Intracellular Signaling Pathways Research Articles

The Role of Kidney Intercalated Cells in the Immune Response Against UTIs

Introduction: Urinary tract infections (UTIs) are among the most prevalent infections. About 3 in 5 women will experience UTIs in their lifetime, resulting from a bacterial infection caused by uropathogenic E. coli (UPEC). Ascending UPEC first adhere to the collecting ducts, where intercalated cells (ICs) are essential for raising an innate immune response activated by UPEC-outer membrane lipopolysaccharide (LPS). Interestingly, individuals with distal renal tubular acidosis (dRTA) who have a mutation on the gene encoding the kidney anion exchanger 1 (kAE1) have a decreased abundance of IC and are at a higher risk for UTIs. The cellular mechanisms contributing to this higher vulnerability to UTIs remain unclear. Objective and Hypothesis: To enhance our understanding of the immune response mechanisms of collecting duct cells in dRTA patients during UTIs, we hypothesize that the loss of intercalated cells (ICs) in dRTA results in an inadequate immune response against UPEC. Methods: Mouse inner medullary collecting duct cells (mIMCD3) and a mouse strain with a knock-in mutation for kAE1 dRTA (specifically, R607H, equivalent to human R589H) were utilized in the study. We identified and quantified innate immunity markers in mIMCD3 cells exposed to vehicle, UPEC or LPS using qRT-PCR and immunoblot. Primary IC were prepared by magnetic-activated cell sorting (MACS). Various incubation conditions, including exposure to vehicle, LPS, control K12 bacteria, and UPEC for different time intervals were assessed to determine the best experimental conditions. Transurethral UPEC inoculation procedures were also performed to validate results obtained in the in vitro approaches. Results: Experiments on mIMCD3 cells showed a significant increase in the innate immunity marker adrenomedullin (ADM) following exposure to UPEC or LPS, compared to the control group. After 17 h, exposing the primary MACS-enriched ICs to UPEC significantly increased the adrenomedullin protein abundance. UPEC transurethral infections in dRTA mice resulted in a higher bacterial load and increased IL-1β, IL-6, and TNF-α mRNA and protein levels compared to WT mice at 24 post-UTI. In the transgenic mice, ADM gene expression was not up-regulated to the same extent as in WT mice, but LCN2/NGAL gene expression was significantly higher. Conclusions: Our study suggests that the loss of IC in a dRTA mouse model results in higher inflammation and reduced immunity against UPEC exposure. Our research will unravel new therapeutic leads on the intracellular signaling pathway(s) and cellular changes that result in IC-mediated renal protection. The findings may contribute to the development of novel therapeutics for UTIs. Natural Sciences and Engineering Research Council of Canada (NSERC), the Canadian Institutes of Health Research (CIHR), University of Alberta Graduate Recruitment Scholarship (2023). This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Non-coding RNAs in Precursor Lesions of Colorectal Cancer: Their Role in Cancer Initiation and Formation.

Colorectal cancer (CRC) is one of the world's most common types of malignancy. The proliferation of precancerous lesions causes this type of cancer. Two distinct pathways for CRC carcinogenesis have been identified: the conventional adenoma-carcinoma pathway and the serrated neoplasia pathway. Recently, evidence has demonstrated the regulatory roles of noncoding RNAs (ncRNAs) in the initiation and progression of precancerous lesions, especially in the adenoma-carcinoma pathway and serrated neoplasia pathway. By expanding the science of molecular genetics and bioinformatics, several studies have identified dysregulated ncRNAs that function as oncogenes or tumor suppressors in cancer initiation and formation by diverse mechanisms via intracellular signaling pathways known to act on tumor cells. However, many of their roles are still unclear. This review summarizes the functions and mechanisms of ncRNAs (such as long non-coding RNAs, microRNAs, long intergenic non-coding RNAs, small interfering RNAs, and circRNAs) in the initiation and formation of precancerous lesions.

Influence of metallic particles and TNF on the transcriptional regulation of NLRP3 inflammasome-associated genes in human osteoblasts.

