Expression Of Inflammatory Proteins Research Articles

Mitochondrial DNA drives neuroinflammation through the cGAS-IFN signaling pathway in the spinal cord of neuropathic pain mice.

Neuroinflammation is pivotal in the development of neuropathic pain (NeP). While mitochondrial deoxyribonucleic acid (mtDNA) and cyclic GMP-AMP synthase (cGAS) are recognized for inducing inflammation in various neurological disorders, their involvement in NeP remains ambiguous. In this study, we examined: (1) the changes in mtDNA and cGAS in mice with NeP induced by chronic constriction injury (CCI) of the sciatic nerve, whether mtDNA triggers inflammation via the cGAS signaling; (2) the effects of RU.521, a cGAS antagonist, on CCI-induced nociception (allodynia and hyperalgesia) and relative inflammatory protein expression; (3) the activation of microglia and the cGAS-IFN pathway mediated by mtDNA in BV2 cell; (4) the effect of RU.521 on mtDNA-induced inflammatory response in BV2 cells. Results revealed reduced mtDNA levels in the sciatic nerve but increased levels in the spinal cord of CCI mice, along with elevated cGAS expression and inflammatory factors. RU.521 alleviated nociceptive behaviors in CCI mice, possibly by normalizing cGAS levels and suppressing inflammation. Neuron-derived mtDNA provoked cellular activation and upregulated cGAS signaling in BV2 cells. Additionally, RU.521 and DNase I effectively inhibited cGAS-induced inflammation. These findings underscore the critical role of mtDNA accumulation and mtDNA-mediated cGAS signaling in NeP development after peripheral nerve injury.

VLDL cholesterol associates with higher plasmatic expression of inflammatory proteins and atherosclerotic pathways compared to LDL cholesterol

Background and aim: High cholesterol in Low-Density Lipoproteins (LDL-C) is the key target of current pharmacological treatments aimed at reducing atherosclerotic cardiovascular disease (ACVD) risk. Increased cholesterol in Very low-density lipoproteins (“VLDL-C”) is an independent predictor of ACVD. VLDL-C was previously associated with markers of inflammation (for instance C-reactive protein). We now tested the relationship between either VLDL-C or LDL-C with a large spectrum of inflammatory proteins in plasma collected from subjects at different ACVD risks. Methods: We measured 276 proteins (OlinkTM) in plasma from a primary ACVD risk prevention cohort (“PLIC” in Milan; n=656 (8.2% on statins)) and a secondary ACVD risk prevention cohort (the Second Manifestations of ARTerial disease, “SMART”, the Netherlands, n=630 (50.8% on statins)). Cohorts were divided into three groups for VLDL-C (“Normal” VLDL-C<15 mg/dL, “High” VLDL-C 15-30 mg/dL, “Very high” VLDL-C >30 mg/dL) and LDL-C (“Normal” LDL-C <115 mg/dL, “High” LDL-C 115-155 mg/dL, “Very high” LDL-C>155 mg/dL). The expression (Normalized Protein eXpression, NPX) of each protein was compared among these groups by artificial intelligence. The performance to discriminate subjects with higher VLDL-C or LDL-C was evaluated by comparing the Areas Under the Curve (AUCs) of the Receiver Operating Characteristics curve (ROC) considering proteomics on top of ACVD risk factors (“CVRFs”: age, body mass index, systolic blood pressure, glycemia, therapies), versus the AUC of the ROCs with CVRFs alone. Results: The number of plasma proteins differentially expressed increased, as a function of higher VLDL-C in PLIC, as the NPXs of 84 were higher in “High” and the NPXs of 136 were higher in “Very high” vs “Normal” VLDL-C respectively. A similar trend was found in SMART, where the NPXs of 30 proteins were higher in “High” and the NPXs of 64 were higher in “Very high” vs “Normal” VLDL-C respectively. 26 proteins were shared between the two populations and recapitulated key atherosclerotic pathways (including chemotaxis of immune cells). The relationship between LDL-C was less marked; in PLIC, 14 proteins were more expressed in “High” and 33 in “Very high” vs “Normal” LDL-C respectively, while in SMART, the NPXs of 11 proteins were higher in “High” and the NPXs of 36 were higher in “Very high” vs “Normal” LDL-C respectively. Only 4 proteins were shared between high and very high LDL-C in the two populations. Finally, none of the proteins were shared between the groups of “High”/“Very high” VLDL-C and “High”/“Very high” LDL-C in the two cohorts. Canonical CVRFs alone slightly improved the ability to identify subjects with increased VLDL-C both in PLIC and SMART (AUCs between 0.6 on average), but adding plasma proteomics markedly improved the performance to identify subjects with “High” VLDL-C, in PLIC (AUC=0.767 (0.709-0.837)) and in SMART (AUC=0.781 (0.681-0.873)), and with “Very high” VLDL-C (AUC=0.950 (0.899-0.976) in PLIC, and AUC=0.938 (0.894-0.971) in SMART). The ROC of plasma proteomics with CVRFs was also superior to the ROC of the CVRFs alone to identify subjects with “High” and “Very high” LDL-C, but, as compared to the ROCs that discriminated subjects with “High” and “Very-high” VLDL-C, the AUCs were attenuated in both cohort (for “High” LDL-C: AUC=0.665 (0.558-0.774) in PLIC and AUC=0.775 (0.704-0.842) in SMART; for “Very high” LDL-C: AUC =0.776 (0.694-0.854) in PLIC and AUC=0.882 (0.825-0.931) in SMART). Conclusion: High VLDL-C associates with a higher number of differentially expressed plasma proteins versus high LDL-C and none of the proteins were in common. Our data do not underestimate the value of LDL-C in ACVD but reinforce the concept that VLDL-C may also promote different atherosclerotic pathways involved in determining ACVD.

