Receptor Protein Levels Research Articles

Targeting calcium signaling by inositol trisphosphate receptors: A novel mechanism for the anti-asthmatic effects of Houttuynia cordata

Asthma is a chronic inflammatory disease characterized by airway hypersensitivity and remodeling. The current treatments provide only short-term benefits and may have undesirable side effects; thus, alternative or supplementary therapy is needed. Because intracellular calcium (Ca2+) signaling plays an essential role in regulating the contractility and remodeling of airway smooth muscle cells, the targeting of Ca2+ signaling is a potential therapeutic strategy for asthma. Houttuynia cordata is a traditional Chinese herb that is used to treat asthma due to its anti-allergic and anti-inflammatory properties. We hypothesized that H. cordata might modulate intracellular Ca2+ signaling and could help relieve asthmatic airway remodeling. We found that the mRNA and protein levels of inositol trisphosphate receptors (IP3Rs) were elevated in interleukin-stimulated primary human bronchial smooth muscle cells and a house dust mite–sensitized model of asthma. The upregulation of IP3R expression enhanced intracellular Ca2+ release upon stimulation and contributed to airway remodeling in asthma. Intriguingly, pretreatment with H. cordata essential oil rectified the disruption of Ca2+ signaling, mitigated asthma development, and prevented airway narrowing. Furthermore, our analysis suggested that houttuynin/2-undecanone could be the bioactive component in H. cordata essential oil because we found similar IP3R suppression in response to the commercially available derivative sodium houttuyfonate. An in silico analysis showed that houttuynin, which downregulates IP3R expression, binds to the IP3 binding domain of IP3R and may mediate a direct inhibitory effect. In summary, our findings suggest that H. cordata is a potential alternative treatment choice that may reduce asthma severity by targeting the dysregulation of Ca2+ signaling.

(074) Association of Androgen Receptor Signaling in Human Corpus Cavernosum with Serum Testosterone Levels

Abstract Introduction Testosterone (T) plays an important role in properly developing and maintaining male reproductive tissues. The normal serum testosterone levels can vary between 300 to 1000 ng/dL or 10.4 to 34.7 nmol/L. However, it is unclear whether androgen receptor signaling can vary based on serum T levels or the half-life of testosterone therapy. What remains unknown is whether there is a uniform serum T level above which androgen receptor signaling within the tissue remains similar (saturation hypothesis). Objective We first evaluated whether varying serum levels of T alter androgen receptor (AR) signaling in human penile tissue. Then, we evaluated and compared AR signaling in corpus cavernosum tissue of men treated with short-acting vs. long-acting testosterone. We hypothesized that we would find similar AR signaling in the penile tissue of both eugonadal and hypogonadal men as well as those men receiving short and long-acting testosterone. Methods We obtained human corpus cavernosum (hCC) tissue biopsies during penile surgery (for erectile dysfunction) from 14 men. We recorded their serum T levels on week prior to surgery before 10AM. We used western blot to evaluate adrenergic receptor (AR) and VEGF protein levels. Men without erectile dysfunction undergoing penile surgery for Peyronie's disease were used as controls. Results The median age of participants was 61 (IQR 12) and the median T level was 325ng/dL (IQR 201.45). The results of western blot showed expression of similar levels of VEGF in hCC samples from men with T >200ng/dL. However, men with serum T levels lower than 200 ng/dL had decreased expression of VEGF and AR. Conclusions This finding is significant because it demonstrates that serum testosterone beyond a certain threshold may have a minimal impact on erectile function. If a serum testosterone threshold is identified, this threshold could be a valuable target for future studies on the effects of testosterone on ED. It may alter the treatment threshold for men who have ED as their primary symptom of hypogonadism. This data also suggests the saturation value for penile androgen receptors. Disclosure Any of the authors act as a consultant, employee or shareholder of an industry for: Acerus Pharmaceuticals, Boston Scientific, Coloplast, Endo Pharmaceuticals, Empower Pharmacy, Nestle Health, Olympus, Hims, Inc