The release of mature interleukin (IL-) 1β from osteoblasts in response to danger signals is tightly regulated by the nucleotide-binding oligomerization domain leucine-rich repeat and pyrin-containing protein 3 (NLRP3) inflammasome. These danger signals include wear products resulting from aseptic loosening of joint arthroplasty. However, inflammasome activation requires two different signals: a nuclear factor-kappa B (NF-κB)-activating priming signal and an actual inflammasome-activating signal. Since human osteoblasts react to wear particles via Toll-like receptors (TLR), particles may represent an inflammasome activator that can induce both signals. Temporal gene expression profiles of TLRs and associated intracellular signaling pathways were determined to investigate the period when human osteoblasts take up metallic wear particles after initial contact and initiate a molecular response. For this purpose, human osteoblasts were treated with metallic particles derived from cobalt-chromium alloy (CoCr), lipopolysaccharides (LPS), and tumor necrosis factor-alpha (TNF) alone or in combination for incubation times ranging from one hour to three days. Shortly after adding the particles, their uptake was observed by the change in cell morphology and spectral data. Exposure of osteoblasts to particles alone increased NLRP3 inflammasome-associated genes. The response was not significantly enhanced when cells were treated with CoCr + LPS or CoCr + TNF, whereas inflammation markers were induced. Despite an increase in genes related to the NLRP3 inflammasome, the release of IL-1β was unaffected after contact with CoCr particles. Although CoCr particles affect the expression of NLRP3 inflammasome-associated genes, a single stimulus was not sufficient to prime and activate the inflammasome. TNF was able to prime the NLRP3 inflammasome of human osteoblasts.

Porcine TLR8 signaling and its anti-infection function are disturbed by immune checkpoint receptor TIM-3 via inhibition of P13K-AKT pathway

Toll-like receptor 8 (TLR8), an important innate immune receptor recognizing single stranded RNA and the antiviral imidazoquinoline compounds, can activate intracellular signaling pathway and produce an inflammatory response to kill and eliminate pathogens. However, the molecular regulation mechanisms of TLR8 signaling and its anti-infection activity are not fully elucidated. Our previous transcriptome analysis of porcine TLR8 (pTLR8) signaling suggested the immune checkpoint receptor TIM-3 as the potential regulator for pTLR8. Here we investigated TIM-3 in the regulation of pTLR8 signaling and its anti-infection activity. Our results showed that porcine TIM-3 is upregulated by pTLR8 signaling and TIM-3 inhibits pTLR8 signaling activity in a negative feedback way. Accordingly, TIM-3 disturbs pTLR8 mediated anti-bacterial and anti-viral activity. Mechanistically, TIM-3 suppresses PI3K-AKT pathway by inhibiting the TLR8-PI3K p85 interaction and subsequent AKT phosphorylation which is essential for TLR8 signaling and anti-infection activity. Therefore, our study reveals new insights into innate immune TLR8 signaling and its anti-infection function.

Vascular endothelial growth factor receptor 2 contributes to shear stress-induced endothelial nitric oxide synthase expression in human lung microvascular endothelial cells

