Ginsenoside Rg1 Alleviates Sepsis-Induced Acute Lung Injury by Reducing FBXO3 Stability in an m6A-Dependent Manner to Activate PGC-1α/Nrf2 Signaling Pathway.

BACKGROUND: Sepsis-induced acute lung injury (ALI) is one of the serious life-threatening complications of sepsis and is pathologically associated with mitochondrial dysfunction. Ginsenoside Rg1 has good therapeutic effects on ALI. Herein, the pharmacological effects of Rg1 in sepsis-induced ALI were investigated. METHODS: Sepsis-induced ALI models were established by CLP operation and LPS treatment. HE staining was adopted to analyze lung pathological changes. The expression and secretion of cytokines were measured by RT-qPCR and ELISA. Cell viability and apoptosis were assessed by MTT assay, flow cytometry and TUNEL staining. ROS level and mitochondrial membrane potential (MMP) were analyzed using DHE probe and JC-1 staining, respectively. FBXO3 m6A level was assessed using MeRIP assay. The interactions between FBXO3, YTHDF1, and PGC-1α were analyzed by Co-IP or RIP. RESULTS: Rg1 administration ameliorated LPS-induced epithelial cell inflammation, apoptosis, and mitochondrial dysfunction in a dose-dependent manner. Mechanically, Rg1 reduced PGC-1α ubiquitination modification level by inhibiting FBXO3 expression m6A-YTHDF1 dependently. As expected, Rg1's mitigative effect on LPS-induced inflammation, apoptosis and mitochondrial dysfunction in lung epithelial cells was abolished by FBXO3 overexpression. Moreover, FBXO3 upregulation eliminated the restoring effect of Rg1 on CLP-induced lung injury in rats. CONCLUSION: Rg1 activated PGC-1α/Nrf2 signaling pathway by reducing FBXO3 stability in an m6A-YTHDF1-dependent manner to improve mitochondrial function in lung epithelial cells during sepsis-induced ALI progression.

Experimental necrotizing enterocolitis using oral lipopolysaccharide and protective role of breastmilk. / Enterocolitis necrotizante experimental con lipopolisacárido oral y función protectora de la leche materna.

INTRODUCTION: Necrotizing enterocolitis (NEC) is a life-threatening condition that afflicts neonates. Breastfeeding has demonstrated to play a protective role against it. By administering lipopolysaccharides (LPS) orally in newborn rats (NBR), we have developed an experimental model to induce NEC-like gut damage. Our aim was to assess the macroscopic and microscopic appearance of the gut, to evaluate the presence of NEC and study the role of breast milk (BM). MATERIALS AND METHODS: NBR were divided into 3 groups: Group A (control, n= 10) remained with the mother, group B (LPS, n= 25) was isolated after birth, gavage-fed with special rat formula and oral LPS, then submitted to stress (hypoxia after gavage) and group c (BM, n= 12) was breastfed once after birth, then isolated, and submitted to stress like group B. On day 4, NBR were sacrificed, and intestine was harvested and assessed. RESULTS: In the control group NEC was not present either macroscopically or histologically. Both groups submitted to stress (B and C) presented a global incidence of NEC of 73%. Most of group B developed histologic signs of NEC (85%) and group C showed a statistically lower incidence of NEC (50%, p= 0.04), playing the BM a protective role against NEC (OR= 0.19; 95% CI: 0.40-0.904). CONCLUSION: Our model showed a significant incidence of NEC in NBR (73%) with the same protective role of BM as in newborn humans, achieving a reliable and reproducible experimental NEC model. This will allow us to investigate new potential therapeutic targets for a devastating disease that currently lacks treatment.

INTRODUCCION: La enterocolitis necrotizante (ECN) es una enfermedad potencialmente mortal que afecta a los neonatos, y frente a la que la leche materna ha demostrado tener un papel protector. Administrando lipopolisacáridos (LPS) por vía oral en ratas recién nacidas (RRN), hemos desarrollado un modelo experimental para inducir un daño intestinal similar al que provoca la ECN con objeto de evaluar el aspecto macroscópico y microscópico del intestino, y de ese modo, analizar la presencia de ECN y estudiar el papel que desempeña la leche materna (LM). MATERIAL Y METODOS: Las RRN se dividieron en tres grupos: el grupo A (control, n= 10) permaneció con su madre; el grupo B (LPS, n= 25) fue aislado tras el nacimiento, alimentado por sonda con una fórmula especial para ratas y LPS oral, y sometido a estrés (hipoxia tras sonda); y el grupo C (LM, n= 12) fue alimentado con leche materna tras el nacimiento y posteriormente aislado y sometido a estrés al igual que el grupo B. El día 4 se sacrificó a las RRN y se recuperaron sus intestinos para su posterior evaluación. RESULTADOS: En el grupo de control, no se observó ECN ni macroscópica ni histológicamente, mientras que los dos grupos sometidos a estrés (B y C) presentaron una incidencia global de la ECN del 73%. La mayoría de los sujetos del grupo B desarrollaron signos histológicos de ECN (85%), y los del grupo C registraron una incidencia de la ECN estadísticamente menor (50%, p= 0,04), lo que significa que la LM desempeña una función protectora frente a la ECN (OR= 0,19; IC 95%: 0,40-0,904). CONCLUSION: Nuestro modelo reveló una incidencia significativa de la ECN en RRN (73%), desempeñando la LM la misma función protectora que en el caso de los humanos recién nacidos, lo que significa que este modelo experimental de ECN es fiable y reproducible. Gracias a dicho logro, podremos investigar nuevos y potenciales objetivos terapéuticos para una peligrosa enfermedad que, a día de hoy, carece de tratamiento.