Folic acid relieves bone cancer pain by downregulating P2X2/3 receptors in rats

BackgroundBone cancer pain (BCP) remains a clinical challenge due to the limited and side effects of therapeutic methods. Folic acid has been known as an FDA approved dietary supplement and proved to have an analgesic effect in neuropathic pain. Here we investigate the role and mechanism of folic acid in bone cancer pain of a rat model. MethodsWalker 256 tumor cells were inoculated into the left tibia of rats to induce bone cancer pain model. Pain reflex were assessed by paw withdrawal threshold (PWT) response to Von Frey filaments and paw withdrawal latency (PWL) response to thermal stimulation. Folic acid was injected intraperitoneally to evaluate its analgesic effect in rats with bone cancer pain. Western blotting and qPCR were used to determine P2X2/3 receptor protein and mRNA levels in ipsilateral L4-6 dorsal root ganglion (DRG) and spinal dorsal horn (SDH). ResultsThe PWT and PWL of rats with bone cancer pain were obviously decreased compared to the naïve and sham rats. Interestingly, continuous folic acid treatment significantly increased the PWT and PWL of rats with bone cancer pain. P2X2 and P2X3 receptors were clearly upregulated at both mRNA and protein expression in L4-6 DRG and SDH of rats with bone cancer pain. P2X2 and P2X3 receptors were mainly localized with CGRP (calcitonin gene-related peptide) or IB4 (isolectin B4) positive neurons in L4-6 DRG of rats with bone cancer pain. Notably, continuous folic acid treatment significantly reduced the expression of P2X2 and P2X3 receptors in L4-6 DRG and SDH of rats with bone cancer pain. Finally, intrathecal injection of A317491 (a selective antagonist of P2X2/3 receptors) markedly elevated the PWT and PWL of rats with bone cancer pain. ConclusionThese results suggest that folic acid has an effective antinociceptive effect on bone cancer pain, which is mediated by downregulating P2X2/3 receptors in L4-6 DRG and SDH of rats with bone cancer pain. Folic acid may be a novel therapeutic strategy in cancer patients for pain relief.

Distinct regulation of iron homeostasis in rat heart and liver in response to systemic iron deficiency

Chronic heart failure (HF) is characterized by severe cardiac mitochondrial dysfunction. Cellular energetic function is importantly regulated by iron since mitochondrial enzymes involved in oxidative phosphorylation, antioxidative defense and oxygen transport require iron-containing cofactors (heme, Fe-S clusters). In HF, systemic iron deficiency (ID) shows high prevalence (30–50%) and is associated with worse exercise capacity and increased mortality. Human and animal studies showed discordant results concerning iron content in failing heart, even reporting mitochondrial iron overload in some instance. This leads to a controversy of using iron chelation or iron supplementation as metabolic therapy for HF. Our aim was to investigate the regulation of key actors of cellular iron metabolism in the left (LV) and the right (RV) ventricles of two rat models of systemic ID. A comparative study was also performed in the liver, the main iron storage organ known to sense systemic ID. We developed two rat models of systemic ID without anaemia and cardiac remodelling, using low iron diet for 60 days, combined either with two distinct blood withdrawals at day 0 and 30 or with iron chelator deferiprone from day 1 to 60. Control rats were fed on a normal iron diet. Rats were sacrificed at day 60, serum and tissue iron contents were determined using a photometric colorimetric method. mRNA and protein levels of transferrin receptor 1 (TfR1), L-ferritin (L-Ft) and ferroportin (Fpn), involved in iron uptake, storage and export respectively were measured in biopsies of RV, LV and liver. Liver hepcidin, regulating systemic iron homeostasis, was analysed at mRNA level as well as cardiac hepcidin required for autonomous cellular iron homeostasis. Our main results showed that LV and RV maintained their iron content in a physiological range while the liver drastically decreased its iron pool in response to ID. In this condition, the RV exhibited a post-transcriptional regulation of both TfR1, L-Ft and Fpn similarly to the one found in the liver. In contrast, the LV showed a post-translational regulation of TfR1, a potential post-transcriptional regulation of L-Ft and no regulation for Fpn in ID. Our study showed that both ventricles initiate an adaptative response to maintain their physiological iron pool under systemic ID. These results should be confirmed in a HF model with systemic ID to better establish the benefits of iron chelation or supplementation in HF patients.

Pre- and postnatal developmental exposure to the polychlorinated biphenyl mixture aroclor 1221 alters female rat pituitary gonadotropins and estrogen receptor alpha levels