Introduction: Physiological shear stress (pSS) helps to maintain endothelial homeostasis via numerous intracellular signaling pathways. We previously showed that pSS upregulates mRNA and protein levels of endothelial nitric oxide synthase (eNOS) in primary human lung microvascular endothelial cells (hLMVECs), and that phosphorylation of Akt (p-Akt), a phosphatidylinositol 3-kinase (PI3K) target, is increased. Upstream regulatory mechanisms for these responses have not been fully studied. Since vascular endothelial growth factor receptor 2 (VEGFR2) is an integral component of the mechanosensory complex that transduces shear stress-mediated signaling pathways in endothelial cells, we hypothesized that VEGFR2 is required for shear stress-induced eNOS expression and PI3K activation in hLMVECs. Methods: We cultured early passage (p≤7) primary hLMVECs from 4 unique donor lots in standard 6-well culture plates using a fixed volume (depth) and viscosity of tissue culture media. We exposed cells to shear stress for 24 hours using an orbital shaker and calculated the magnitude of shear stress exerted on hLMVECs using the extended solution to Stokes’ second problem. We defined the magnitude of pSS as 12 dyn/cm2. In some experiments, we exposed cells to pSS in the presence of chemical inhibitors of VEGFR2 (SU5416; 5 or 10 μM) or PI3K (LY294002; 10 μM). We collected whole cell lysates and performed immunoblotting to probe for phosphorylated eNOS (p-eNOS; Serine 1177), total eNOS, p-Akt (Serine 473), total Akt and housekeeping protein β-tubulin. Potential interactions between chemical inhibitors and pSS on protein expression and phosphorylation were analyzed by two-way ANOVA. Results: After 24 hours of pSS exposure, total eNOS levels were significantly (p<0.05) increased in hLMVECs, although the p-eNOS/eNOS ratio was not changed (p=0.85). Conversely, 24 hours of pSS exposure increased the p-Akt/Akt ratio (p<0.05) but did not increase total Akt levels (p=0.88). Inhibition of VEGFR2 activity with SU5416 attenuated pSS-induced changes in total eNOS levels (p<0.05 for interaction) but did not attenuate pSS-induced increases in the p-Akt/Akt ratio (p=0.91 for interaction). Inhibition of PI3K activity using LY294002 did not modify the effects of pSS on total eNOS levels (p=0.25 for interaction) or p-Akt/Akt ratio (p=0.47 for interaction). Conclusions: We conclude that VEGFR2 activity is necessary for pSS-mediated increases in eNOS protein levels, but not PI3K activity, in hLMVECs. Furthermore, neither VEGFR2 nor PI3K activity appear to be required for pSS-mediated eNOS phosphorylation in these cells. Collectively, these data suggest that shear stress regulates hLMVEC eNOS signaling via multiple signaling pathways. This complex regulation may have important implications for local nitric oxide bioavailability under pathological conditions that modify shear stress (e.g., pulmonary embolism, pulmonary hypertension). This work is supported by NIH grants HL126514 and HL159906. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

High-Resolution and Dynamic Visualization of Intracellular Redox Potential Using a Metal-Organic Framework-Functionalized Nanopotentiometer.

Redox potential plays a key role in regulating intracellular signaling pathways, with its quantitative analysis in individual cells benefiting our understanding of the underlying mechanism in the pathophysiological events. Here, a metal organic framework (MOF)-functionalized SERS nanopotentiometer has been developed for the dynamic monitoring of intracellular redox potential. The approach is based on the encapsulation of zirconium-based MOF (Uio-66-F4) on a surface of gold-silver nanorods (Au-Ag NRs) that is modified with the newly synthesized redox-sensitive probe ortho-mercaptohydroquinone (HQ). Thanks to size exclusion of MOF as the chemical protector, the nanopotentiometer can be adapted to long-term use and possess high anti-interference ability toward nonredox species. Combining the superior fingerprint identification of SERS with the electrochemical activity of the quinone/hydroquinone, the nanopotentiometer shows a reversible redox responsivity and can quantify redox potential with a relatively wide range of -250-100 mV. Furthermore, the nanopotentiometer allows for dynamic visualization of intracellular redox potential changes induced by drugs' stimulation in a high-resolution manner. The developed approach would be promising for offering new insights into the correlation between redox potential and tumor proliferation-involved processes such as oxidative stress and hypoxia.

Exploring the Role of Neuroplasticity in Stroke Rehabilitation: Mechanisms, Interventions and Clinical Implications

Neuroplasticity, the brain's remarkable ability to reorganize and adapt throughout life, has emerged as a central focus in neuroscience research. This abstract delves into the mechanisms underlying neuroplasticity and its profound implications for learning, memory, rehabilitation, and neurological disorders. At the cellular level, neuroplasticity involves synaptic plasticity, where the strength and efficacy of connections between neurons can be modified through activity-dependent processes such as long-term potentiation (LTP) and long-term depression (LTD). Molecular mechanisms, including changes in neurotransmitter release, receptor expression, and intracellular signaling pathways, mediate these synaptic changes. Beyond synaptic plasticity, structural plasticity encompasses alterations in neuronal morphology, including dendritic branching, spine density, and axonal sprouting. These structural changes facilitate the formation of new neural circuits and underlie learning and memory processes. Neuroplasticity is not limited to developmental stages but persists throughout life, with experience and environmental factors continuously shaping the brain's structure and function. Experience-dependent plasticity is evident in various contexts, from skill acquisition to recovery from brain injuries. Understanding neuroplasticity has profound implications for education, as it highlights the importance of enriched environments and active learning strategies in promoting cognitive development. Additionally, neuroplasticity forms the basis for rehabilitative interventions following brain damage, with therapies focusing on promoting adaptive neural rewiring and functional recovery. Moreover, dysregulation of neuroplasticity is implicated in numerous neurological and psychiatric disorders, including Alzheimer's disease, stroke, and depression. Elucidating the mechanisms underlying aberrant plasticity holds promise for developing targeted interventions to treat these conditions. In conclusion, neuroplasticity represents a fundamental property of the nervous system, allowing for adaptation and optimization in response to environmental demands. Continued research into the mechanisms governing neuroplasticity promises to unveil new therapeutic avenues and deepen our understanding of brain function and dysfunction.

