The use of minor cannabinoids has been advanced, in part, by the idea of providing relief from pain and inflammation without the unwanted psychogenic effects associated with delta-9-tetrahydro-cannabinol (ΔTHC). With a f...The use of minor cannabinoids has been advanced, in part, by the idea of providing relief from pain and inflammation without the unwanted psychogenic effects associated with delta-9-tetrahydro-cannabinol (ΔTHC). With a focus on peripheral nociception, 4 common minor cannabinoids: cannabidiol (CBD), cannabigerol (CBG), cannabinol (CBN), and cannabichromene (CBC) were studied in primary cultures of mouse dorsal root ganglion (DRG) neurons. We queried whether calcium responses induced by the 4 cannabinoids differed in potency of activation, neuronal size preference, and dose-response relationships. Additionally, we determined the dependence of CBD and CBN on key channel receptors that are known to mediate pain and/or antinociception. Individually, CBD, CBG, and CBC directed greater response magnitudes when compared with CBN. All 4 minor cannabinoids activated overlapping size populations of sensory neurons. CBD and CBG activated the widest range of DRG neuron sizes (smaller-larger), overlapping with smaller capsaicin-sensitive neurons. In contrast, CBN and CBC activated predominantly larger sensory neurons. CBD diverged from other minor cannabinoids in directing a linear dose-response profile, whereas CBG and CBC directed sigmoidal profiles and CBN activated DRG neurons with an inverted U-shaped dose-response relationship. CBD-induced activation of DRG neurons was dependent on coexpression of the nociceptive channel transient receptor potential cation channel subfamily Vanilloid member 1 (TRPV1) plus cannabinoid receptor 1 (CBR), whereas CBN-induced activation was independent of TRPV1 and CBR. Overall, we observed that minor cannabinoids CBD, CBG, CBN, and CBC directed unique activation properties across a diverse population of sensory neurons. Such differences underlie the hypothesis that a combination of minor cannabinoids can direct complementary antinociceptive activity. SIGNIFICANCE STATEMENT: Minor cannabinoids CBD, CBG, CBN, and CBC differ in their dose-dependent properties of sensory neuronal activation. CBD-induced activation of small to large sensory neurons was dependent on coexpression of the nociceptive channel TRPV1 plus CBR and directed a linear dose-response profile. In contrast, CBN activated predominantly larger sensory neurons in an inverted U-shaped dose-response profile and was independent of TRPV1 and CBR. A combination of minor cannabinoids is hypothesized to direct complementary antinociceptive activity.
Osteosarcoma (OS) is a very aggressive malignant neoplasm characterized by significant metastases and a deficiency of treatment targets. Therefore, it is urgent to find new therapeutic mechanisms and targets for OS. We a...Osteosarcoma (OS) is a very aggressive malignant neoplasm characterized by significant metastases and a deficiency of treatment targets. Therefore, it is urgent to find new therapeutic mechanisms and targets for OS. We assessed the capability of chromobox homolog 4 (CBX4) to modulate OS growth using in vivo and in vitro dual tests utilizing short hairpin RNA technology, 3-dimensional tumor sphere building, immunofluorescence co-localization, and other biotechnological methods. Our findings indicated that CBX4 was overexpressed in OS tissues, and the silencing of CBX4 markedly reduced OS cell proliferation, migration, and invasion. CBX4 modulates vasculogenic mimicry through its influence on yes1 associated transcriptional regulator (YAP1) and neuropilin-2/vascular endothelial growth factor A (NRP2/VEGFA) pathway, hence promoting angiogenic activity, a mechanism perhaps linked to CBX4's role in regulating mitochondrial autophagy. CBX4 affects vasculogenic mimicry by regulating the YAP1 and NRP2/VEGFA pathways and promotes the OS process. CBX4 may serve as a novel biomarker for OS, potentially offering innovative approaches for early diagnosis and tailored therapy of OS. SIGNIFICANCE STATEMENT: Osteosarcoma (OS) is a very aggressive malignant neoplasm characterized by significant metastases and a deficiency of treatment targets. Therefore, it is urgent to find new therapeutic mechanisms and targets for OS. Chromobox homolog 4 may serve as a novel biomarker for OS, potentially offering innovative approaches for early diagnosis and tailored therapy of OS.
