mg·kg-1·d-1, i.g.) for another 8 weeks. By conducting transcriptomic analysis in palmitic acid-treated HepG2 cells and primary hepatocytes as well as lipidomic analysis in liver tissues, we demonstrated that HFD activated the intestinal farnesoid X receptor (FXR) pathway, leading to the release of FGF15/19, which in turn promoted hepatic FXR-SHP binding with cAMP-responsive element-binding protein H (CREBH), thereby inhibiting CREBH-mediated fatty acid oxidation (FAO) and ketogenesis. Intriguingly, we found that CHS improved lipid metabolism in HFD mice by suppressing the enterohepatic crosstalk of FXR-SHP to enhance CREBH transactivation. Among these, lysine-specific demethylase 1 (LSD1)-mediated histone demethylation played a crucial role in lipid metabolic reprogramming. Moreover, we identified LSD1 as a critical cellular target of CHS, directly binding to Lys661 and Tyr761 of LSD1 to inhibit its histone demethylation activity. Our results suggest that targeting intestinal LSD1 with CHS could be a promising strategy for MAFLD treatment, offering new insights into the bioavailability and efficacy of natural products.

Synapse-to-nucleus ERK-CREB transcriptional signaling requires dendrite-to-soma Ca2+propagation mediated by L-type voltage-gated Ca2+ channels.

The cAMP-response element binding protein (CREB) transcription factor controls the expression of the neuronal immediate early genes c-Fos, Arc, and Bdnf and is essential for long-lasting synaptic plasticity underlying learning and memory. Despite this critical role, there is still ongoing debate regarding the synaptic excitation-transcription (E-T) coupling mechanisms mediating CREB activation in the nucleus. Here we employed optical uncaging of glutamate to mimic synaptic excitation of distal dendrites in conjunction with simultaneous imaging of intracellular Ca2+ dynamics and transcriptional reporter gene expression to elucidate CREB E-T coupling mechanisms in hippocampal neurons cultured from both male and female rats. Using this approach, we found that CREB-dependent transcription was engaged following dendritic stimulation of N-methyl, D-aspartate receptors (NMDARs) only when Ca2+ signals propagated to the soma via subsequent activation of L-type voltage-gated Ca2+ channels resulting in activation of Extracellular signal-Regulated Kinase (ERK) MAP kinase signaling to sustain CREB phosphorylation in the nucleus. In contrast, dendrite-restricted Ca2+ signals generated by NMDARs failed to stimulate CREB-dependent transcription. Furthermore, Ca2+-CaM-dependent kinase (CaMK)-mediated signaling pathways that may transiently contribute to CREB-phosphorylation following stimulation were ultimately dispensable for downstream CREB-dependent transcription and c-Fos induction. These findings emphasize the essential role that L-type Ca2+ channels play in rapidly relaying signals over long distances from synapses located on distal dendrites to the nucleus to control gene expression.Significance Statement The transcription factor CREB controls gene expression programs required for long-lasting synaptic plasticity and learning and memory, yet the synapse-to-nucleus signaling mechanisms mediating CREB activation are still unclear. Using glutamate uncaging to mimic synaptic input to dendrites, this study shows that Ca2+ signals propagated to the soma by L-type voltage-gated Ca2+ channels engage the ERK MAP kinase cascade to mediate CREB phosphorylation and CREB-dependent transcription. In contrast, dendrite-restricted Ca2+ signals generated primarily by NMDARs failed to effectively engage this signaling pathway or CREB-dependent transcription. In addition, we found that while ERK and CaMK pathways may both contribute to increased CREB phosphorylation immediately following neuronal stimulation, sustained ERK signaling to CREB was necessary to effectively drive CREB-dependent transcription.