Systemic Sclerosis-Associated Pulmonary Arterial Hypertension: From Bedside to Bench and Back Again.

Systemic sclerosis (SSc) is a heterogeneous disease characterized by autoimmunity, vasculopathy, and fibrosis which affects the skin and internal organs. One key aspect of SSc vasculopathy is pulmonary arterial hypertension (SSc-PAH) which represents a leading cause of morbidity and mortality in patients with SSc. The pathogenesis of pulmonary hypertension is complex, with multiple vascular cell types, inflammation, and intracellular signaling pathways contributing to vascular pathology and remodeling. In this review, we focus on shared molecular features of pulmonary hypertension and those which make SSc-PAH a unique entity. We highlight advances in the understanding of the clinical and translational science pertinent to this disease. We first review clinical presentations and phenotypes, pathology, and novel biomarkers, and then highlight relevant animal models, key cellular and molecular pathways in pathogenesis, and explore emerging treatment strategies in SSc-PAH.

The IC87201 (a PSD95/nNOS Inhibitor) Attenuates Post- Stroke Injuries.

N-methyl-D-aspartate receptor-dependent excitotoxicity is one of the most important mechanisms underlying stroke injury and the resulting neuronal death. In the present study, in order to reduce post-stroke brain injury and improve behavioral performance, a new molecule named IC87201, which acts as an inhibitor of PSD95/nNOS interaction in the intracellular signaling pathway of NMDA receptors, was administered. Using the middle cerebral artery occlusion (MCAO) technique, 24 adult male rats were subjected to one hour of cerebral ischemia. Animals were randomly divided into sham, MCAO, MCAO + DXM, and MCAO + IC87201 groups, and in the last two groups, intraperitoneal injection of dextromethorphan hydrobromide monohydrate (DXM), as an NMDA antagonist, and IC87201 was performed after ischemia. Neurobehavioral scores were evaluated for seven days, and on the last two days, the rats' memory performance was appraised using the passive avoidance test. On seventh day, the brain tissue was properly prepared for stereological analysis. Stereological studies of the hippocampus CA1 and CA3 regions revealed that changes in the total and infarcted volumes, total number of neurons, non-neurons, and dead neurons are the consequences of cerebral ischemia. Also, following cerebral ischemia, neurobehavioral and memory function impairments which were assessed by modified neurological severity scores (mNSS) and passive avoidance test, were observed. The aforementioned impairments were recovered after administration of IC87201 significantly and more potently than DXM. Based on our findings, IC87201 successfully attenuated post-ischemia damages. Therefore, this molecule can be considered as a new therapeutic approach in future research.

Predicting FFAR4 agonists using structure-based machine learning approach based on molecular fingerprints