Methotrexate (MTX), a commonly used chemotherapeutic and immunosuppressive agent, induces dose-dependent neurotoxicity characterized by oxidative stress, glial reactivity, and dysregulation of apoptotic signaling, contri...Methotrexate (MTX), a commonly used chemotherapeutic and immunosuppressive agent, induces dose-dependent neurotoxicity characterized by oxidative stress, glial reactivity, and dysregulation of apoptotic signaling, contributing to chemotherapy-related cognitive impairment. Agomelatine (AGO), a melatonergic melatonin receptor type 1/melatonin receptor type 2 receptor agonist and 5-hydroxytryptamine receptor 2C antagonist, exhibits antioxidant, anti-inflammatory, and antiapoptotic properties, suggesting potential neuroprotection. Twenty-four female Wistar rats were randomly divided into control, MTX, MTX + AGO, and AGO groups (n = 6 each). AGO (20 mg/kg/d, by mouth) was administered for 7 days, with a single MTX injection (20 mg/kg, i.p.) on day 2. Cortical tissues were examined histopathologically (Hematoxylin and Eosin), immunohistochemically (glial fibrillary acidic protein and oligodendrocyte transcription factor 2), and molecularly via RT-quantitative polymerase chain reaction (qPCR) (Bcl-2-associated X protein and B-cell lymphoma 2 [BCL2]). MTX-induced marked cortical neurodegeneration, vascular hyperemia, gliosis, and parenchymal hemorrhage, accompanied by elevated glial fibrillary acidic protein and oligodendrocyte transcription factor 2 expression. MTX also increased proapoptotic Bcl-2-associated X protein and decreased antiapoptotic BCL2 expression, shifting the Bcl-2-associated X protein/BCL2 ratio toward apoptosis. AGO cotreatment significantly mitigated these alterations, preserving cortical cytoarchitecture, reducing glial activation, and restoring apoptotic balance. AGO alone upregulated BCL2 expression without histopathological abnormalities. AGO confers significant neuroprotection against MTX-induced cortical toxicity by attenuating glial reactivity and re-establishing apoptosis survival equilibrium. These findings highlight AGO as a promising adjunct therapeutic strategy for reducing chemotherapy-related neurotoxicity and improving neurological outcomes in MTX-treated patients. SIGNIFICANCE STATEMENT: Agomelatine, a clinically available melatonergic antidepressant, mitigated methotrexate-induced cortical neurotoxicity in rats by dampening astrocytic/oligodendrocytic reactivity and restoring the Bcl-2-associated X protein/B-cell lymphoma 2 apoptotic balance. These findings identify glial apoptotic homeostasis as a tractable target and support repurposing agomelatine as an adjunct to reduce chemotherapy-related neurotoxicity and potentially improve cognitive outcomes in methotrexate-treated patients.
Deoxylapachol (DLP), a natural product of marine origin, firstly isolated from the New Zealand brown algae, was firstly found has potent anti-inflammatory activities both in vitro and in vivo experiments. The study's obj...Deoxylapachol (DLP), a natural product of marine origin, firstly isolated from the New Zealand brown algae, was firstly found has potent anti-inflammatory activities both in vitro and in vivo experiments. The study's objective is aimed at its prospective molecular mechanisms. Firstly, the anti-inflammatory effect of DLP was assessed in zebrafish models induced by copper sulfate, tail-cutting and lipopolysaccharide exposure, respectively. The results showed that DLP effectively alleviated the acute inflammatory response by inhibiting the migration of immune cells, and the elevation of reactive oxygen species and nitric oxide. Moreover, DLP regulated the mRNA expression of tumor necrosis factor-α, matrix metalloproteinase 9, signal transducer and activator of transcription 3, nuclear factor of kappa light polypeptide gene enhancer in B-cells, cyclooxygenase-2, caspase-3, myeloperoxidase, glycogen synthase kinase 3 beta, mitogen-activated protein kinase 14a, mitogen-activated protein kinase 1, and hypoxia-inducible factor 1 alpha genes related to the interleukin (IL)-17/Toll-like receptor/TNF signaling pathway, and inflammatory cytokines (transforming growth factor beta, nucleotide-binding domain, leucine-rich repeat containing family, pyrin domain containing 3, IL-1β, IL-4, IL-6, IL-8, IL-10, Toll-like receptor 2 [TLR2], TLR3, TLR4, IL-17a, and IL-17b). DLP also regulates the expressions of proteins such as Tumor Necrosis Factor-α, nuclear factor of kappa light polypeptide gene enhancer in B-cells, matrix metalloproteinase 9, cyclooxygenase-2, p-P38, signal transducer and activator of transcription 3, GSK3α/β, caspase-3, inducible nitric oxide synthase, IL-18, nucleotide-binding domain, leucine-rich repeat containing family, pyrin domain containing 3, peroxisome proliferator-activated receptor and nuclear factor (erythroid-derived 2)-like 2. In conclusion, our finding indicated that DLP has the potential to be a candidate drug for the effective treatment of inflammation. SIGNIFICANCE STATEMENT: This study has shown that deoxylapachol exerts a potent anti-inflammatory effect in zebrafish inflammatory models. Deoxylapachol, a natural product of marine origin, has the potential to be an effective agent for anti-inflammatory treatment.