Polychlorinated-biphenyls (PCBs) are industrial compounds, which were widely used in manufacturing of electrical parts and transformers. Despite being banned in 1979 due to human health concerns, they persist in the environment. In humans and experimental model systems, PCBs elicit toxicity in part by acting as endocrine-disrupting chemicals (EDCs). Aroclor 1221 (A1221) is a weakly estrogenic PCB mixture known to alter reproductive function in rodents. EDCs can impact hormone signaling at any level of the hypothalamic-pituitary-gonadal (HPG) axis, and we investigated the effects of A1221 exposure during the prenatal and postnatal developmental periods on pituitary hormone and steroid receptor expression in female rats. Examining offspring at 3 ages, postnatal day 8 (P8), P32 and P60, we found that prenatal exposure to A1221 increased P8 neonate pituitary luteinizing hormone beta (Lhb) mRNA and LHβ gonadotrope cell number while decreasing LH serum hormone concentration. No changes in pituitary hormone or hormone receptor gene expression were observed peri-puberty at P32. In reproductively mature rats at P60, we found pituitary follicle stimulating hormone beta (Fshb) mRNA levels increased by prenatal A1221 exposure with no corresponding alterations in FSH hormone or FSHβ expressing cell number. Estrogen receptor alpha (ERα) mRNA and protein levels were also increased at P60, but only following postnatal A1221 dosing. Together, these data illustrate that exposure to the PCB A1221, during critical developmental windows, alters pituitary gonadotropin hormone subunits and ERα levels in offspring at different phases of maturation, potentially impacting reproductive function in concert with other components of the HPG axis.

Exogenous growth hormone administration during total sleep deprivation changed the microRNA-9 and dopamine D2 receptor expressions followed by improvement in the hippocampal synaptic potential, spatial cognition, and inflammation in rats.

Disorders caused by total sleep deprivation can be modulated by the administration of growth hormone, which could affect the expression of microRNA-9 and dopamine D2 receptor expressions followed by improvement in the hippocampal synaptic potential, spatial cognition, and inflammation in rats. The present study aimed to elucidate the putative effects of exogenous growth hormone (GH) against total sleep deprivation (TSD)-induced learning and memory dysfunctions and possible involved mechanisms. To induce TSD, rats were housed in homemade special cages equipped with stainless steel wire conductors to induce general and inconsistent TSD. They received a mild repetitive electric shock to their paws every 10 min for 21 days. GH (1 mg/kg, sc) was administered to adult young male rats once daily for 21-day-duration induction of TSD. Spatial learning and memory performance, inflammatory status, microRNA-9 (miR-9) expression, dopamine D2 receptor (DRD2) protein level, and hippocampal histological changes were assayed at scheduled times after TSD. The results indicated that TSD impaired spatial cognition, increased TNF-α, decreased level of miR-9, and increased DRD2 levels. Treatment with exogenous GH improved spatial cognition, decreased TNF-α, increased level of miR-9, and decreased DRD2 levels after TSD. Our findings suggest that GH may play a key role in the modulation of learning and memory disorders as well as the ameliorating abnormal DRD2-related functional disorders associated with miR-9 in TSD.

NLRP3 mediates the neuroprotective effects of SVHRSP derived from scorpion venom in rotenone-induced experimental Parkinson’s disease model

Ethnopharmacological relevanceIn traditional Chinese medicine, scorpion is used to treat diseases with symptoms such as trembling, convulsion and dementia. Our laboratory employs patented technology to extract and purify the active single component from scorpion venom. We then utilize mass spectrometry to determine the amino acid sequence of the polypeptide and synthesize it artificially to acquire the polypeptide with a purity of 99.3%, named SVHRSP (Scorpion Venom Heat-Resistant Peptide). SVHRSP has been demonstrated to display potent neuroprotective efficacy in Parkinson’s disease. Aim of the studyTo explore the molecular mechanisms and potential molecular targets of SVHRSP-afforded neuroprotection in PD mouse models, as well as to investigate the role of NLRP3 in SVHRSP-mediated neuroprotection. Materials and methodsThe PD mouse model was induced by rotenone and the neuroprotective role of SVHRSP on the PD mouse model was measured using the gait test, rotarod test, the number of dopaminergic neurons, and the activation of microglia. RNA sequencing and GSEA analysis were performed to find the differentially biological pathways regulated by SVHRSP. Primary mid-brain neuron-glial cultures and NLRP3−/− mice were applied to verify the role of NLRP3 by using qRT-PCR, western blotting, enzyme-linked immunosorbent assay (ELISA) and immunostaining. ResultsSVHRSP-afforded dopaminergic neuroprotection was accompanied with inhibition of microglia-mediated neuroinflammatory pathways. Importantly, depletion of microglia markedly reduced the neuroprotective efficacy of SVHRSP against rotenone-induced dopaminergic neurotoxicity in vitro. SVHRSP inhibited microglial NOD-like receptor pathway, mRNA expression and protein level of NLRP3 in rotenone PD mice. SVHRSP also reduced rotenone-induced caspse-1 activation and IL-1β maturation, indicating that SVHRSP mitigated activation of NLRP3 inflammasome. Moreover, inactivation of NLRP3 inflammasome by MCC950 or genetic deletion of NLRP3 almost abolished SVHRSP-afforded anti-inflammatory, neuroprotective effects and improvement of motor performance in response to rotenone. ConclusionsNLRP3 mediated the neuroprotective effects of SVHRSP in rotenone-induced experimental PD model, providing additional evidence for the mechanisms of SVHRSP-afforded anti-inflammatory and neuroprotective effects in PD.