Free Fatty Acid Receptor 4 (FFAR4), a G-protein-coupled receptor, is responsible for triggering intracellular signaling pathways that regulate various physiological processes. FFAR4 agonists are associated with enhancing insulin release and mitigating the atherogenic, obesogenic, pro-carcinogenic, and pro-diabetogenic effects, normally associated with the free fatty acids bound to FFAR4. In this research, molecular structure-based machine-learning techniques were employed to evaluate compounds as potential agonists for FFAR4. Molecular structures were encoded into bit arrays, serving as molecular fingerprints, which were subsequently analyzed using the Bayesian network algorithm to identify patterns for screening the data. The shortlisted hits obtained via machine learning protocols were further validated by Molecular Docking and via ADME and Toxicity predictions. The shortlisted compounds were then subjected to MD Simulations of the membrane-bound FFAR4-ligand complexes for 100 ns each. Molecular analyses, encompassing binding interactions, RMSD, RMSF, RoG, PCA, and FEL, were conducted to scrutinize the protein–ligand complexes at the inter-atomic level. The analyses revealed significant interactions of the shortlisted compounds with the crucial residues of FFAR4 previously documented. FFAR4 as part of the complexes demonstrated consistent RMSDs, ranging from 3.57 to 3.64, with minimal residue fluctuations 5.27 to 6.03 nm, suggesting stable complexes. The gyration values fluctuated between 22.8 to 23.5 nm, indicating structural compactness and orderliness across the studied systems. Additionally, distinct conformational motions were observed in each complex, with energy contours shifting to broader energy basins throughout the simulation, suggesting thermodynamically stable protein–ligand complexes. The two compounds CHEMBL2012662 and CHEMBL64616 are presented as potential FFAR4 agonists, based on these insights and in-depth analyses. Collectively, these findings advance our comprehension of FFAR4’s functions and mechanisms, highlighting these compounds as potential FFAR4 agonists worthy of further exploration as innovative treatments for metabolic and immune-related conditions.

Methylglyoxal and Advanced Glycation End Products (AGEs): Targets for the Prevention and Treatment of Diabetes-Associated Bladder Dysfunction?

Methylglyoxal (MGO) is a highly reactive α-dicarbonyl compound formed endogenously from 3-carbon glycolytic intermediates. Methylglyoxal accumulated in plasma and urine of hyperglycemic and diabetic individuals acts as a potent peptide glycation molecule, giving rise to advanced glycation end products (AGEs) like arginine-derived hydroimidazolone (MG-H1) and carboxyethyl-lysine (CEL). Methylglyoxal-derived AGEs exert their effects mostly via activation of RAGE, a cell surface receptor that initiates multiple intracellular signaling pathways, favoring a pro-oxidant environment through NADPH oxidase activation and generation of high levels of reactive oxygen species (ROS). Diabetic bladder dysfunction is a bothersome urological complication in patients with poorly controlled diabetes mellitus and may comprise overactive bladder, urge incontinence, poor emptying, dribbling, incomplete emptying of the bladder, and urinary retention. Preclinical models of type 1 and type 2 diabetes have further confirmed the relationship between diabetes and voiding dysfunction. Interestingly, healthy mice supplemented with MGO for prolonged periods exhibit in vivo and in vitro bladder dysfunction, which is accompanied by increased AGE formation and RAGE expression, as well as by ROS overproduction in bladder tissues. Drugs reported to scavenge MGO and to inactivate AGEs like metformin, polyphenols, and alagebrium (ALT-711) have shown favorable outcomes on bladder dysfunction in diabetic obese leptin-deficient and MGO-exposed mice. Therefore, MGO, AGEs, and RAGE levels may be critically involved in the pathogenesis of bladder dysfunction in diabetic individuals. However, there are no clinical trials designed to test drugs that selectively inhibit the MGO-AGEs-RAGE signaling, aiming to reduce the manifestations of diabetes-associated bladder dysfunction. This review summarizes the current literature on the role of MGO-AGEs-RAGE-ROS axis in diabetes-associated bladder dysfunction. Drugs that directly inactivate MGO and ameliorate bladder dysfunction are also reviewed here.

IQGAP3 Is an Important Mediator of Skin Inflammatory Diseases.

IQGAP3 (IQ Motif Containing GTPase Activating Protein 3) is member of the IQGAP family of scaffold proteins, which are essential for assembling multiprotein complexes that coordinate various intracellular signaling pathways. Previous research has shown that IQGAP3 is overexpressed in psoriatic skin lesions. Given its involvement in processes like cell proliferation and chemokine signaling, we sought to explore its molecular role in driving the psoriatic phenotype of keratinocytes. By conducting transcriptome profiling of HaCaT keratinocytes, we identified numerous psoriasis-associated pathways that were affected when IQGAP3 was knocked down. These included alterations in NFkB signaling, EGFR signaling, activation of p38/MAPK and ERK1/ERK2, lipid metabolism, cytokine production, and the response to inflammatory cytokine stimulation. Real-time analysis further revealed changes in cell growth dynamics, including proliferation and wound healing. The balance between cell proliferation and apoptosis was altered, as were skin barrier functions and the production of IL-6 and IFNγ. Despite these significant findings, the diversity of the alterations observed in the knockdown cells led us to conclude that IQGAP3 may not be the best target for the therapeutic inhibition to normalize the phenotype of keratinocytes in psoriasis.