Alzheimer's disease is a progressive neurodegenerative disorder defined by the presence of extracellular amyloid-β (Aβ) plaques and intracellular neurofibrillary tangles, which together drive cognitive decline and memory...Alzheimer's disease is a progressive neurodegenerative disorder defined by the presence of extracellular amyloid-β (Aβ) plaques and intracellular neurofibrillary tangles, which together drive cognitive decline and memory impairment. Current therapies provide only symptomatic relief, highlighting the urgent need for disease-modifying strategies targeting the underlying pathology. Piperine, a plant-derived alkaloid with neuroprotective, antioxidant, and antiamyloidogenic properties, has shown preclinical efficacy against Aβ toxicity. However, its clinical translation is limited by poor aqueous solubility and low bioavailability. The present study aimed to develop and characterize spray-dried piperine-loaded solid dispersion (PIP@SDs) to overcome these pharmacokinetic barriers and enhance neuroprotective efficacy. PIP@SDs were prepared using spray drying with leucine as a carrier to improve solubility and stability. Solid-state characterization was performed using differential scanning calorimetry, Fourier-transform infrared spectroscopy, and powdered X-ray diffraction to confirm amorphization and molecular interactions. PIP@SDs demonstrated enhanced solubility, achieving 97.5% drug release in 24 hours, compared with pure drug and improved flow properties suitable for pharmaceutical manufacturing. In SH-SY5Y cells, PIP@SDs markedly increased viability after Aβ insult (from 50.2% to 80.8%, P < .0001), reduced oxidative stress, stabilized mitochondrial membrane potential, restored calcium equilibrium, and lowered apoptosis levels relative to free piperine. Notably, PIP@SDs inhibited cholinesterase activity and prevented Aβ aggregation, with all effects confirmed through quantitative and imaging analyses. In closing, by substantially enhancing solubility and bioavailability and providing superior neuroprotection against Aβ-mediated toxicity, spray-dried PIP@SDs offer a novel, multifunctional platform that supports their continued research as a promising candidate for disease-modifying interventions in Alzheimer's and related neurodegenerative disorders. SIGNIFICANCE STATEMENT: Poor aqueous solubility and limited bioavailability restrict the therapeutic potential of many neuroprotective natural compounds. This study demonstrates that spray-dried piperine-leucine solid dispersions markedly enhance piperine dissolution and significantly attenuate amyloid-β-induced oxidative stress, mitochondrial dysfunction, and apoptosis in neuronal cells. These findings highlight a pharmacologically relevant formulation strategy that improves the bioactivity of piperine and supports its development as a multifunctional therapeutic candidate targeting oxidative and mitochondrial pathways implicated in Alzheimer disease.
Obesity-associated hepatic steatosis is a major metabolic consequence of chronic dietary lipid excess and a critical precursor to progressive liver disease. In the present study, we investigated the therapeutic potential...Obesity-associated hepatic steatosis is a major metabolic consequence of chronic dietary lipid excess and a critical precursor to progressive liver disease. In the present study, we investigated the therapeutic potential of betulinic acid (BA), a plant-derived pentacyclic triterpenoid, in high-fat diet (HFD)-induced hepatic steatosis in rats, with specific emphasis on hepatic lipid handling and bile acid-regulated nuclear receptor signaling. Male Sprague-Dawley rats were fed an HFD for 16 weeks to induce obesity and subsequently treated with BA (40 mg/kg, oral) for 6 weeks. BA treatment markedly improved systemic dyslipidemia and reduced hepatic accumulation of triglycerides, cholesterol, free fatty acids, and phospholipids. These metabolic improvements were accompanied by normalization of serum liver marker enzymes and restoration of bile acid homeostasis. Histopathological assessment demonstrated pronounced attenuation of hepatic steatosis, lobular inflammation, hepatocyte ballooning, and a significant reduction in the Nonalcoholic Fatty Liver Disease Activity Score. At the molecular level, BA suppressed HFD-induced de novo lipogenesis by downregulating sterol regulatory element-binding protein-1c, acetyl-CoA carboxylase, and fatty acid synthase while concurrently restoring the expression of apolipoprotein B and microsomal triglyceride transfer protein. In parallel, BA re-established bile acid-regulated nuclear receptor signaling by restoring farnesoid X receptor and small heterodimer partner expression and normalizing cholesterol 7α-hydroxylase levels. Molecular docking analyses further supported these findings by revealing stable interactions of BA with multiple regulators of lipid and bile acid metabolism. Taken together, these findings identify BA as a multitarget metabolic modulator that alleviates obesity-associated hepatic steatosis through coordinated regulation of lipid metabolism and bile acid signaling pathways. SIGNIFICANCE STATEMENT: Obesity-associated hepatic steatosis lacks effective pharmacological interventions that target the underlying metabolic dysregulation. This study demonstrates that betulinic acid exerts hepatoprotective effects by coordinately suppressing hepatic lipogenesis, restoring lipid export, and normalizing bile acid-regulated nuclear receptor signaling in a HFD model. These findings identify betulinic acid as a multitarget metabolic modulator with potential therapeutic relevance for obesity-driven fatty liver disease.