Upregulated ATF1 Promotes Lipopolysaccharide Induced Inflammatory Response and Inhibits Osteogenic Differentiation of Human Periodontal Ligament Cells by Regulating NF-κB Pathway.

BACKGROUND: Periodontitis is a chronic inflammatory disease resulting from bacterial plaque infection. While the involvement of activating transcription factor 1 (ATF1) has been extensively explored in various human diseases, its specific role in periodontitis remains unclear. This study aims to elucidate the expression and biological function of ATF1 in the context of periodontitis. METHODS: Primary human periodontal ligament cells (hPDLCs) were procured from clinical samples and subsequently characterized. Following treatment with P. gingivalis lipopolysaccharide (LPS, 10 µg/mL), hPDLCs underwent transfection with either ATF1 vector or siRNA. The expression levels of ATF1 in LPS-treated hPDLCs or transfected cells were evaluated through real-time quantitative polymerase chain reaction (RT-qPCR) and western blot assay. Inflammatory factors, including interleukin-6 (IL-6), interleukin-8 (IL-8), tumor necrosis factor-alpha (TNF-α), and interleukin-1beta (IL-1ß), were quantified using Enzyme-linked Immunosorbent Assay (ELISA). The assessment of osteogenic proteins, such as runt-related transcription factor 2 (Runx2), osteopontin (OPN), and osteoprotegerin (OPG), as well as noncanonical nuclear factor-kappaB (NF-κB) pathway-related proteins (p65, p-p65, IkBα, p-IkBα), was conducted using western blot assay. 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry assays were employed to detect cell viability. RESULTS: LPS induced an inflammatory response and hindered the osteogenic differentiation of hPDLCs (p < 0.05, p < 0.01). Furthermore, ATF1 silencing enhanced cell proliferation and suppressed apoptosis in LPS-stimulated hPDLCs (p < 0.05, p < 0.01). ATF1 silencing not only restrained the inflammatory response but also promoted the osteogenic differentiation of LPS-stimulated hPDLCs (p < 0.05, p < 0.01). Importantly, ATF1 silencing effectively blocked the LPS-induced activation of the NF-κB signaling pathway (p < 0.05, p < 0.01, p < 0.001). CONCLUSIONS: ATF1 emerges as a promising treatment option, inhibiting the osteogenic differentiation of hPDLCs and mitigating the inflammatory response by preventing the phosphorylation of the NF-κB signaling pathway.

In vitro and in vivo anti-inflammatory properties of an active fucoidan fraction from Sargassum fusiforme and a fraction-based hydrogel.

In a previous study, we separated an active fucoidan (JHCF4) from acid-processed Sargassum fusiforme, then analyzed and confirmed its structure. In the present study, we investigated the potential anti-inflammatory properties of JHCF4 and a JHCF4-based hydrogel in vitro and in vivo. JHCF4 reliably inhibited nitric oxide (NO) production in LPS-induced RAW 264.7 macrophages, with an IC50 of 22.35 µg/ml. Furthermore, JHCF4 attenuated the secretion of prostaglandin E2, tumor necrosis factor-α, interleukin (IL)-1ß, and IL-6, indicating that JHCF4 regulates inflammatory reactions. In addition, JHCF4 downregulated iNOS and COX-2 and inhibited the activation of the MAPK pathway. According to further in vivo analyses, JHCF4 significantly reduced the generation of reactive oxygen species (ROS), NO production, and cell death in an LPS-induced zebrafish model, suggesting that JHCF4 exhibits anti-inflammatory effects. Additionally, a JHCF4-based hydrogel was developed, and its properties were evaluated. The hydrogel significantly decreased inflammatory and nociceptive responses in carrageenan (carr)-induced mouse paws by reducing the increase in paw thickness and decreasing neutrophil infiltration in the basal and subcutaneous layers of the toe epidermis. These results indicate that JHCF4 exhibits potential anti-inflammatory activity in vitro and in vivo and that JHCF4-based hydrogels have application prospects in the cosmetic and pharmaceutical fields.

The protective effects of naringenin, a citrus flavonoid, non-complexed or complexed with hydroxypropyl-ß-cyclodextrin against multiorgan damage caused by neonatal endotoxemia.