Cyclic adenosine monophosphate-elevating agents inhibit amyloid-beta internalization and neurotoxicity: their action in Alzheimer's disease prevention.

Cyclic adenosine monophosphate-elevating agents inhibit amyloid-beta internalization and neurotoxicity: their action in Alzheimer's disease prevention.

Abstract 3620: N-myristoylation inhibition reduces angiogenesis and cancer cell migration

Abstract Myristoylation is the modification of proteins by the 14-carbon fatty acid myristate. In humans, two N-myristoyltransferases (NMT1 and NMT2) catalyze myristate transfer onto >200 proteins to regulate membrane binding and signal transduction. The pan-NMT inhibitor PCLX-001 has been in phase I clinical trials over a year for the treatment of lymphoma and advanced solid malignancies. Lack of myristoylation promotes the degradation of numerous normally myristoylated proteins like the proto-oncogenic Src family kinases (SFKs). SFKs like Src and Lyn are essential for downstream signaling of Vascular Endothelial Growth Factor (VEGF) and Epidermal Growth Factor (EGF) receptor tyrosine kinases. Thus, SFKs are critical for angiogenesis and cell motility. Angiogenesis is essential for tumor growth and dissemination of cancer cells to distal locations. VEGF-induced angiogenesis requires Src activation, suggesting that PCLX-001 may have antiangiogenic properties. SFKs also promote cell migration/metastasis, the leading cause of cancer-associated mortality. Lyn SFK knockdown was shown to reduce breast cancer migration and neighbouring tissue invasion, suggesting that PCLX-001 may also inhibit cell migration/metastasis. We investigated the antiangiogenic properties of PCLX-001 in vitro using HUVEC umbilical cord cells. PCLX-001 treatment significantly reduced HUVEC sprouting and tube-formation to levels lower than two FDA-approved angiogenesis inhibitors Sorafenib and Sunitinib. PCLX-001 also significantly reduced neo-vascularisation in vivo at lower concentrations than these two drugs using a chicken chorioallantoic membrane model, demonstrating the high potential of myristoylation inhibitors to reduce angiogenesis. We then validated our observations in vivo using a HT-1376 bladder mouse xenograft model. Tumors from mice treated for 28 days at increasing PCLX-001 concentrations showed significant reduction of Src, Lyn, and VEGF receptor protein levels as measured by immunohistochemistry, further validating PCLX-001 effect in vivo. We hypothesize that PCLX-001 acts downstream of VEGF/EGF receptors by inducing degradation of un-myristoylated SFKs. We confirmed this by knocking down NMT1 or treating breast cancer cells with PCLX-001 and found that both significantly reduced SFK protein levels. In a wound-healing assay using PCLX-001, we observed significant dose-dependent cell migration inhibition. PCLX-001 also dose-dependently reduced migration and invasion in transwell migration/invasion assays, indicating that myristoylation is required for optimal migration and potentially cancer cell metastasis. Overall, by lowering SFK levels, PCLX-001 reduces angiogenesis, cell migration/invasion, and thus potentially cancer metastasis. Reducing these classical cancer hallmarks in malignant cells may benefit a wide array of cancer patients and ultimately improve cancer treatment outcomes. Citation Format: Rony Pain, Erwan Beauchamp, Katia Carmine-Simmen, Jay Gamma, Rebecca Reif, Abul Azad, Allan Murray, John Lewis, Luc Berthiaume. N-myristoylation inhibition reduces angiogenesis and cancer cell migration. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3620.