Long-term proton pump inhibitor use is associated with cognitive and motor impairments and nutrient deficiencies. Whether these impairments result directly from proton pump inhibitors or indirectly from nutrient deficien...Long-term proton pump inhibitor use is associated with cognitive and motor impairments and nutrient deficiencies. Whether these impairments result directly from proton pump inhibitors or indirectly from nutrient deficiencies remains unclear. Chronic pantoprazole treatment (0 or vehicle, 1, 10, or 100 mg/kg/day, by mouth, 38 days) was evaluated across tests of anxiety-like behavior, learning and memory behavior, and motor function and coordination in 4-month-old male C57BL/6 mice. Serum levels of holotranscobalamin and magnesium, as well as poly(A) RNA sequencing of brain tissue, were subsequently evaluated. Pantoprazole (100 mg/kg) reduced the percentage of spontaneous alternations in the Y-maze (P < .05 vs 0 and 1 mg/kg) and decreased the distance traveled in Rotarod testing (P < .05 vs vehicle), suggesting that it impaired short-term working memory behavior and motor coordination, respectively. All pantoprazole doses increased anxiety-like behavior in the open field (P < .05 vs vehicle). In novel (N) object recognition testing, pantoprazole (1 mg/kg) reduced the discrimination index at 24 hours (P < .05, vs vehicle), suggesting it impaired recognition memory behavior. Magnesium levels were similar across chronic pantoprazole groups. Holotranscobalamin, the bioavailable form of vitamin B, was lower at 10 mg/kg pantoprazole (P < .05 vs vehicle). Poly(A) RNA sequencing identified 15 differentially expressed genes in hippocampus (10 upregulated and 5 downregulated) and 32 in FC (14 upregulated and 18 downregulated) with 100 mg/kg pantoprazole treatment compared to vehicle. There was no correspondence between behavioral changes and alterations in serum nutrient levels with the respective doses of pantoprazole. SIGNIFICANCE STATEMENT: Long-term proton pump inhibitor use may affect cognition, motor function, and nutrient status. This study examined the behavioral impact of chronic pantoprazole treatment in young male mice and whether declines in serum magnesium or holotranscobalamin levels could explain behavioral changes. Doses of chronic pantoprazole that altered behavior were not associated with declines in either nutrient.
Kidney diseases represent one of the most prevalent and rapidly growing global health burdens, with rising mortality linked to both acute kidney injury and chronic kidney disease (CKD). Despite the increasing incidence o...Kidney diseases represent one of the most prevalent and rapidly growing global health burdens, with rising mortality linked to both acute kidney injury and chronic kidney disease (CKD). Despite the increasing incidence of CKD affecting 8%-16% of the world's population, therapeutic options remain limited, largely because of an incomplete understanding of underlying pathophysiology and the lack of reliable biomarkers. Recent evidence highlights that free-fatty acids (FFA), particularly short-chain and long-chain fatty acids, are beneficial modulators of renal health, associated with reduced CKD risk and slower kidney function decline. Although effects of FFA have historically been linked to their intracellular activity, they can also engage a family of cell-surface G protein-coupled FFA receptors (FFAR), including FFARs FFA2/GPR43 and FFA3/GPR41, which are activated by short-chain fatty acids, and FFA1/GPR40 and FFA4/GPR120, which are agonized by long-chain fatty acids. This review summarizes current knowledge of FFAR-mediated mechanisms in acute kidney injury, CKD, and associated renal pathologies, emphasizing their potential as therapeutic targets to improve kidney outcomes. We also discuss the therapeutic relevance of dietary FFA related to these FFAR in relation to renal disease. SIGNIFICANCE STATEMENT: Fatty acid sensing G protein-coupled receptors have emerged as important regulators of renal inflammation, fibrosis, and tubular survival across acute and chronic models of kidney injury. This review synthesizes current evidence for short- and long-chain free-fatty acid receptors in kidney disease, highlights translational relevance, and outlines challenges that must be addressed to advance free-fatty acid receptor-targeted therapies.