BACKGROUND: Endotoxemia is a severe and dangerous clinical syndrome that results in elevated morbidity, especially in intensive care units. Neonates are particularly susceptible to endotoxemia due to their immature immune systems. There are few effective treatments for neonatal endotoxemia. One group of compounds with potential in the treatment of neonatal inflammatory diseases such as endotoxemia is the flavonoids, mainly due to their antioxidant and anti-inflammatory properties. Among these, naringenin (NGN) is a citrus flavonoid which has already been reported to have anti-inflammatory, antioxidant, anti-nociceptive and anti-cancer effects. Unfortunately, its clinical application is limited by its low solubility and bioavailability. However, cyclodextrins (CDs) have been widely used to improve the solubility of nonpolar drugs and enhance the bioavailability of these natural products. OBJECTIVE: We, therefore, aimed to investigate the effects of NGN non-complexed and complexed with hydroxypropyl-ß-cyclodextrin (HPßCD) on neonatal endotoxemia injuries in a rodent model and describe the probable molecular mechanisms involved in NGN activities. METHOD: We used exposure to a bacterial lipopolysaccharide (LPS) to induce neonatal endotoxemia in the mice. RESULTS: It was found that NGN (100 mg/kg i.p.) exposure during the neonatal period reduced leukocyte migration and decreased pro-inflammatory cytokine (TNF-α, IL-1ß and IL-6) levels in the lungs, heart, kidneys or cerebral cortex. In addition, NGN upregulated IL-10 production in the lungs and kidneys of neonate mice. The administration of NGN also enhanced antioxidant enzyme catalase and SOD activity, reduced lipid peroxidation and protein carbonylation and increased the reduced sulfhydryl groups in an organ-dependent manner, attenuating the oxidative damage caused by LPS exposure. NGN decreased ERK1/2, p38MAPK and COX-2 activation in the lungs of neonate mice. Moreover, NGN complexed with HPßCD was able to increase the animal survival rate. CONCLUSION: NGN attenuated inflammatory and oxidative damage in the lungs, heart and kidneys caused by neonatal endotoxemia through the MAPK signaling pathways regulation. Our results show that NGN has beneficial effects against neonatal endotoxemia and could be useful in the treatment of neonatal inflammatory injuries.

Network pharmacology analysis and experimental validation of Xiao-Qing-Long-Tang's therapeutic effects against neutrophilic asthma.

BACKGROUND: Xiao-Qing-Long-Tang (XQLT), a classical Chinese herbal medicine formula, has been extensively used for allergic asthma treatment. However, there is limited research on its anti-inflammatory effects and mechanisms specifically in neutrophilic asthma (NA). PURPOSE: This study aims to investigate the potential therapeutic effects of XQLT against NA using a combination of network pharmacology and experimental validation. STUDY DESIGN: By utilizing traditional Chinese medicine and disease databases, we constructed an XQLT-asthma network to identify potential targets of XQLT for NA. In the experimental phase, we utilized an ovalbumin (OVA)/lipopolysaccharide (LPS)-induced model for neutrophilic asthma and examined the therapeutic effects of XQLT. RESULTS: Our research identified 174 bioactive components within XQLT and obtained 140 target genes of XQLT against asthma. Functional enrichment analysis revealed that these target genes were primarily associated with inflammation and cytokines. In the experimental validation, mice induced with OVA-LPS showcased eosinophilic and neutrophilic cell infiltration in peri-bronchial areas, elevated levels of IL-4 and IL-17 in both serum and lung, increased percentages of Th2 and Th17 cells in the spleen, as well as elevated levels of CD11b+ and CD103+ dendritic cells (DCs) within the lung. Treatment with XQLT effectively reduced IL-4 and IL-17 levels, decreased the percentages of Th2, Th17, CD11b+, and CD103+ DCs, and improved inflammatory cell infiltrations in lung tissues. These findings serve as a foundation for the potential clinical application of XQLT in neutrophilic asthma.

Possible pharmacological targets and mechanisms of sivelestat in protecting acute lung injury.

Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is a life-threatening syndrome induced by various diseases, including COVID-19. In the progression of ALI/ARDS, activated neutrophils play a central role by releasing various inflammatory mediators, including elastase. Sivelestat is a selective and competitive inhibitor of neutrophil elastase. Although its protective effects on attenuating ALI/ARDS have been confirmed in several models of lung injury, clinical trials have presented inconsistent results on its therapeutic efficacy. Therefore, in this report, we used a network pharmacology approach coupled with animal experimental validation to unravel the concrete therapeutic targets and biological mechanisms of sivelestat in treating ALI/ARDS. In bioinformatic analyses, we found 118 targets of sivelestat against ALI/ARDS, and identified six hub genes essential for sivelestat treatment of ALI/ARDS, namely ERBB2, GRB2, PTK2, PTPN11, ESR1, and CCND1. We also found that sivelestat targeted several genes expressed in human lung microvascular endothelial cells after lipopolysaccharide (LPS) treatment at 4 h (ICAM-1, PTGS2, RND1, BCL2A1, TNF, CA2, and ADORA2A), 8 h (ICAM-1, PTGS2, RND1, BCL2A1, MMP1, BDKRB1 and SLC40A1), and 24 h (ICAM-1). Further animal experiments showed that sivelestat was able to attenuate LPS-induced ALI by inhibiting the overexpression of ICAM-1, VCAM-1, and PTGS2 and increasing the phosphorylation of PTK2. Taken together, the bioinformatic findings and experimentative data indicate that the therapeutic effects of sivelestat against ALI/ARDS mainly focus on the early stage of ALI/ARDS by pharmacological modulation of inflammatory reaction, vascular endothelial injury, and cell apoptosis-related molecules.