Anti-cancer activity and cellular uptake of 7,3′,4′- and 7,8,4′-trihydroxyisoflavone in HepG2 cells under hypoxic conditions

Transarterial chemoembolisation (TACE) is used for unresectable hepatocellular carcinoma (HCC) treatment, but TACE-induced hypoxia leads to poor prognosis. The anti-cancer effects of soybean isoflavones daidzein derivatives 7,3′,4′-trihydroxyisoflavone (734THIF) and 7,8,4′-trihydroxyisoflavone (784THIF) were evaluated under hypoxic microenvironments. Molecular docking of these isomers with cyclooxygenase-2 (COX-2) and vascular endothelial growth factor receptor 2 (VEGFR2) was assessed. About 40 μM of 734THIF and 784THIF have the best effect on inhibiting the proliferation of HepG2 cells under hypoxic conditions. At a concentration of 40 μM, 784THIF significantly inhibits COX-2 expression in pre-hypoxia conditions compared to 734THIF, with an inhibition rate of 67.73%. Additionally, 40 μM 784THIF downregulates the expression of hypoxic, inflammatory, and metastatic-related proteins, regulates oxidative stress, and inhibits the expression of anti-apoptotic proteins. The uptake by HepG2 confirmed higher 784THIF level and slower degradation characteristics under post- or pre-hypoxic conditions. In conclusion, our results showed that 784THIF had better anti-cancer effects and cellular uptake than 734THIF.

Gene-expression profiling of laser-dissected islets and studies in deficient mice reveal chemokines as differential driving force of type 1 diabetes

Although type 1 diabetes (T1D) results from the autoimmune destruction of the insulin-producing β-cells, its treatment is largely restricted to exogenous insulin administration. Only few therapies targeting the autoaggressive immune system have been introduced into clinical practice or are considered in clinical trials. Here, we provide a gene expression profile of the islet microenvironment obtained by laser-dissection microscopy in an inducible mouse model. Thereby, we have identified novel targets for immune intervention. Increased gene expression of most inflammatory proteins was apparent at day 10 after T1D induction and largely paralleled the observed degree of insulitis. We further focused on genes involved in leukocyte migration, including chemokines and their receptors. Besides the critical chemokine CXCL10, we found several other chemokines upregulated locally in temporary or chronic manner. Localization of the chemokine ligand/receptor pairs to the islet microenvironment has been confirmed by RNAscope. Interference with the CXCL16-CXCR6 and CX3CL1-CX3CR1 axes, but not the CCL5-CCR1/3/5 axis, resulted in reduced insulitis and lower T1D incidence. Further, we found that the receptors for the differentially expressed chemokines CXCL10, CXCL16 and CX3CL1 are distributed unevenly among islet autoantigen-specific T cells, which explains why the interference with just one chemokine axis cannot completely abrogate insulitis and T1D.

Mechanistic role of Syzygium cumini (L.) Skeels in glycation induced diabetic nephropathy via RAGE–NF–κB pathway and extracellular proteins modifications: A molecular approach

Ethnopharmacology relevanceSyzygium cumini (L.) Skeels (SC), an ancient medicinal plant, is used as a complementary and alternative medicine for treating diabetes mellitus and its associated complications, such as diabetic nephropathy (DN). Phytochemicals present in SC homeopathic formulations possess anti-glycemic, anti-glycation, anti-inflammatory, and antioxidant properties. Additionally, the non-enzymatic formation of advanced glycation end products (AGEs) increases during hyperglycemia in diabetes. AGEs interaction with their receptor of AGEs (RAGE) promotes inflammation via Nuclear Factor-κB (NF-κB) and the accumulation of Extracellular Matrix (ECM) proteins, contributing to the renal dysfunction in DN. However, the molecular mechanism through which SC formulations interact with the AGEs-RAGE–NF–κB pathway has not yet been investigated. AimThis study aims to examine the impact of SC formulations on the RAGE–NF–κB pathway and ECM protein modifications in glycation-induced DN using a molecular approach. Materials and methodsHuman serum albumin (10 mg/ml) was glycated with MGO (55 mM) in the presence of SC formulations - Mother tincture (MT), 30C, 200C for 7 days. Glycated samples were added to renal cells (HEK 293) for 24 h. Subsequently, cellular gene and protein expressions of RAGE, NF-κB, vascular endothelial growth factor (VEGF), interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), collagen IV (Col IV), and fibronectin were determined using RT-qPCR and Western blot analysis. The immunofluorescence, luciferase assay, and chromatin immunoprecipitation techniques were employed to gain insights into glycation-induced NF-κB nuclear translocation, transcriptional activity, and its effect on RAGE promoter activity in SC-treated cells. ResultsSC formulations significantly downregulated glycation-induced elevated levels of RAGE and NF-κB. Mechanistically, SC formulations prevented NF-κB nuclear translocation, transcriptional activity, and RAGE promoter activity. Also, SC formulations significantly attenuated glycation-enhanced expressions of inflammatory cytokines (IL-6, TNF-α, and VEGF) and ECM proteins (Col IV and fibronectin). ConclusionOur findings enlighten the molecular mechanism of SC in DN by targeting the AGEs-RAGE–NF–κB signaling pathway, inflammatory responses, and ECM accumulation. Hence, the study validates the protective role of SC formulations and signifies its novel potential for treating DN.