Changes in cytokine and cytokine receptor levels during postnatal development of the human dorsolateral prefrontal cortex

In addition to their traditional roles in immune cell communication, cytokines regulate brain development. Cytokines are known to influence neural cell generation, differentiation, maturation, and survival. However, most work on the role of cytokines in brain development investigates rodents or focuses on prenatal events. Here, we investigate how mRNA and protein levels of key cytokines and cytokine receptors change during postnatal development of the human prefrontal cortex. We find that most cytokine transcripts investigated (IL1B, IL18, IL6, TNF, IL13) are lowest at birth and increase between 1.5 and 5 years old. After 5 years old, transcriptional patterns proceeded in one of two directions: decreased expression in teens and young adults (IL1B, p = 0.002; and IL18, p = 0.004) or increased mean expression with maturation, particularly in teenagers (IL6, p = 0.004; TNF, p = 0.002; IL13, p < 0.001). In contrast, cytokine proteins tended to remain elevated after peaking significantly around 3 years of age (IL1B, p = 0.012; IL18, p = 0.026; IL6, p = 0.039; TNF, p < 0.001), with TNF protein being highest in teenagers. An mRNA-only analysis of cytokine receptor transcripts found that early developmental increases in cytokines were paralleled by increases in their ligand-binding receptor subunits, such as IL1R1 (p = 0.033) and IL6R (p < 0.001) transcripts. In contrast, cytokine receptor-associated signaling subunits, IL1RAP and IL6ST, did not change significantly between age groups. Of the two TNF receptors, the ‘pro-death’ TNFRSF1A and ‘pro-survival’ TNFRSF1B, only TNFRSF1B was significantly changed (p = 0.028), increasing first in toddlers and again in young adults. Finally, the cytokine inhibitor, IL13, was elevated first in toddlers (p = 0.006) and again in young adults (p = 0.053). While the mean expression of interleukin-1 receptor antagonist (IL1RN) was highest in toddlers, this increase was not statistically significant. The fluctuations in cytokine expression reported here support a role for increases in specific cytokines at two different stages of human cortical development. The first is during the toddler/preschool period (IL1B, IL18, and IL13), and the other occurs at adolescence/young adult maturation (IL6, TNF and IL13).

Long-term potentiation and spatial memory training stimulate the hippocampal expression of RyR2 calcium release channels.

Introduction: Neuronal Ca2+ signals generated through the activation of Ca2+-induced Ca2+ release in response to activity-generated Ca2+ influx play a significant role in hippocampal synaptic plasticity, spatial learning, and memory. We and others have previously reported that diverse stimulation protocols, or different memory-inducing procedures, enhance the expression of endoplasmic reticulum-resident Ca2+ release channels in rat primary hippocampal neuronal cells or hippocampal tissue. Methods and Results: Here, we report that induction of long-term potentiation (LTP) by Theta burst stimulation protocols of the CA3-CA1 hippocampal synapse increased the mRNA and protein levels of type-2 Ryanodine Receptor (RyR2) Ca2+ release channels in rat hippocampal slices. Suppression of RyR channel activity (1 h preincubation with 20 μM ryanodine) abolished both LTP induction and the enhanced expression of these channels; it also promoted an increase in the surface expression of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunits GluR1 and GluR2 and caused a moderate but significant reduction of dendritic spine density. In addition, training rats in the Morris water maze induced memory consolidation, which lasted for several days after the end of the training period, accompanied by an increase in the mRNA levels and the protein content of the RyR2 channel isoform. Discussion: We confirm in this work that LTP induction by TBS protocols requires functional RyR channels. We propose that the increments in the protein content of RyR2 Ca2+ release channels, induced by LTP or spatial memory training, play a significant role in hippocampal synaptic plasticity and spatial memory consolidation.

Effect of acupuncture on the opening time of implantation window and endometrial receptivity in controlled ovarian hyperstimulation rats during peri-implantation period.

To investigate the effect of acupuncture for improving the pregnancy rate of COH rats from the viewpoint of regulating the opening time of the implantation window and endometrial receptivity. Experimental rats were randomly divided into normal group (N), model group (M) and acupuncture group(A), and samples were collected on Day 4, 5 and 6 after mating. COH rats were treated with acupuncture at SP6, LR3, and ST36 once a day for 7 times. The pinopodes were observed under a scanning electron microscope. Serum estrogen and progesterone levels were measured via ELISA. The protein and mRNA levels of estrogen receptor (ER), progesterone receptor (PR), leukemia inhibitory factor (LIF), integrin β3, vascular endothelial growth factor (VEGF), and fibroblast growth factor 2 (FGF-2) in the endometrium were evaluated via West-blot, immunohistochemistry, and PCR. Compared with group N, the pregnancy rate of group M was significantly decreased (P<0.05), and the abnormal serum hormone levels and implantation window advancement were observed. Compared with group M, the pregnancy rate of group A was significantly increased (P<0.05), the supraphysiological serum progesterone levels were restored to normalcy (P<0.05), and the advanced implantation window was restored to a certain extent. Further, the abnormal ER, PR, LIF, integrin β3, VEGF, and FGF-2 expression levels of the endometrium got recovered to varying degrees. Acupuncture may restore the estrogen and progesterone balance in COH rats and the forward shift of the implantation window to a certain extent, improving the endometrial receptivity and finally improving the pregnancy rate of COH rats.