Synthesis of urolithin derivatives and their anti-inflammatory activity.

Two series of urolithin derivatives, totally 38 compounds, were synthesized. Their anti-inflammatory activity was investigated by detecting the inhibitory effects on the expression of TNF-α in bone marrow-derived macrophages (BMDMs), showing that 24 of 38 ones reduced the expression of TNF-α. Compound B2, the ring C opened derivative of urolithin B with a butoxycarbonyl substitution in ring A, showed the strongest inhibitory activity compared with that of indomethacin. Furthermore, B2 treatment decreased the expression of pro-inflammatory factors IL-1ß, IL-6, iNOS and COX-2. Mechanically, the anti-inflammatory effect of B2 was related to the inhibition of NF-κB signaling pathway. These results clearly illustrated that B2 hold potential for application as an anti-inflammatory agent. The present study provided a viable approach to modify the gut metabolites for anti-inflammatory drug development.

Phase II Study of Eribulin plus Pembrolizumab in Metastatic Soft-tissue Sarcomas: Clinical Outcomes and Biological Correlates.

PURPOSE: Eribulin modulates the tumor-immune microenvironment via cGAS-STING signaling in preclinical models. This non-randomized phase II trial evaluated the combination of eribulin and pembrolizumab in patients with soft-tissue sarcomas (STS). PATIENTS AND METHODS: Patients enrolled in one of three cohorts: leiomyosarcoma (LMS), liposarcomas (LPS), or other STS that may benefit from PD-1 inhibitors, including undifferentiated pleomorphic sarcoma (UPS). Eribulin was administered at 1.4 mg/m2 i.v. (days 1 and 8) with fixed-dose pembrolizumab 200 mg i.v. (day 1) of each 21-day cycle, until progression, unacceptable toxicity, or completion of 2 years of treatment. The primary endpoint was the 12-week progression-free survival rate (PFS-12) in each cohort. Secondary endpoints included the objective response rate, median PFS, safety profile, and overall survival (OS). Pretreatment and on-treatment blood specimens were evaluated in patients who achieved durable disease control (DDC) or progression within 12 weeks [early progression (EP)]. Multiplexed immunofluorescence was performed on archival LPS samples from patients with DDC or EP. RESULTS: Fifty-seven patients enrolled (LMS, n = 19; LPS, n = 20; UPS/Other, n = 18). The PFS-12 was 36.8% (90% confidence interval: 22.5-60.4) for LMS, 69.6% (54.5-89.0) for LPS, and 52.6% (36.8-75.3) for UPS/Other cohorts. All 3 patients in the UPS/Other cohort with angiosarcoma achieved RECIST responses. Toxicity was manageable. Higher IFNα and IL4 serum levels were associated with clinical benefit. Immune aggregates expressing PD-1 and PD-L1 were observed in a patient that completed 2 years of treatment. CONCLUSIONS: The combination of eribulin and pembrolizumab demonstrated promising activity in LPS and angiosarcoma.

Intranasal irbesartan reverts cognitive decline and activates the PI3K/AKT pathway in an LPS-induced neuroinflammation mice model.

BACKGROUND: New strategies are urgently needed to manage and delay the development of Alzheimer's disease (AD). Neuroinflammation is a significant contributor to cognitive decline in neurodegenerative diseases, including AD. Angiotensin receptor blockers (ARBs) and angiotensin converting enzyme inhibitors (ACEIs) protect hypertensive patients against AD, but the cellular and molecular mechanisms underlying these effects remain unknown. In light of this, the protective effects of three ARBs and three ACEIs against neuroinflammation and cognitive decline were investigated through comprehensive pharmacologicalin vitro/in vivoscreening. METHODS: BV-2 microglia cells were exposed tolipopolysaccharide (LPS) and treated with ARBs and ACEIs to provide initial insights into the anti-inflammatory properties of the drugs. Subsequently, irbesartan was selected, and its efficacy was evaluated inC57/BL6 male miceintranasally administered with irbesartan and injected with LPS. Long-term memory and depressive-like behavior were evaluated; dendritic spines were measured as well as neuroinflammation, neurodegeneration and cognitive decline biomarkers. RESULTS: Irbesartan mitigated memory loss and depressive-like behavior in mice treated with LPS, probably because itincreased spine density, ameliorated synapsis dysfunction and activated the PI3K/AKT pathway. Irbesartan elevated the levels of hippocampalsuperoxide dismutase2 andglutathione peroxidaseandsuppressed LPS-induced astrogliosis. CONCLUSIONS: Overall, this study provides compelling evidence that multiple intranasal administrations of irbesartan can effectively prevent LPS-induced cognitive decline by activating pathways involved in neuroprotection and anti-inflammatory events. These findings underscore the potential of irbesartan as a preventive strategy against the development of AD and other neurodegenerative conditions associated with neuroinflammation.

Peppermint as a promising treatment agent in inflammatory conditions: A comprehensive systematic review of literature.