Unripe Pear Extract Suppresses UVB-Induced Skin Photoaging in Hairless Mice and Keratinocytes.

Our study aimed to investigate whether unripe pear extract (UP) could provide protection against UVB-induced damage to both mouse skin and keratinocytes. We observed that UVB exposure, a common contributor to skin photoaging, led to wrinkle formation, skin dryness, and inflammation in mice. Nevertheless, these effects were mitigated in the groups of UVB-irradiated mice treated with UP. Moreover, UP treatment at 400 μg/mL increased the antioxidant enzyme activities (sodium dodecyl sulfate, 2.22-fold higher; catalase, 2.91-fold higher; GPx, 1.96-fold higher) along with sphingomyelin (1.58-fold higher) and hyaluronic acid (1.31-fold higher) levels in UVB-irradiated keratinocytes. In the keratinocytes irradiated with UVB, UP 400 μg/mL resulted in reduced cytokine production (TNF-α, 33.2%; IL-1β, 45.3%; IL-6, 33.4%) and the expression of inflammatory pathway-related proteins. The findings indicate that UP has a direct protective effect on UVB-irradiated keratinocytes and is also able to shield against photoaging induced by UVB. Hence, it is suggested that UP could contribute to improved skin health by averting skin photoaging.

Activin suppresses the expression of inflammatory genes and signaling proteins in human leukemia monocytic THP-1 cells.

Activin regulates inflammation, cell proliferation, immune response, wound repair, and endocrine function. In this study, we investigated the effect of activin on inflammatory genes in THP-1 cells and the involvement of NF-κB, AKT, and mitogen-activated protein kinase (MAPK) signaling. Cell viability was determined using a colorimetric assay with the MTS/PES solution. The mRNA levels were analyzed using reverse transcription-quantitative polymerase chain reaction. The expression of NF-κB, AKT, and MAPK signaling proteins was measured using immunoblot analysis. Activin A did not affect THP-1 cell viability at concentrations below 50 ng/ml. Activin decreased the mRNA expression of cytokines (interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α), toll-like receptor 4 (TLR4), and matrix metallo-proteinases (MMP)-9 proteins but did not affect IL-8 expression. Activin increased the expression of TLR2 and MMP-2. In addition, activin inhibited the phosphorylation of NF-κB p65, AKT, and MAPK (c-Jun N-terminal kinase (JNK), extracellular signal-regulated kinase (ERK), and p38 MAPK) signaling proteins. Our results suggest that activin may be involved in anti-inflammation by inhibiting inflammatory gene expression and regulating NF-κB and AKT/MAPK signaling.

Jingfang Granule mitigates Coxsackievirus B3-induced myocardial damage by modulating mucolipin 1 expression

Ethnopharmacological relevanceJingfang Granules (JFG) originate from the traditional herbal formula Jingfang Baidu powder. It has the effects of inducing sweating and dispelling wind. It is a classic medication used for treating external pathogenic factors and viral diseases. However, the therapeutic mechanism of JFG for viral myocarditis needs further clarification. Aim of the studyThis study aimed to explore the therapeutic efficacy of JFG on coxsackievirus B3-induced viral myocarditis (VMC), along with the elucidation of its underlying mechanisms. Materials and methodsC57 BL/6JNifdc mice were divided randomly into several groups: control, model, Jingfang Granule groups (0.23, 0.46, and 0.69 g/20g, respectively), and a positive group (oseltamivir, 19.33 mg/kg). Following the establishment of the VMC model, the mice underwent an 8 -week treatment regimen. Pathological alterations in cardiac tissues and inflammatory protein expression were monitored. Differential gene analysis was conducted utilizing transcriptomic techniques. The differential gene mucolipin 1 (Mcoln1) was knocked down by transfection with siRNA in H9C2 cell, and investigative techniques such as immunoblotting, qRT-PCR, immunofluorescence, JC-1 staining, reactive oxygen species (ROS) detection, and mitochondrial stress testing were employed to examine its mechanism of action. ResultsJFG significantly mitigates the pathological damage observed in the cardiac tissues of CVB3-induced VMC mice and attenuates the expression of inflammatory genes. Subsequently, differentially expressed genes are identified through transcriptomic analysis and validated via PCR. Among these, the upregulation of Mcoln1 promotes autophagy, facilitating the clearance of damaged mitochondria and excessive ROS. This has been substantiated through in vitro experiments. Excessive ROS precipitates a reduction in mitochondrial membrane potential, instigating cell apoptosis. In accordance with TUNEL staining results, JFG acts to inhibit cell apoptosis. To ascertain whether Mcoln1 is a crucial target for JFG in treating VMC, Mcoln1 was suppressed in H9C2 cells. The suppression of Mcoln1 hinders the elevation in autophagy levels post-JFG treatment, obstructs the enhancement of mitochondrial function, and impedes the clearance of ROS. Furthermore, the inhibitory effect of JFG on cell apoptosis is attenuated. ConclusionThe research findings indicate that JFG has a protective effect on CVB3-induced H9C2 cell injury. JFG may exert its effects in VMC treatment by enhancing autophagy to suppress cell apoptosis through the mitochondrial pathway, thereby counteracting cell damage.