Colchicine reduces the activation of NLRP3 inflammasome in COVID-19 patients.

To evaluate whether colchicine treatment was associated with the inhibition of NLRP3 inflammasome activation in patients with COVID-19. We present a post hoc analysis from a double-blinded placebo-controlled randomized clinical trial (RCT) on the effect of colchicine for the treatment of COVID-19. Serum levels of NOD-like receptor protein 3 (NLRP3) inflammasome products-active caspase-1 (Casp1p20), IL-1β, and IL-18-were assessed at enrollment and after 48-72h of treatment in patients receiving standard-of-care (SOC) plus placebo vs. those receiving SOC plus colchicine. The colchicine regimen was 0.5mg tid for 5days, followed by 0.5mg bid for another 5days. Thirty-six patients received SOC plus colchicine, and thirty-six received SOC plus placebo. Colchicine reduced the need for supplemental oxygen and the length of hospitalization. On Days 2-3, colchicine lowered the serum levels of Casp1p20 and IL-18, but not IL-1β. Treatment with colchicine inhibited the activation of the NLRP3 inflammasome, an event triggering the 'cytokine storm' in COVID-19. RBR-8jyhxh.

Tilianin Protects against Nonalcoholic Fatty Liver Disease in Early Obesity Mice.

Non-alcoholic fatty liver disease (NAFLD) has emerged as one of the most frequent types of liver disease in pediatric populations with obesity. Tilianin has multiple biological activities including anti-inflammatory and antioxidant. Here, we aim to explore the functions and possible mechanisms of tilianin on NAFLD in obese children. A high-fat high-carbohydrate (HFHC) diet was used to feed 21-d-old mice. Tilianin was administered at a dose of 10 or 20 mg/kg daily. HFHC-fed mice gained weight, increased liver index. The liver showed hepatocyte ballooning, inflammatory infiltration, and steatosis. Elevated levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl transpeptidase (GGT), low-density lipoprotein cholesterol (LDL-C) and total cholesterol (TC) and reduced the high-density lipoprotein cholesterol (HDL-C) level were found in HFHC-fed mice. Administration of tilianin significantly reduced these impairments. We further evaluated proteins related to lipid metabolism and observed that LXRα, SREBP-1c, FAS and ACC1 expression were blunted following tilianin administration. In addition, tilianin suppressed reactive oxygen species (ROS) overproduction and lipid peroxide 4-Hydroxynonenal expression, ascribed to its oxidative stress-modulating capacity. Tilianin also reversed the increase in F4/80 expression and proinflammatory cytokine levels. Of note, tilianin administration resulted in decreased protein levels of active caspase-1 and NOD-like receptor protein 3 (NLRP3) in HFHC-fed mice. Our study suggests that tilianin may ameliorate NAFLD in early obese mice by modulating lipids metabolism, oxidative stress, and inflammation, which may in part involve inhibiting NLRP3 inflammasome activation.

Transcription factor GATA1 represses oxidized-low density lipoprotein-induced pyroptosis of human coronary artery endothelial cells.

Atherosclerosis (AS) is defined as a chronic inflammatory disorder underly the pathogenesis of cardiovascular diseases (CVDs). Endothelial pyroptosis is associated with AS-like diseases and other CVDs. This work was designed to expound on the effect of GATA-binding protein 1 (GATA1) on pyroptosis of human coronary artery endothelial cells (HCAECs) in AS. HCAECs were treated with oxidized-low density lipoprotein (ox-LDL) to establish HCAEC injury models. Plasmids for overexpressing GATA1 or silencing retinoic acid-related orphan receptor α (RORα) were transfected into HCAECs. Thereafter, the mRNA levels of GATA1 and RORα in HCAECs were detected using real-time quantitative polymerase chain reaction. HCAEC viability was examined using the cell counting kit-8 method. The levels of pyroptosis-related proteins NOD-like receptor protein 3 (NLRP3), cleaved-Caspase-1, N-terminal of gasdermin D (GSDMD-N), and pyroptosis-related inflammatory cytokines interleukin (IL)-1β and IL-18 were determined using Western blot and enzyme-linked immunosorbent assays, respectively. The targeting relationship between GATA1 and RORα was verified using the chromatin-immunoprecipitation assay. Then, the rescue experiment was conducted to explore the effect of RORα on pyroptosis of ox-LDL-treated HCAECs. In ox-LDL-treated HCAECs, GATA1 and RORα expressions were decreased, HCAEC viability was reduced, and the levels of NLRP3, cleaved-Caspase1, GSDMD-N, IL-1β, and IL-18 were elevated. GATA1 overexpression increased HCAEC viability and attenuated pyroptosis. GATA1 bound to the RORα promoter region to stimulate RORα transcription, and RORα suppression facilitated ox-LDL-induced pyroptosis of HCAECs. GATA1 activated RORα transcription and therefore limited pyroptosis of ox-LDL-treated HCAECs.