Topical application of 666-15, a potent inhibitor of CREB, alleviates alkali-induced corneal neovascularization

Corneal neovascularization (CNV) is a dynamically regulated process that arises due to a disruption in the equilibrium between pro-angiogenic and anti-angiogenic factors. Various cytokines are released by vascular endothelial cells and macrophages in damaged cornea, ultimately inducing CNV. The cAMP-response element-binding protein (CREB), a nuclear transcription factor, potentially impacts tumor angiogenesis by modulating the secretion of angiogenic proteins. This study aimed to assess the impact of 666-15, a potent inhibitor of CREB, on angiogenesis using human microvascular retinal endothelial cells (HMRECs), RAW 264.7 macrophage cell line and alkali-induce CNV mouse model. In vivo, the topical application of 666-15 (0.05mg/mL) to the alkali-burn corneas led to 45% reduction in CNV. Additionally, in vitro treatment with 666-15 is effective in suppressing the migration, proliferation, and tube formation by HMRECs. Furthermore, treatment with 666-15 resulted in a down-regulation of pro-angiogenic cytokines expression, including VEGF-A, TGF-β1, b-FGF, and MMP-2 but simultaneously increasing anti-angiogenic cytokines expression, such as ADAMTS-1, Thrombospondin-1 (Tsp-1) and Tsp-2, both in alkali-burn corneas and HMRECs. And 666-15 inhibited the recruitment and the cytokines expression (VEGF-A, MMP-2, IL-1β, TNF-α, MCP-1 and MIP-1) of macrophage. Our findings revealed that 666-15 may suppress the function of endothelial cells and angiogenesis by restoring the homeostasis of pro-angiogenic stimuli, suggesting its potential as a therapeutic agent in the treatment of CNV and other angiogenesis-driven diseases.

The Effect of Caffeine Exposure on Sleep Patterns in Zebrafish Larvae and Its Underlying Mechanism.

The effect of caffeine on the behavior and sleep patterns of zebrafish larvae, as well as its underlying mechanisms, has been a topic of great interest. This study aimed to investigate the impact of caffeine on zebrafish larval sleep/wake behavior and the expression of key regulatory genes such as cAMP-response element binding protein (CREB) and adenosine (ADA) in the sleep pathway. To begin, the study determined the optimal dose and duration of caffeine exposure, with the optimal doses found to be 31.25 μM, 62.5 μM, and 120 μM. Similarly, the optimal exposure time was established as no more than 120 h, ensuring a mortality rate of less than 10%. The confirmation of these conditions was achieved through the assessment of angiogenesis and the inflammatory reaction. As a result, the treatment time point of 24 h post-fertilization (hpf) was selected to examine the effects of caffeine on zebrafish larval sleep rhythm (48 h, with a light cycle of 14:10). Furthermore, the study analyzed the expression of clock genes (bmal1a, per1b, per2, per3, cry2), adenosine receptor genes (adora1a, adora1b, adora2aa, adora2ab, adora2b), and key regulatory factors (CREB and ADA). The research confirmed that caffeine could induce sleep pattern disorders, significantly upregulate adenosine receptor genes (adora1a, adora1b, adora2a, adora2ab, adora2b) (p < 0.05), and markedly decrease the total sleep time and sleep efficiency of the larvae. Additionally, the activity of ADA significantly increased during the exposure (p < 0.001), and the tissue-specific expression of CREB was also significantly increased, as assessed by immunofluorescence. Caffeine may regulate circadian clock genes through the ADA/ADORA/CREB pathway. These findings not only enhance our understanding of the effects of caffeine on zebrafish larvae but also provide valuable insights into the potential impact of caffeine on human behavior and sleep.

Diosmin alleviates NLRP3 inflammasome-dependent cellular pyroptosis after stroke through RSK2/CREB pathway

In the context of our previous analyses on the main active ingredients of Jieyudan, a classic formula targeting aphasia in stroke, we further delve into the function and mechanisms of its active ingredient, Diosmin (DM), which may exert neuroprotective effects, in ischemic stroke. Herein, bioinformatics analysis revealed targets of DM and their intersection with differentially expressed genes in ischemic stroke. Middle cerebral artery occlusion (MCAO) rats and oxygen-glucose deprivation (OGD) cells were used to construct in vivo and in vitro models of ischemic stroke. The effects of DM on MCAO rats were assessed by Zea-Longa score, Morris water maze, TTC staining, Nissl staining, immunohistochemistry, and Western blot. At the cellular level, cell counting kit-8 assay and Western blot were carried out to verify the mechanism of DM in ischemic stroke. In vivo, DM decreased neurological deficit score, cerebral infarct volume and neuronal damage, and improved cognitive function in MCAO rats. In vitro, DM increased the viability of OGD-treated cells. In addition, DM down-regulated the expressions of NLR family pyrin domain containing 3 (NLRP3) and pyroptosis-associated proteins, while up-regulating ribosomal protein S6 kinase A3 (RSK2) level and activating cyclic-AMP response element-binding protein (CREB) signaling. Conversely, RSK2 inhibitor LJH685 reduced the viability and promoted pyroptosis-associated protein levels, which also partially reversed the effects of DM in vitro. Collectively, DM plays a therapeutic role in ischemic stroke by inhibiting NLRP3 inflammasome-mediated cellular pyroptosis via the RSK2/CREB pathway.