(E)-2-methoxy-4-(3-(4-methoxyphenyl)prop-1-en-1-yl)phenol alleviates inflammatory responses in LPS-induced mice liver sepsis through inhibition of STAT3 phosphorylation

Sepsis is a life-threatening disease with limited treatment options, and the inflammatory process represents an important factor affecting its progression. Many studies have demonstrated the critical roles of signal transducer and activator of transcription 3 (STAT3) in sepsis pathophysiology and pro-inflammatory responses. Inhibition of STAT3 activity may therefore represent a promising treatment option for sepsis. We here used a mouse model to demonstrate that (E)-2-methoxy-4-(3-(4-methoxyphenyl)prop-1-en-1-yl)phenol (MMPP) treatment prevented the liver sepsis-related mortality induced by 30 mg/kg lipopolysaccharide (LPS) treatment and reduced LPS-induced increase in alanine transaminase, aspartate transaminase, and lactate dehydrogenase levels, all of which are markers of liver sepsis progression. These recovery effects were associated with decreased LPS-induced STAT3, p65, and JAK1 phosphorylation and proinflammatory cytokine (interleukin 1 beta, interleukin 6, and tumor necrosis factor alpha) level; expression of cyclooxygenase-2 and induced nitric oxide synthase were also reduced by MMPP. In an in vitro study using the normal liver cell line THLE-2, MMPP treatment prevented the LPS-induced increase of STAT3, p65, and JAK1 phosphorylation and inflammatory protein expression in a dose-dependent manner, and this effect was enhanced by combination treatment with MMPP and STAT3 inhibitor. The results clearly indicate that MMPP treatment prevents LPS-induced mortality by inhibiting the inflammatory response via STAT3 activity inhibition. Thus, MMPP represents a novel agent for alleviating LPS-induced liver sepsis.

Comparative Morphology of Island and Inland Agastache rugosa and Their Gastroprotective Effects in EtOH/HCl-Induced Gastric Mucosal Gastritis.

Agastache rugosa Kuntze (Lamiaceae; Labiatae), a medicinal and functional herb used to treat gastrointestinal diseases, grows well both on islands and inland areas in South Korea. Thus, we aimed to reveal the morphological and micromorphological differences between A.rugosa grown on island and inland areas and their pharmacological effects on gastritis in an animal model by combining morphological and mass spectrophotometric analyses. Morphological analysis showed that island A.rugosa had slightly smaller plants and leaves than inland plants; however, the density of all types of trichomes on the leaves, petioles, and stems of island A.rugosa was significantly higher than that of inland plants. The essential oil component analysis revealed that pulegone levels were substantially higher in island A.rugosa than in inland A.rugosa. Despite the differences between island and inland A.rugosa, treatment with both island and inland A.rugosa reduced gastric damages by more than 40% compared to the gastritis induction group. In addition, expression of inflammatory protein was reduced by about 30% by treatment of island and inland A.rugosa. The present study demonstrates quantitative differences in morphology and volatile components between island and inland plants; significant differences were not observed between the gastritis-inhibitory effects of island and inland A.rugosa, and the efficacy of island A.rugosa was found to be similar to that of A.rugosa grown in inland areas.

Diethylaminoethyl-dextran and monocyte cell membrane coated 1,8-cineole delivery system for intracellular delivery and synergistic treatment of atherosclerosis