Previous studies demonstrated that CNS4, a glutamate concentration biased N-methyl-D-aspartate receptor modulator, produces central analgesic and stress mitigating effects in mice. To translate these findings to a nonrod...Previous studies demonstrated that CNS4, a glutamate concentration biased N-methyl-D-aspartate receptor modulator, produces central analgesic and stress mitigating effects in mice. To translate these findings to a nonrodent species, we evaluated the pharmacokinetics (PK) and safety of CNS4 in 21 adult beagle dogs. A custom formulation was developed for subcutaneous (SC) delivery, and an initial 2 dose PK study (5 and 10 mg/kg) was conducted. CNS4 achieved rapid absorption, with T values of 1.50 ± 0.87 hour (5 mg/kg) and 1.67 ± 0.58 hour (10 mg/kg). Pharmacokinetics were comparable across the limited doses studied, and apparent volumes of distribution for the extravascular route were 33.5 ± 12.5 and 25.6 ± 3.67 L/kg, respectively. To assess oral feasibility, a 50 mg/kg dose of CNS4 suspended in 0.5% carboxymethyl cellulose was first tested in mice and then in dogs. Oral administration in dogs resulted in rapid absorption, with a T of ∼1 hour, comparable to the SC route. Systemic tolerability was assessed in an acute toxicity study involving 4 groups (vehicle, 5, 10, and 25 mg/kg SC) of dogs. No changes in behavior, food intake, or body weight were observed over 14 days. Hematology, lipid panels, and electrolyte measurements collected on days 0 (predose), 7, and 14 revealed no significant CNS4 related abnormalities. Together, these findings demonstrate that CNS4 has an acceptable safety profile and predictable PK across SC and oral administration routes in dogs. This first-in-dog evaluation supports further development of CNS4 as a potential non opioid analgesic and stress mitigating therapeutic agent. SIGNIFICANCE STATEMENT: N-methyl-D-aspartate receptor subtypes are attractive therapeutic targets for chronic pain and post-traumatic stress disorder because of their critical role in emotional memory formation. This translational pharmacology study demonstrates the safety and predictable pharmacokinetics of a centrally acting analgesic and stress mitigating N-methyl-D-aspartate receptor modulator (CNS4) in dogs after single dose subcutaneous and oral administration.
Preclinical studies indicate that blocking diacylglycerol kinase (DGK) family members DGKα or DGKζ can improve antitumor immunity, prompting the development of clinical-stage, potent and selective small molecule inhibito...Preclinical studies indicate that blocking diacylglycerol kinase (DGK) family members DGKα or DGKζ can improve antitumor immunity, prompting the development of clinical-stage, potent and selective small molecule inhibitors of DGKα and/or DGKζ. DGKα and DGKζ are the most widely expressed DGK family members by immune cells, and both enzymes convert the signaling lipid diacylglycerol (DAG) to phosphatidic acid. DAG is a critical second messenger downstream of T cell receptor (TCR) stimulation that promotes activation and effector function. Blocking DGKα or DGKζ activity enhances DAG-mediated signaling, potentiating immune cell activity. Because DGKα and DGKζ functionally overlap, we sought to compare the effects of pharmacological inhibition strategies targeting DGKα or DGKζ individually or simultaneously (DGKα/ζ) to determine which approach maximized immune cell activation. Evaluation of TCR downstream signaling using primary human and mouse cells revealed that dual DGKα/ζ inhibition promoted the greatest increase in cellular activity, including antigen-dependent cytokine production and tumor cell killing. In contrast, pharmacological inhibition of DGKζ alone had modest effects, and inhibition of DGKα alone had minimal bearing on TCR-mediated activity. Notably, loss of DGKα and DGKζ protein was observed following inhibitor treatment and may point to an additional mechanism of action for DGK targeting small molecule inhibitors. Finally, analysis of biopsies from patients with nonsmall cell lung cancer showed that tumor infiltrating lymphocytes expressed both DGKα and DGKζ and exhibited increased activation and cytokine production ex vivo upon DGKα/ζ coinhibition in conjunction with TCR stimulation, indicating that tumor infiltrating lymphocytes are sensitive to DGKα/ζ coinhibition. SIGNIFICANCE STATEMENT: This work directly compares pharmacological inhibition of DGKα and DGKζ, affirming that DGKα/ζ coinhibition is required to maximally increase TCR responses. Importantly, DGKα/ζ inhibition increased activation and cytokine production in both healthy T cells and tumor infiltrating lymphocytes.