Ketamine attenuates kidney damage and depression-like behaviors in mice with cisplatin-induced acute kidney injury

Acute kidney injury (AKI) is a serious condition characterized by decreased urine output, often accompanied by psychiatric symptoms like depression. However, there are limited pharmacological treatments available for AKI and its associated depressive symptoms. In this study, we investigated whether cisplatin-induced AKI in mice leads to depression-like behaviors and whether ketamine could alleviate both the kidney injury and these behaviors. Mice with cisplatin-induced AKI exhibited elevated levels of creatinine and urea, kidney damage, increased kidney injury molecule-1 protein, and pathological changes in the liver, colon, and spleen. They also showed depression-like behaviors and reduced expression of synaptic proteins in the prefrontal cortex. Remarkably, a single dose of ketamine significantly reduced these symptoms and pathological changes. Interestingly, the beneficial effects of ketamine on the kidneys, other organs, and depression-like behaviors, were reversed by the tropomyosin receptor kinase B (TrkB) inhibitor ANA-12. Western blot analysis revealed the involvement of the TrkB and ERK (extracellular signal-regulated kinase)-CREB (cAMP response element binding protein) signaling pathway. Additionally, metabolomics analysis indicated that blood metabolites, such as C16-ceramide, may contribute to the effects of ketamine in this model. These findings suggest that cisplatin-induced nephrotoxicity in AKI mice contributes to depression-like behaviors, and ketamine can alleviate both kidney damage and depression-like symptoms by modulating the TrkB and ERK-CREB signaling pathways, as well as altering blood metabolites. However, the role of the kidney-brain axis in these depression-like behaviors remains unclear. Furthermore, ketamine may have therapeutic potential for treating kidney diseases such as AKI, along with associated depressive symptoms.

Validation of selective catalytic BmCBP inhibitors that regulate the Bm30K-24 protein expression in silkworm, Bombyx mori.

The cAMP response element binding protein (CREB)-binding protein (CBP) is a histone acetyltransferase that plays an indispensable role in regulating the acetylation of histone and non-histone proteins. Recently, it has been discovered that chemical inhibitors A485 and C646 can bind to Bombyx mori's CBP (BmCBP) and inhibit its acetyltransferase activity. Notably, the binding ability of A485 with BmCBP showed a very low Kd value of 48 nM by surface plasmon resonance (SPR) test. Further identification showed that both A485 and C646 can decrease the acetylation level of known substrate H3K27 and only 1 μM of A485 can almost completely inhibit the acetylation of H3K27, suggesting that A485 is an effective inhibitor of BmCBP's acetyltransferase activity. Moreover, it was confirmed that A485 could downregulate the expression of acetylated Bm30K-24 protein at a post-translational level through acetylation modification by BmCBP. Additionally, it was found that A485 can downregulate the stability of Bm30K-24 and improve its ubiquitination level, suggesting that the acetylation modification by BmCBP could compete with ubiquitination modification at the same lysine site on Bm30K-24, thereby affecting its protein stability. Here, we predict that A485 may be a potent CBP acetyltransferase inhibitor which could be utilized to inhibit acetyltransferase activity in insects, including silkworms.