Cognitive and hippocampal effects of adult male mice exposure to environmentally relevant doses of phthalates

We recently showed that chronic exposure of adult male mice to environmental doses of DEHP alone or in a phthalate mixture altered blood brain barrier integrity and induced an inflammatory profile in the hippocampus. Here, we investigate whether such exposure alters hippocampus-dependent behavior and underlying cellular mechanisms. Adult C57BL/6 J male mice were continuously exposed orally to the vehicle or DEHP alone (5 or 50 μg/kg/d) or to DEHP (5 μg/kg/d) in a phthalate mixture. In the Morris water maze, males showed reduced latencies across days to find the platform in the cue and spatial reference memory tasks, regardless of their treatment group. In the probe test, DEHP-50 exposed males displayed a higher latency to find the platform quadrant. In the temporal order memory test, males exposed to DEHP alone or in a phthalate mixture were unable to discriminate between the most recently and previously seen objects. They also displayed reduced ability to show a preference for the new object in the novel object recognition test. These behavioral alterations were associated with a lowered dendritic spine density and protein levels of glutamate receptors and postsynaptic markers, and increased protein levels of the presynaptic synaptophysin in the hippocampus. Metabolomic analysis of the hippocampus indicated changes in amino acid levels including reduced tryptophan and L-kynurenine and elevated NAD + levels, respectively, a precursor, intermediate and endproduct of the kynurenine pathway of tryptophan metabolism. Interestingly, the protein amounts of the xenobiotic aryl hydrocarbon receptor, a target of this metabolic pathway, were elevated in the CA1 area. These data indicate that chronic exposure of adult male mice to environmental doses of DEHP alone or in a phthalate mixture impacted hippocampal function and structure, associated with modifications in amino acid metabolites with a potential involvement of the kynurenine pathway of tryptophan metabolism.

Association of Androgen-Receptor Gene Mutations with the Copy Number of Androgen-Receptor Silk Protein A Complex and Glutathione-S-Transferases T1 and M1 in Prostate Cancer Patients.

The purpose of our work was to explore the association of mutations in the androgen receptor gene and copy numbers of the androgen-receptor silk protein A complex with glutathione-S-transferases T1 and M1 in prostate cancer patients. Eighty-five patients with PC and 85 healthy controls were included in the study. Fasting peripheral venous blood was collected, whole blood genomic DNA was extracted, and AR gene-receptor genotype was detected by a high-resolution melting curve analysis detection technology. Expression levels of androgen receptor (AR) and filamin protein A (FlnA) were detected by Western blotting. RT-PCR was used to detect the copy number of T1 and M1 glutathione-S-transferases. The wild-type androgen receptor gene rs5918762 is of TT type. The frequencies of CC and TC genes in the prostate cancer group were significantly higher than those in the normal control group (P < 0.05). Compared with TT-type PC patients, PC patients with TC-type and CC-type had higher expression levels of sex hormone receptor silk protein A complex and higher copy numbers of GSTT1 and GSTM1 (P < 0.05). Androgen-receptor gene mutation (T ⟶ C) was significantly positively correlated with the expression level of androgen-receptor silk protein A complex and the copy number of GSTT1 and GSTM1. Androgen-receptor gene polymorphisms were significantly associated with expression levels of androgen receptor complex A and silk proteins, and copy numbers of T1 and M1 glutathione-S-transferases. A combination of four factors can be used to identify prostate cancer susceptibility and disease progression.