We have developed a biomimetic delivery system termed the Monocyte Cell Membrane-Coated 1,8-Cineole Biomimetic Delivery System (MM-CIN-BDS or BDS), which integrates diethylaminoethyl-dextran (DEAE) and monocyte cell membrane (MM). This innovative approach enhances the cellular uptake efficiency of 1,8-cineole (CIN) and facilitates targeted therapy for atherosclerosis. Our findings demonstrate the successful modification of the drug carrier with DEAE and MM, as validated by measurements of particle size, zeta potential, microscopic morphology, and western blotting analyses. Notably, cellular uptake experiments unveil a significant enhancement in cellular uptake efficiency due to DEAE modification. However, the introduction of monocyte cell membranes diminishes this effect in normal human umbilical vein endothelial cells (HUVECs), although this efficiency is notably restored in HUVECs activated with lipopolysaccharide (LPS). Through in vivo imaging investigations, we observe that the MM coating augments distribution in the spleen, brain, and atherosclerotic plaques, while concurrently diminishing distribution in the heart and kidneys. Animal studies corroborate these findings, illustrating that MM-CIN-BDS treatment curtails lipid parameters, dampens the expression of inflammatory factors and proteins, mitigates vascular tissue damage, and ultimately reduces the extent of atherosclerotic lesion areas. To encapsulate, DEAE emerges as an especially adept agent for modifying drug carriers with suboptimal cellular uptake efficiency in the realm of cardiovascular diseases. The potential therapeutic promise of MM-CIN-BDS for atherosclerosis treatment is evident from our research.

Cerclage prevents ascending intrauterine infection in pregnant mice

The treatment for cervical insufficiency is cerclage surgery. Although cerclage is a common therapy for prevention of preterm birth, there is no consensus about its mechanism of efficacy. Previous investigators have hypothesized that cerclage prevents preterm birth by improving the cervical barrier to ascending infection. However, this hypothesis is difficult to study in human pregnancy. In a mouse model of ascending infection, we hypothesized that a cerclage improves the cervical barrier leading to decreased ascending intrauterine infection and inflammation. An abdominal cerclage was studied because a vaginal cerclage is not feasible in mice. To perform an abdominal cerclage, laparotomy was performed on timed, pregnant C57BL/6 mice on gestational day 10 (E10). A 6-0 silk suture was placed around the cervix just below the junction of the 2 uterine horns. Sham controls received the same surgery, but no cerclage was placed. To track ascending infection nonpathogenic E coli K12 was genetically modified to express bioluminescence. On E15, bioluminescent E coli K12 (20 μL of 1×109 bacteria) was inoculated into the vagina. Whole-body bioluminescence imaging was performed 0.5 hours and 24 hours after inoculation. To assess intrauterine inflammation, pathogenic E coli K1 was used. On E15, bacterial inoculums of E coli K1 (20 μL of 1×104 bacteria) were vaginally administered. Samples of uterus, placenta, and fetal membranes were collected 24 hours after inoculation. Gene expression of inflammation-related proteins was compared between 3 groups: (1) sham control surgery+ inoculation of phosphate-buffered saline (PBS), (2) sham control surgery+ inoculation of E coli K1, and (3) cerclage surgery+ inoculation of E coli K1. Abdominal cerclage was well tolerated. No cases of preterm birth were seen following abdominal cerclage. Whole-body bioluminescent imaging performed 0.5 hours post inoculation showed a strong luminescence signal in the vaginal region of mice in both control and experimental groups indicating successful bacteria inoculation. Twenty-four hours after inoculation, bioluminescent signal was seen ascending into the uterine horns in all control mice. However, in mice with abdominal cerclages, no bioluminescent signal was seen after 24 hours. When the reproductive tissues were imaged separately in control mice, strong bioluminescence signal was detected in the placenta, fetal membranes, and uterus. Gene expression studies showed that cerclage significantly decreased the expression of inflammatory proteins induced by E coli K1 in the uterus, placenta, and fetal membranes. In this mouse model of ascending intrauterine infection, abdominal cerclage prevented ascending infection of E coli. In addition, abdominal cerclage prevented expression of inflammatory cytokines in the uterus, placenta, and membranes of mice. The study provides evidence for a potential mechanism of cerclage success in human pregnancy.

PEDF inhibits LPS-induced acute lung injury in rats and promotes lung epithelial cell survival by upregulating PPAR-γ

BackgroundThe progression of acute lung injury (ALI) involves numerous pathological factors and complex mechanisms, and cause the destruction of epithelial and endothelial barriers. Pigment epithelium-derived factor (PEDF) is an angiogenesis inhibitor and a potential anti-inflammatory factor. The purpose of this study was to investigate the effect of PEDF on lipopolysaccharide (LPS)-induced ALI in rats.MethodsIn vivo, pathological and injury related factors examination were performed on rat lung to investigate the effect of PEDF on ALI. In vitro, the effect of PEDF on inflammatory injury and apoptosis of lung epithelial type II RLE-6TN cell was evaluated, and the expression of inflammatory factors and related pathway proteins and PPAR-γ (in the presence or absence of PPAR-γ inhibitors) were analyzed.ResultsIn vivo results showed that PEDF inhibited the inflammatory factor expression (TNF-α, IL-6 and IL-1β) and progression of ALI and reduced lung cell apoptosis in rats. In vitro results showed that PEDF could effectively inhibit LPS-stimulated inflammatory damage and apoptosis of RLE-6TN cells. PEDF inhibited the RLE-6TN cell injury by enhancing the expression of PPAR-γ.ConclusionsPEDF is an anti-inflammatory factor, which can inhibit apoptosis of lung epithelial cells by upregulating the expression of PPAR-γ and reducing LPS-induced ALI in rats.