A Phase 1b, Dose Escalation/Dose Expansion, Multicenter, Open-Label Study to Assess the Safety and Tolerability of OPN-6602 Monotherapy and in Combination with Dexamethasone in Subjects with Relapsed and/or Refractory Multiple Myeloma

Background and Significance: OPN-6602 is a potent, selective, and orally active small molecule dual cyclic-AMP response element-binding protein (CBP) and E1A binding protein (EP300) bromodomain (BRD) inhibitor. It has low nanomolar potency against EP300 and CBP, with greater than 100-fold less activity against BRD3, which was the most potent among all the other BRDs evaluated. OPN-6602 suppresses known oncogenes interferon regulatory factor 4 (IRF4) with downstream suppression of MYC resulting in antitumor activity through EP300/CBP inhibition. It is a potent inhibitor of multiple myeloma (MM) cell growth in the MM cell lines, OPM-2 and MM.1S, and a potent inhibitor of histone acetylation via BRD intramolecular modulation of histone acetyltransferase (HAT) activity. EP300 and CBP are transcription coactivators that contain both epigenetic writer HAT and reader BRDs that regulate transcription of genes via chromatin remodeling. EP300 and CBP are highly conserved and ubiquitously expressed enzymes that belong to the KAT3 family of acetyltransferases. EP300 and CBP interact with numerous other proteins to function as global transcriptional coactivators through the modification of lysines on both histone and non-histone nuclear proteins, affecting proteins such as IRF4 and MYC. IRF4/MYC act as lineage-specific transcription factors controlling the survival and growth of MM. EP300/CBP inhibition causes cell cycle arrest and apoptosis in MM through suppression of IRF4 and MYC, rendering EP300/CBP as a novel target for MM (Conery 2016). Recently, unbiased analysis of vulnerability genes identified EP300 as a specific and highly relevant target for MM (de Matos Simoes 2023). OPN-6602 pharmacokinetic (PK) studies in mice, rats, and dogs consistently showed moderate clearance and short terminal half-life. In definitive 28-day GLP toxicology studies, OPN-6602 showed near dose-proportional increases in exposure over the dose range evaluated. These PK properties were specifically engineered to enable daily ‘pulsatile’ dosing. The potent selective inhibition of EP300 and CBP can be exploited to address several unmet medical needs through different mechanisms. In particular, a selective inhibitor of EP300 and CBP such as OPN-6602 has the potential to be a promising treatment for MM. Study Design and Methods: Study OPN6602-C01 is a Phase 1b, open-label study evaluating the safety, tolerability, PK, preliminary antitumor activity, and pharmacodynamics of OPN-6602 monotherapy and in combination with dexamethasone in subjects with relapsed and/or refractory MM. OPN-6602 as monotherapy or in combination with dexamethasone will be administered orally once daily in 21-day cycles. This study consists of 3 parts: Part 1a OPN-6602 monotherapy dose escalation (Part 1a), Part 1b OPN-6602 combination with dexamethasone dose escalation (Part 1b), and Part 2 dose expansion of the preferred regimen (Part 2) to select and optimize the dose with 1:1 randomization of at least 2 potential dose levels of 20 subjects at each dose level (ie, the provisional recommended Phase 2 dose [RP2D] identified in Part 1a or Part 1b and 1 additional dose level). Either OPN-6602 monotherapy or OPN-6602 in combination with dexamethasone will be selected for Part 2. Up to 130 total evaluable subjects will be enrolled across dose escalation and dose expansion. The primary objective and endpoint for Part 1a and Part 1b is to characterize the tolerability of OPN-6602 (with and without dexamethasone). Dose limiting toxicities, treatment-emergent adverse events and changes in safety and lab parameters will be characterized in order to identify the RP2D. The primary objective and endpoint for dose expansion of OPN-6602 monotherapy or in combination with dexamethasone is to evaluate the preliminary antitumor activity using overall response rate based on the 2011 International Myeloma Working Group (IMWG) response criteria. Eligibility criteria include a confirmed diagnosis of MM per IMWG criteria, disease that is relapsed or refractory to 3 or more different prior lines of therapy that include immunomodulatory agents, proteasome inhibitors and an anti-CD38 antibody, measurable disease and adequate hematologic, renal and liver function. This multi-center study (NCT06433947) is currently open in the United States with additional sites planned to open in Switzerland, United Kingdom and the European Union.