Potential role of the hydroxyl carboxylic acid receptor type 2 (HCAR2) in microglia pathophysiology: A possible cross-talk with C-X-C chemokine receptor 1 (CXCR1)

Following insults or injury, microglia cells are activated contributing to the cytotoxic response or by promoting an immune-mediated damage resolution. Microglia cells express HCA2R, a hydroxy carboxylic acid (HCA) receptor, which has been shown to mediate neuroprotective and anti-inflammatory effects.In this study we found that HCAR2 expression levels were increased in cultured rat microglia cells after Lipopolysaccharide (LPS) exposure. In a similar fashion, the treatment with MK 1903, a potent full agonist of HCAR2, increased the receptor protein levels. Moreover, HCAR2 stimulation prevented i) cells viability ii) morphological activation iii) pro/anti-inflammatory mediators production in LPS-treated cells. Likewise, HCAR2 stimulation reduced the proinflammatory mediators mRNA expression induced by neuronal chemokine fractalkine (FKN), a neuronal derived chemokine activating its unique receptor, chemokine receptor 1 (CX3CR1) on microglia surface. Interestingly, electrophysiological recordings in vivo revealed that MK1903 was able to prevent the increase of the nociceptive neurons (NS) firing activity mediated by the spinal FKN application in healthy rats.Collectively, our data demonstrate that HCAR2 is functionally expressed in microglia, by showing its capability to shift microglia toward an anti-inflammatory phenotype. Moreover, we indicated the contribute of HCAR2 in the FKN signaling and suggested a possible HCAR2/CX3CR1 functional interaction.This study paves the way for further investigations aimed at understanding the role HCAR2 as potential target in neuroinflammation-based CNS disorders.This article is part of the Special Issue on “The receptor-receptor interaction as a new target for therapy”.

Investigation of the effects of atorvastatin and Lactobacillus acidophilus on some hormones and oxidative stress in experimental hypercholesterolemia

AimThe investigation of serum leptin, ghrelin, insulin, seratonin hormones, NO, total oxidant/antioxidant status and brain cannaboid 1 receptor protein and apoptotic cell levels in atorvastatin and Lactobacillus acidophilus administrated experimental hypercholesterolemia was aimed in the project. MethodsIn the study, 5 experimental groups were formed. Group 1 was fed with standard rat chow, and Group 2 was fed with 2% cholesterol added standard rat chow for 8 weeks. Group 3 was fed with 2% cholesterol feed and received atorvastatin (20 mg/kg/day) for the last 4 weeks. Group 4 was given L. acidophilus (2 ×108 cfu/kg/day). Group 5 was given atorvastatin and L. acidophilus probiotic in the last 4 weeks of the experiment period. After the experimental period, blood samples were taken from each rat. Rats were sacrificed and brain tissues were taken for analyzes. In sera samples, leptin, ghrelin, insulin, serotonin hormones and NO levels were measured with ELISA. In brain samples, cannabinoid 1 receptor proteins and apoptosis levels were measured by ELISA. Total oxidant and antioxidant levels were investigated with using Rel Assay Kits. ResultsThe addition of cholesterol to feeds increased the levels of serum cholesterol, insulin and leptin levels; on the other hand, reduced the levels of serotonin and ghrelin. In hypercholesterolemia, total oxidant and NO levels were increased, and total antioxidant levels were decreased. ConclusionThe results showed that administrations of L. acidophilus and atorvastatin might be recommended for treatment of hypercholesterolemia.

Lavender essential oil accelerates lipopolysaccharide-induced chronic wound healing by inhibiting caspase-11-mediated macrophage pyroptosis.

Chronic wounds seriously affect the quality of life of the elderly, obese people, and diabetic patients. The excessive inflammatory response is a key driver of delayed chronic wound healing. Although lavender essential oil (EO [lav]) has been proven to have anti-inflammatory and accelerate wound curative effects, the specific molecular mechanism involved is still ambiguous. The results showed that the wounds treated with lipopolysaccharide (LPS) not only had delayed healing, but also the expression levels of pro-inflammatory cytokines, such as tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), interleukin-1β (IL-1β), and the inflammatory mediator protein, high-mobility group box 1 protein (HMGB-1), in the wound tissues were significantly increased. However, treatment of LPS-induced chronic wounds with EO (lav) accelerated wound healing and decreased IL-1β and HMGB-1 expression levels. It was further found that LPS induced macrophage pyroptosis to produce IL-1β. After treatment with EO (lav), the expression level of macrophage pyroptosis marker Gasdermin D (GSDMD) and pyroptosis-related cytotoxic effects were significantly reduced. Immunofluorescence results also directly indicate that EO (lav) can protect macrophages from LPS-induced pyroptosis. Moreover, EO (lav) can down-regulate expression levels of IL-1β, GSDMD, and nucleotide-binding oligomerization domain-like receptor protein 3 (NLRP3) in the caspase-11-related pyroptotic signaling pathway. This study demonstrates that EO (lav) can reduce proinflammatory factor production and ameliorate inflammatory response by inhibiting macrophage pyroptosis, which accelerates LPS-induced chronic wound healing.