Degradation profile of environmental pollutant 17β-estradiol by human intestinal fungus Aspergillus niger RG13B1 and characterization of genes involved in its degradation

Environmental hormones have attracted more attention because of their adverse impact on the health and ecological security of human. Biodegradation is still an efficient tactics to remove environmental hormones, but human intestinal microbes remain to be elucidated in the role of their degradation. In the present work, we intended to perform the in vitro experiment for investigating the degradation of 17β-estradiol, the main environmental estrogen, by human intestinal microflora Aspergillus niger RG13B1. Its degradation led to the production of eighteen metabolites characterized by 1H, 13C, and 2D NMR, and HRMS spectra, including nine new (1–9) and nine known metabolites (10–18). Based on their structures, the degradation pathway of 17β-estradiol mediated by A. niger RG13B1 involved hydroxylation, oxidation, methylation, acetylation, and dehydrogenation, especially infrequent lactylation, and the key degradation enzymes were found in the gene cluster of A. niger. In addition, we found that metabolite 12 interacted with amino acid residues Lys37, Gln39, Lys93, and Asn115 of NF-κB p65 to suppress expressions of inflammatory genes or proteins, exerting its anti-inflammatory effect. This study first illustrated the role of human gut microbe in 17β-estradiol degradation and provided new insights into its degradation mechanism by A. niger RG13B1.

Genipin relieves diabetic retinopathy by down-regulation of advanced glycation end products via the mitochondrial metabolism related signaling pathway.

Glycation is an important step in aging and oxidative stress, which can lead to endothelial dysfunction and cause severe damage to the eyes or kidneys of diabetics. Inhibition of the formation of advanced glycation end products (AGEs) and their cell toxicity can be a useful therapeutic strategy in the prevention of diabetic retinopathy (DR). Gardenia jasminoides Ellis (GJE) fruit is a selective inhibitor of AGEs. Genipin is an active compound of GJE fruit, which can be employed to treat diabetes. To confirm the effect of genipin, a vital component of GJE fruit, in preventing human retinal microvascular endothelial cells (hRMECs) from AGEs damage in DR, to investigate the effect of genipin in the down-regulation of AGEs expression, and to explore the role of the CHGA/UCP2/glucose transporter 1 (GLUT1) signal pathway in this process. In vitro, cell viability was tested to determine the effects of different doses of glucose and genipin in hRMECs. Cell Counting Kit-8 (CCK-8), colony formation assay, flow cytometry, immunofluorescence, wound healing assay, transwell assay, and tube-forming assay were used to detect the effect of genipin on hRMECs cultured in high glucose conditions. In vivo, streptozotocin (STZ) induced mice were used, and genipin was administered by intraocular injection (IOI). To explore the effect and mechanism of genipin in diabetic-induced retinal dysfunction, reactive oxygen species (ROS), mitochondrial membrane potential (MMP), and 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-d-glucose (2-NBDG) assays were performed to explore energy metabolism and oxidative stress damage in high glucose-induced hRMECs and STZ mouse retinas. Immunofluorescence and Western blot were used to investigate the expression of inflammatory cytokines [vascular endothelial growth factor (VEGF), SCG3, tumor necrosis factor-alpha (TNF-α), interleukin (IL)-1β, IL-18, and nucleotide-binding domain, leucine-rich-containing family, pyrin domain-containing 3 (NLRP3)]. The protein expression of the receptor of AGEs (RAGE) and the mitochondria-related signal molecules CHGA, GLUT1, and UCP2 in high glucose-induced hRMECs and STZ mouse retinas were measured and compared with the genipin-treated group. The results of CCK-8 and colony formation assay showed that genipin promoted cell viability in high glucose (30 mmol/L D-Glucose)-induced hRMECs, especially at a 0.4 μmol/L dose for 7 d. Flow cytometry results showed that high glucose can increase apoptosis rate by 30%, and genipin alleviated cell apoptosis in AGEs-induced hRMECs. A high glucose environment promoted ATP, ROS, MMP, and 2-NBDG levels, while genipin inhibited these phenotypic abnormalities in AGEs-induced hRMECs. Furthermore, genipin remarkably reduced the levels of the pro-inflammatory cytokines TNF-α, IL-1β, IL-18, and NLRP3 and impeded the expression of VEGF and SCG3 in AGEs-damaged hRMECs. These results showed that genipin can reverse high glucose induced damage with regard to cell proliferation and apoptosis in vitro, while reducing energy metabolism, oxidative stress, and inflammatory injury caused by high glucose. In addition, ROS levels and glucose uptake levels were higher in the retina from the untreated eye than in the genipin-treated eye of STZ mice. The expression of inflammatory cytokines and pathway protein in the untreated eye compared with the genipin-treated eye was significantly increased, as measured by Western blot. These results showed that IOI of genipin reduced the expression of CHGA, UCP2, and GLUT1, maintained the retinal structure, and decreased ROS, glucose uptake, and inflammation levels in vivo. In addition, we found that SCG3 expression might have a higher sensitivity in DR than VEGF as a diagnostic marker at the protein level. Our study suggested that genipin ameliorates AGEs-induced hRMECs proliferation, apoptosis, energy metabolism, oxidative stress, and inflammatory injury, partially via the CHGA/UCP2/GLUT1 pathway. Control of advanced glycation by IOI of genipin may represent a strategy to prevent severe retinopathy and vision loss.