Inflammation, a type of the body's defense against injury or infection, causes many chronic disorders including diabetes, cardiovascular disease, and cancer. Therefore, discovering natural compounds with numerous biological activities for the management of inflammation is highly recommended. Out of natural compounds, peppermint and its main component, menthol, has been suggested to possess antiinflammatory potential. Four databases including Web of Sciences, PubMed, Scopus, and Embase were searched to identify articles about peppermint and its antiinflammatory effects up to March 2023. Out of 3805 records screened, 14 articles met the study criteria. The evidence reviewed here proposed peppermint as an antiinflammatory agent. Peppermint may suppress inflammation by activating the AMP-activated protein kinase/unc-51 like kinase 1/nuclear factor-E2 associated factor 2 autophagy pathway, downregulating extracellular signal-regulated kinase-nuclear factor kappa B and mitogen activated protein kinases pathways, attenuating oxidative stress, suppressing the production of pro-inflammatory mediators and nitric oxide, and inducing the production of antiinflammatory prostaglandins. Due to the promising antiinflammatory effects of peppermint and the lack of human studies in this regard, future randomized clinical trials examining the effects of peppermint on inflammation and its related maladies are warranted.

Effects of low-intensity extracorporeal shock wave therapy on lipopolysaccharide cystitis in a rat model of interstitial cystitis/bladder pain syndrome.

PURPOSE: To investigate the effect of low-intensity extracorporeal shock wave therapy (LiESWT) on lipopolysaccharide (LPS)-induced cystitis in an animal model of interstitial cystitis/bladder pain syndrome (IC/BPS). METHODS: Sprague-Dawley rats were divided into three groups: control, cystitis (LPS group, intravesical injection of LPS (1 mg) twice), and cystitis with LiESWT (LiESWT group). On the third and fourth days, LiESWT was administered (0.12 mJ/mm2, 300 shots each time) on the lower abdomen toward the bladder. On the seventh day, the rats underwent pain assessment and a metabolic cage study. Subsequently, a continuous cystometrogram (CMG) was performed under urethane anaesthesia. Immunohistochemical studies were also performed, including S-100 staining, an immunohistochemical marker of Schwann cells in the bladder. RESULTS: In the LPS group, the pain threshold in the lower abdomen was significantly lower than that in the control group. In the metabolic cage study, the mean voided volume in the LPS group significantly increased. The CMG also revealed a significant decrease in bladder contraction amplitude, compatible with detrusor underactivity in the LPS group. Immunohistochemical studies showed inflammatory changes in the submucosa, increased fibrosis, and decreased S-100 stain-positive areas in the muscle layer of the LPS group. In the LiESWT group, tactile allodynia and bladder function were ameliorated, and S-100 stain-positive areas were increased. CONCLUSION: By restoring nerve damage, LiESWT improved lower abdominal pain sensitivity and bladder function in an LPS-induced cystitis rat model. This study suggests that LiESWT may be a new therapeutic modality for IC/BPS.

Handelin alleviates cachexia- and aging-induced skeletal muscle atrophy by improving protein homeostasis and inhibiting inflammation.

BACKGROUND: Handelin is a bioactive compound from Chrysanthemum indicum L. that improves motor function and muscle integrity during aging in Caenorhabditis elegans. This study aimed to further evaluate the protective effects and molecular mechanisms of handelin in a mouse muscle atrophy model induced by cachexia and aging. METHODS: A tumour necrosis factor (TNF)-α-induced atrophy model was used to examine handelin activity in cultured C2C12 myotubes in vitro. Lipopolysaccharide (LPS)-treated 8-week-old model mice and 23-month-old (aged) mice were used to examine the therapeutic effects of handelin on cachexia- and aging-induced muscle atrophy, respectively, in vivo. Protein and mRNA expressions were analysed by Western blotting, ELISA and quantitative PCR, respectively. Skeletal muscle mass was measured by histological analysis. RESULTS: Handelin treatment resulted in an upregulation of protein levels of early (MyoD and myogenin) and late (myosin heavy chain, MyHC) differentiation markers in C2C12 myotubes (P < 0.05), and enhanced mitochondrial respiratory (P < 0.05). In TNF-α-induced myotube atrophy model, handelin maintained MyHC protein levels, increased insulin-like growth factor (Igf1) mRNA expression and phosphorylated protein kinase B protein levels (P < 0.05). Handelin also reduced atrogin-1 expression, inhibited nuclear factor-κB activation and reduced mRNA levels of interleukin (Il)6, Il1b and chemokine ligand 1 (Cxcl1) (P < 0.05). In LPS-treated mice, handelin increased body weight (P < 0.05), the weight (P < 0.01) and cross-sectional area (CSA) of the soleus muscle (P < 0.0001) and improved motor function (P < 0.05). In aged mice, handelin slightly increased the weight of the tibialis anterior muscle (P = 0.06) and CSA of the tibialis anterior and gastrocnemius muscles (P < 0.0001). In the tibialis anterior muscle of aged mice, handelin upregulated mRNA levels of Igf1 (P < 0.01), anti-inflammatory cytokine Il10 (P < 0.01), mitochondrial biogenesis genes (P < 0.05) and antioxidant-related enzymes (P < 0.05) and strengthened Sod and Cat enzyme activity (P < 0.05). Handelin also reduced lipid peroxidation and protein carbonylation, downregulated mRNA levels of Fbxo32, Mstn, Cxcl1, Il1b and Tnf (P < 0.05), and decreased IL-1ß levels in serum (P < 0.05). Knockdown of Hsp70 or using an Hsp70 inhibitor abolished the ameliorating effects of handelin on myotube atrophy. CONCLUSIONS: Handelin ameliorated cachexia- and aging-induced skeletal muscle atrophy in vitro and in vivo, by maintaining homeostasis of protein synthesis and degradation, possibly by inhibiting inflammation. Handelin is a potentially promising drug candidate for the treatment of muscle wasting.