A randomized, placebo-controlled, trial to assess the photosensitizing, phototoxic and carcinogenic potential of hydrochlorothiazide in healthy volunteers.

Pharmacovigilance reports, associating hydrochlorothiazide (HCT) with skin cancer, resulted in a significant decrease of HCT prescriptions for hypertension and heart failure. Whether HCT exhibits phototoxic properties thereby causing skin cancer remains unknown. This study aimed to examine the photosensitizing, phototoxic and carcinogenic potential of HCT in a randomized, placebo-controlled, double-blind trial in vivo and also in vitro . The trial assigned 30 healthy, normotensive adult volunteers in a 2:1 ratio to either HCT 25 mg/day or placebo for 15 days. Photosensitivity of the skin with and without the effect of HCT treatment were assessed. Following whole-body ultraviolet A (UVA) and B (UVB, 311 nm) irradiation, phototoxic and carcinogenic reactions by measuring urinary excretion of pyrimidine dimers were evaluated. For the in-vitro studies, human keratinocytes (HaCaT) were incubated with HCT, irradiated with UVB, and analysed for markers of inflammation, apoptosis and carcinogenesis. Skin photosensitivity following exposure to UVA and UVB remained unchanged from baseline to 15-day follow-up in both groups (UVA change HCT 0.0 J/cm 2 vs. placebo 0.0 J/cm 2 ; P = 0.99; UVB change HCT 0.0 J/cm 2 vs. placebo -0.2 J/cm 2 ; P = 0.06). Pyrimidine dimers were not detected in either group. In vitro , combination of HCT and UVB irradiation did not induce the expression of oxidative stress marker proteins, inflammatory proteins, apoptotic proteins or activation of oncoproteins. HCT did not increase photosensitivity for UVA or UVB in healthy volunteers compared with placebo, and was not associated with phototoxic or carcinogenic reactions. In vitro , HCT was also not associated with phototoxicity or carcinogenesis (NCT04654312).

Methyl lucidone inhibits airway inflammatory response by reducing TAK1 activity in human bronchial epithelial NCI–H292 cells

BackgroundMethyl lucidone (ML), a methyl derivative of lucidone, has anti-inflammatory properties. However, the molecular mechanisms that reduce the inflammatory effect of ML in human lung epithelial cells remain unkown. This study aimed to elucidate the molecular mechanisms underlying the anti-inflammatory effect of ML. MethodsFour compounds (ML, methyl linderone, kanakugiol, and linderone) from Lindera erythrocarpa Makino were evaluated for their ability to reduce MUC5AC secretion levels in phorbol-12-myristate-13-acetate (PMA)-stimulated NCI–H292 cells using ELISA. The expression and secretion levels of inflammatory response-related proteins were analyzed using quantitative reverse transcription-PCR, ELISA, and western blotting. To determine whether ML directly regulates TGF-β-activated kinase 1 (TAK1), we performed an in vitro kinase assay. ResultsML treatment effectively reduced the levels of inflammatory cytokines, including interleukin-1β and TNF-α, increased by stimulation. Furthermore, ML downregulated the pathway cascade of both IκB kinase (IKK)/NF-κB and p38 mitogen-activated protein (MAP) kinase/CREB by inhibiting the upstream kinase TAK1. An in vitro kinase analysis confirmed that ML treatment significantly reduced the kinase activity of TAK1. ConclusionML pretreatment repressed the PMA-stimulated inflammation reaction by reducing the TAK1-mediated IKK/NF-κB and p38 MAP kinase/CREB signaling. These findings suggest that ML may improve respiratory health and can be used as a dietary supplement or functional food to prevent inflammatory lung diseases.

Matcha-silver nanoparticles reduce gamma radiation-induced oxidative and inflammatory responses by activating SIRT1 and NLRP-3 signaling pathways in the Wistar rat spleen.

The biogenic synthesis of nanoparticles has drawn significant attention. The spleen is the largest lymphatic organ that is adversely impacted during irradiation. The current study was designated to evaluate the possible anti-inflammatory effect of matcha-silver nanoparticles (M-AgNPs) to reduce inflammation associated with γ-radiation induced-oxidative stress and inflammation in rats' spleen. Silver nanoparticles (AgNPs) were synthesized by biogenic synthesis using a green sonochemical method from matcha (M) green tea. The obtained M-AgNPs were extensively characterized by dynamic light scattering, transmission electron microscopy, thermogravimetric analysis, and Fourier-transform infrared spectroscopy. Using zetasizer analysis, the surface charge, particle size, and radical scavenging DPPH assay of M-AgNPs were also examined. Biocompatibility and cytotoxicity were analyzed by MTT assay, and the IC50 was calculated. Four groups of 24 Wistar rats each had an equal number of animals. The next step involved measuring the levels of oxidative stress markers in the rat splenic tissue. Additionally, the amounts of inflammatory protein expression were evaluated using the ELISA analysis. The results indicated the formation of spherical nanoparticles of pure Ag° coated with matcha polyphenols at the nanoscale, as well as uniform monodisperse particles suited for cellular absorption. Results revealed that M-AgNPs improved all biochemical parameters. Furthermore, M-AgNPs relieve inflammation by reducing the expression of NOD-like receptor family pyrin domain-containing 3 (NLRP3), interleukin-1β (IL-1β), and enhancing the levels of ileSnt information regulator 1 (SIRT1). Histopathological examinations demonstrated the ability of M-AgNPs to overcome the damage consequent to irradiation and recover the spleen's cellular structure. These results confirmed that matcha is a potential biomaterial for synthesizing AgNPs, which can be exploited for their anti-inflammatory activity.