Research on the Inhibitory Effect of Doxorubicin-loaded Liposomes Targeting GFAP for Glioma Cells.

BACKGROUND: Glioma is the most common and devastating brain tumor. In recent years, doxorubicin (DOX) is one of the drugs used in the treatment of gliomas, but it has side effects and poor clinical outcomes. Therefore, the delivery of drugs to the tumor site by targeted transport is a new approach to tumor treatment. OBJECTIVE: This study focuses on the anti-tumor effects of GFAP-modified drug-carrying liposomes loaded with DOX (GFAP-DOX-LPs) on gliomas. METHODS: GFAP-DOX-LPs were prepared by solvent evaporation method. After characterization analysis of GFAP-DOX-LPs, the encapsulation efficiency, the drug loading capacity and in vitro release performance were determined. Then, the MTT method was used to investigate the cytotoxicity and proliferative behavior of U251 and U87 cell lines. After that, flow cytometry was used to investigate the effect of the drug administration group on tumor cell apoptosis. Eventually, the anti-tumor activity was tested in vivo. RESULTS: The average particle size of GFAP-DOX-LPs was determined to be 116.3 ± 6.2 nm, and the average potential was displayed as 22.8 ± 7.2 mv. Besides, the morphology of the particle indicated a spherical shape. The encapsulation rate and drug loading were calculated and determined, which were 91.84 ± 0.41% and 9.27 ± 0.55%. In an acidic medium, the DOX release rate reached about 87%. GFAP-DOX-LPs could target glioma cells with low cytotoxicity and inhibit glioma cell proliferation with high efficiency, resulting in promoting apoptosis. The anti-tumor effect of GFAP-DOX-LPs was significantly enhanced. At the same time, the number of GFAPpositive cells in tumor tissues was significantly lower after treatment. Therefore, the overall survival time could be significantly prolonged. CONCLUSION: The prepared GFAP-DOX-LPs had good targeting and glioma cell inhibition ability. This demonstrated the promising application of the prepared liposomes in tumor targeting, especially in the field of targeted drug delivery for the treatment of brain tumor.

Anesthesia/surgery-induced learning and memory dysfunction by inhibiting mitophagy-mediated NLRP3 inflammasome inactivation in aged mice.

Postoperative cognitive dysfunction (POCD) is a common postoperative complication, not only affects the quality of life of the elderly and increases the mortality rate, but also brings a greater burden to the family and society. Previous studies demonstrated that Nod-like receptor protein 3 (NLRP3) inflammasome participates in various inflammatory and neurodegenerative diseases. However, possible mitophagy mechanism in anesthesia/surgery-elicited NLRP3 inflammasome activation remains to be elucidated. Hence, this study clarified whether mitophagy dysfunction is related to anesthesia/surgery-elicited NLRP3 inflammasome activation. POCD model was established in aged C57BL/6 J mice by tibial fracture fixation under isoflurane anesthesia. Morris Water Maze (MWM) was used to evaluate learning and memory abilities. We found that in vitro experiments, lipopolysaccharide (LPS) significantly facilitated NLRP3 inflammasome activation and mitophagy inhibition in BV2 cells. Rapamycin restored mitophagy and improved mitochondrial function, and inhibited NLRP3 inflammasome activation induced by LPS. In vivo experiments, anesthesia and surgery caused upregulation of hippocampal NLRP3, caspase recruitment domain (ASC) and interleukin-1ß (IL-1 ß), and downregulation of microtubule-associated protein light chain 3II (LC3II) and Beclin1 in aged mice. Olaparib inhibited anesthesia/surgery-induced NLRP3, ASC, and IL-1ß over-expression in the hippocampus, while upregulated the expression of LC3II and Beclin1. Furthermore, Olaparib improved cognitive impairment in older mice. These results revealed that mitophagy was involved in NLRP3 inflammasome-mediated anesthesia/surgery-induced cognitive deficits in aged mice. Overall, our results suggested that mitophagy was related in NLRP3 inflammasome-induced cognitive deficits after anesthesia and surgery in aged mice. Activating mitophagy may have clinical benefits in the prevention of cognitive impairment induced by anesthesia and surgery in elderly patients.

Oral angiotensin-converting enzyme inhibitor captopril protects the heart from Porphyromonas gingivalis LPS-induced cardiac dysfunction in mice.

Although angiotensin converting enzyme (ACE) inhibitors are considered useful for the treatment of human heart failure, some experimental failing-heart models have shown little beneficial effect of ACE inhibitors in animals with poor oral health, particularly periodontitis. In this study, we examined the effects of the ACE inhibitor captopril (Cap; 0.1 mg/mL in drinking water) on cardiac dysfunction in mice treated with Porphyromonas gingivalis lipopolysaccharide (PG-LPS) at a dose (0.8 mg/kg/day) equivalent to the circulating level in patients with periodontal disease. Mice were divided into four groups: 1) Control, 2) PG-LPS, 3) Cap, and 4) PG-LPS + Cap. After1 week, we evaluated cardiac function by echocardiography. The left ventricular ejection fraction was significantly decreased in PG-LPS-treated mice compared to the control (from 66 ± 1.8 to 59 ± 2.5%), while Cap ameliorated the dysfunction (63 ± 1.1%). The area of cardiac fibrosis was significantly increased (approximately 2.9-fold) and the number of apoptotic myocytes was significantly increased (approximately 5.6-fold) in the heart of PG-LPS-treated group versus the control, and these changes were suppressed by Cap. The impairment of cardiac function in PG-LPS-treated mice was associated with protein kinase C δ phosphorylation (Tyr-311), leading to upregulation of NADPH oxidase 4 and xanthine oxidase, and calmodulin kinase II phosphorylation (Thr-286) with increased phospholamban phosphorylation (Thr-17). These changes were also suppressed by Cap. Our results suggest that the renin-angiotensin system might play an important role in the development of cardiac diseases induced by PG-LPS.

[Protective effect of recombinant Schistosoma japonicum cystatin against acute kidney injury associated with acute liver failure in mice].

OBJECTIVE: To evaluate the protective effect of recombinant Schistosoma japonicum cystatin (rSj-Cys) against acute kidney injury induced by acute liver failure and unravel the underlying mechanism, so as to provide insights into the clinical therapy of acute kidney injury. METHODS: Twenty-four male C57BL/6J mice at ages of 6 to 8 weeks were randomly divided into the normal control group, rSj-Cys control group, lipopolysaccharide (LPS)/D-galactosamine (D-GaIN) model group and LPS/D-GaIN + rSj-Cys treatment group, of 6 mice each group. Mice in the LPS/D-GaIN group and LPS/D-GaIN + rSj-Cys group were intraperitoneally injected with LPS (10 µg/kg) and D-GaIN (700 mg/kg), and mice in the LPS/D-GaIN + rSj-Cys group were additionally administered with rSj-Cys (1.25 mg/kg) by intraperitoneal injection 30 min post-modeling, while mice in the rSj-Cys group were intraperitoneally injected with rSj-Cys (1.25 mg/kg), and mice in the normal control group were injected with the normal volume of PBS. All mice were sacrificed 6 h post-modeling, and mouse serum and kidney samples were collected. Serum creatinine (Cr) and urea nitrogen (BUN) levels were measured, and the pathological changes of mouse kidney specimens were examined using hematoxylin-eosin (HE) staining. Serum tumor necrosis factor (TNF)-α and interleukin (IL)-6 levels were detected using enzyme-linked immunosorbent assay (ELISA), and the expression of inflammatory factors and pyroptosis-related proteins was quantified in mouse kidney specimens using immunohistochemistry. In addition, the expression of pyroptosis-related proteins and nuclear factor-kappa B (NF-κB) signaling pathway-associated proteins was determined in mouse kidney specimens using Western blotting assay. RESULTS: HE staining showed no remarkable abnormality in the mouse kidney structure in the normal control group and the rSj-Cys control group, and renal tubular injury was found in LPS/D-GaIN group, while the renal tubular injury was alleviated in LPS/D-GaIN+rSj-Cys treatment group. There were significant differences in serum levels of Cr (F = 46.33, P < 0.001), BUN (F = 128.60, P < 0.001), TNF-α (F = 102.00, P < 0.001) and IL-6 (F = 202.10, P < 0.001) among the four groups, and lower serum Cr [(85.35 ± 32.05) µmol/L], BUN [(11.90 ± 2.76) mmol/L], TNF-α [(158.27 ± 15.83) pg/mL] and IL-6 levels [(56.72 ± 4.37) pg/mL] were detected in the in LPS/D-GaIN + rSj-Cys group than in the LPS/D-GaIN group (all P values < 0.01). Immunohistochemical staining detected significant differences in TNF-α (F = 24.16, P < 0.001) and IL-10 (F = 15.07, P < 0.01) expression among the four groups, and lower TNF-α [(106.50 ± 16.57)%] and higher IL-10 expression [(91.83 ± 5.23)%] was detected in the LPS/D-GaIN + rSj-Cys group than in the LPS/D-GaIN group (both P values < 0.01). Western blotting and immunohistochemistry detected significant differences in the protein expression of pyroptosis-related proteins NOD-like receptor thermal protein domain associated protein 3 (NLRP3) (F = 24.57 and 30.72, both P values < 0.001), IL-1ß (F = 19.24 and 22.59, both P values < 0.001) and IL-18 (F = 16.60 and 19.30, both P values < 0.001) in kidney samples among the four groups, and lower NLRP3, IL-1ß and IL-18 expression was quantified in the LPS/D-GaIN + rSj-Cys treatment group than in the LPS/D-GaIN group (P values < 0.05). In addition, there were significant differences in the protein expression of NF-κB signaling pathway-associated proteins p-NF-κB p-P65/NF-κB p65 (F = 71.88, P < 0.001), Toll-like receptor (TLR)-4 (F = 45.49, P < 0.001) and p-IκB/IκB (F = 60.87, P < 0.001) in mouse kidney samples among the four groups, and lower expression of three NF-κB signaling pathway-associated proteins was determined in the LPS/D-GaIN + rSj-Cys treatment group than in the LPS/D-GaIN group (all P values < 0.01). CONCLUSIONS: rSj-Cys may present a protective effect against acute kidney injury caused by acute liver failure through inhibiting inflammation and pyroptosis and downregulating the NF-κB signaling pathway.