Prolyl oligopeptidase inhibition ameliorates experimental pulmonary fibrosis both in vivo and in vitro

BackgroundPulmonary fibrosis is a progressive disease characterized by lung remodeling due to excessive deposition of extracellular matrix. Although the etiology remains unknown, aberrant angiogenesis and inflammation play an important role in the development of this pathology. In this context, recent scientific research has identified new molecules involved in angiogenesis and inflammation, such as the prolyl oligopeptidase (PREP), a proteolytic enzyme belonging to the serine protease family, linked to the pathology of many lung diseases such as pulmonary fibrosis. Therefore, the aim of this study was to investigate the effect of a selective inhibitor of PREP, known as KYP-2047, in an in vitro and in an in vivo model of pulmonary fibrosis.MethodsThe in vitro model was performed using human alveolar A549 cells. Cells were exposed to lipopolysaccharide (LPS) 10 μg/ml and then, cells were treated with KYP-2047 at the concentrations of 1 μM, 10 μM and 50 μM. Cell viability was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium (MTT) bromide colorimetric assay, while inflammatory protein expression was assessed by western blots analysis. The in vivo model was induced in mice by intra-tracheal administration of bleomycin (1 mg/kg) and then treated intraperitoneally with KYP-2047 at doses of 1, 2.5 and 5 mg/kg once daily for 12 days and then mice were sacrificed, and lung tissues were collected for analyses.ResultsThe in vitro results demonstrated that KYP-2047 preserved cell viability, reduced inflammatory process by decreasing IL-18 and TNF-α, and modulated lipid peroxidation as well as nitrosative stress. The in vivo pulmonary fibrosis has demonstrated that KYP-2047 was able to restore histological alterations reducing lung injury. Our data demonstrated that KYP-2047 significantly reduced angiogenesis process and the fibrotic damage modulating the expression of fibrotic markers. Furthermore, KYP-2047 treatment modulated the IκBα/NF-κB pathway and reduced the expression of related pro-inflammatory enzymes and cytokines. Moreover, KYP-2047 was able to modulate the JAK2/STAT3 pathway, highly involved in pulmonary fibrosis.ConclusionIn conclusion, this study demonstrated the involvement of PREP in the pathogenesis of pulmonary fibrosis and that its inhibition by KYP-2047 has a protective role in lung injury induced by BLM, suggesting PREP as a potential target therapy for pulmonary fibrosis. These results speculate the potential protective mechanism of KYP-2047 through the modulation of JAK2/STAT3 and NF-κB pathways.Graphical abstract

Renal ischemia alters the transcriptomic and epigenetic profile of inflammatory genes in swine scattered tubular-like cells.

Scattered tubular-like cells (STCs) are differentiated renal tubular cells that during recovery from ischemic injury dedifferentiate to repair other injured renal cells. Renal artery stenosis (RAS), often associated with chronic inflammatory injury, compromises the integrity and function of STCs, but the underlying mechanisms remain unknown. We hypothesized that RAS alters the transcriptomic and epigenetic profile of inflammatory genes in swine STCs. STCs were harvested from pig kidneys after 10 weeks of RAS or sham (n=6 each). STC mRNA profiles of inflammatory genes were analyzed using high-throughput mRNA-sequencing (seq) and their DNA methylation (5mC) and hydroxymethylation (5hmC) profiles by DNA immunoprecipitation and next-generation sequencing (MeDIP-seq) (n=3 each), followed by an integrated (mRNA-seq/MeDIP-seq) analysis. STC protein expression of candidate differentially expressed (DE) genes and common proinflammatory proteins were subsequently assessed in vitro before and after epigenetic (Bobcat339) modulation. mRNA-seq identified 57 inflammatory genes up-regulated in RAS-STCs versus Normal-STCs (>1.4 or <0.7-fold, P<0.05), of which 14% exhibited lower 5mC and 5% higher 5hmC levels in RAS-STCs versus Normal-STCs, respectively. Inflammatory gene and protein expression was higher in RAS-STCs compared with Normal-STCs but normalized after epigenetic modulation. These observations highlight a novel modulatory mechanism of this renal endogenous repair system and support development of epigenetic or anti-inflammatory therapies to preserve the reparative capacity of STCs in individuals with RAS.