Small Extracellular Vesicles Secreted by iPSC-Derived MSCs Ameliorate Pulmonary Inflammation and Lung Injury Induced by Sepsis through Delivery of miR-125b-5p.

Background: Sepsis-induced acute lung injury is a common critical illness in intensive care units with no effective treatment is currently available. Small extracellular vesicles, secreted by mesenchymal stem cells (MSCs), derived from human-induced pluripotent stem cells (iMSC-sEV), possess striking advantages when incorporated MSCs and iPSCs, which are considered extremely promising cell-free therapeutic agents. However, no studies have yet been conducted to systemically examine the effects and underlying mechanisms of iMSC-sEV application on attenuated lung injury under sepsis conditions. Method: iMSC-sEV were intraperitoneally administered in a rat septic lung injury model induced by cecal ligation and puncture (CLP). The efficacy of iMSC-sEV was assessed by histology, immunohistochemistry, and pro-inflammatory cytokines of bronchoalveolar lavage fluid. We also evaluated the in vitro effects of iMSC-sEV on the activation of the inflammatory response in alveolar macrophages (AMs). Small RNA sequencing was utilized to detect changes in the miRNA expression profile in lipopolysaccharide (LPS)-treated AMs after iMSC-sEV administration. The effects of miR-125b-5p on the function of AMs were studied. Results: iMSC-sEV were able to attenuate pulmonary inflammation and lung injury following CLP-induced lung injury. iMSC-sEV were internalized by AMs and alleviated the release of inflammatory factors by inactivating the NF-κB signaling pathway. Moreover, miR-125b-5p showed a fold-change in LPS-treated AMs after iMSC-sEV administration and was enriched in iMSC-sEV. Mechanistically, iMSC-sEV transmitted miR-125b-5p into LPS-treated AMs to target TRAF6. Conclusion: Our findings demonstrated that iMSC-sEV treatment protects against septic lung injury and exerts anti-inflammatory effects on AMs at least partially through miR-125b-5p, suggesting that iMSC-sEV may provide a novel cell-free strategy for the treatment of septic lung injury.

The role of intestinal alkaline phosphatase in the development of necrotizing enterocolitis.

Necrotizing enterocolitis (NEC) is a devastating neonatal disease that affects neonates worldwide and often leads to high morbidity and mortality rates. Despite extensive research, the cause of NEC remains unclear, and current treatment options are limited. An important novel finding is the potential role of intestinal Alkaline Phosphatase (IAP) in both pathogenesis and treatment of NEC. IAP can play a vital role in detoxifying liposaccharides (LPS), a key mediator of many pathological processes, thereby reducing the inflammatory response associated with NEC. Furthermore, IAP can help prevent dysbiosis, improve intestinal perfusion, and promote autophagy. In this comprehensive review, we present evidence of the possible connection between IAP and the LPS/Toll-like receptor 4 (TLR4) pathway, impaired gut immunity, and dysbiosis in the preterm gut. Based on these findings, the administration of exogenous IAP might provide promising preventive and therapeutic avenues for the management of NEC.

Gut barrier dysfunction and endotoxemia in heart failure: A dangerous connubium?

Heart failure (HF) is a leading cause of death worldwide despite recent advances in pharmacological treatments. Gut microbiota dysbiosis and gut barrier dysfunction with consequent bacterial translocation and increased blood endotoxemia has gained much attention as one of the key pathogenetic mechanisms contributing to increased mortality of patients at risk or with cardiovascular disease. Indeed, increased blood levels of lipopolysaccharide (LPS), a glycolipid of outer membrane of gut gram-negative bacteria, have been detected in patients with diabetes, obesity and nonalcoholic fatty liver disease or in patients with established coronary disease such as myocardial infarction or atrial fibrillation, suggesting endotoxemia as aggravating factor via systemic inflammation and eventually vascular damage. Upon interaction with its receptor Toll-like receptor 4 (TLR4) LPS may, in fact, act at different cellular levels so eliciting formation of proinflammatory cytokines or exerting a procoagulant activity. Increasing body of evidence pointed to endotoxemia as factor potentially deteriorating the clinical course of patients with HF, that, in fact, is associated with gut dysbiosis-derived changes of gut barrier functionality and eventually bacteria or bacterial product translocation into systemic circulation. The aim of this review is to summarize current experimental and clinical evidence on the mechanisms linking gut dysbiosis-related endotoxemia with HF, its potential negative impact with HF progression, and the therapeutic strategies that can counteract endotoxemia.