Unlocking the Power of Light on the Skin: A Comprehensive Review on Photobiomodulation.

Photobiomodulation (PBM) is a procedure that uses light to modulate cellular functions and biological processes. Over the past decades, PBM has gained considerable attention for its potential in various medical applications due to its non-invasive nature and minimal side effects. We conducted a narrative review including articles about photobiomodulation, LED light therapy or low-level laser therapy and their applications on dermatology published over the last 6 years, encompassing research studies, clinical trials, and technological developments. This review highlights the mechanisms of action underlying PBM, including the interaction with cellular chromophores and the activation of intracellular signaling pathways. The evidence from clinical trials and experimental studies to evaluate the efficacy of PBM in clinical practice is summarized with a special emphasis on dermatology. Furthermore, advancements in PBM technology, such as novel light sources and treatment protocols, are discussed in the context of optimizing therapeutic outcomes and improving patient care. This narrative review underscores the promising role of PBM as a non-invasive therapeutic approach with broad clinical applicability. Despite the need for further research to develop standard protocols, PBM holds great potential for addressing a wide range of medical conditions and enhancing patient outcomes in modern healthcare practice.

MiR-141/200c contributes to ethanol-mediated hepatic glycogen metabolism

ObjectiveHepatic glucose metabolism is profoundly perturbed by excessive alcohol intake. miR-141/200c expression is significantly induced by chronic ethanol feeding. This study aimed at identifying the role of miR-141/200c in glucose homeostasis during chronic ethanol exposure. MethodsWT and miR-141/200c KO mice were fed a control or an ethanol diet for 30 days, followed by a single binge of maltose dextrin or ethanol, respectively. Untargeted metabolomics analysis of hepatic primary metabolites was performed along with analyses for liver histology, gene expression, intracellular signaling pathways, and physiological relevance. Primary hepatocytes were used for mechanistic studies. ResultsmiR-141/200c deficiency rewires hepatic glucose metabolism during chronic ethanol feeding, increasing the abundance of glucose intermediates including G6P, an allosteric activator for GS. miR-141/200c deficiency replenished glycogen depletion during chronic ethanol feeding accompanied by reduced GS phosphorylation in parallel with increased expression of PP1 glycogen targeting subunits. Moreover, miR-141/200c deficiency prevented ethanol-mediated increases in AMPK and CaMKK2 activity. Ethanol treatment reduced glycogen content in WT-hepatocytes, which was reversed by dorsomorphin, a selective AMPK inhibitor, while KO-hepatocytes displayed higher glycogen content than WT-hepatocytes in response to ethanol treatment. Furthermore, treatment of hepatocytes with A23187, a calcium ionophore activating CaMKK2, lowered glycogen content in WT-hepatocytes. Notably, the suppressive effect of A23187 on glycogen deposition was reversed by dorsomorphin, demonstrating that the glycogen depletion by A23187 is mediated by AMPK. KO-hepatocytes exhibited higher glycogen content than WT-hepatocytes in response to A23187. Finally, miR-141/200c deficiency led to improved glucose tolerance and insulin sensitivity during chronic ethanol feeding. ConclusionsmiR-141/200c deficiency replenishes ethanol-mediated hepatic glycogen depletion through the regulation of GS activity and calcium signaling coupled with the AMPK pathway, improving glucose homeostasis and insulin sensitivity. These results underscore miR-141/200c as a potential therapeutic target for the management of alcohol intoxication.

Bupleurum in Treatment of Depression Disorder: A Comprehensive Review.

The incidence of depression has been steadily rising in recent years, making it one of the most prevalent mental illnesses. As the pursuit of novel antidepressant drugs captivates the pharmaceutical field, the therapeutic efficacy of Traditional Chinese Medicine (TCM) has been widely explored. Chaihu (Bupleurum) has been traditionally used for liver conditions such as hepatitis, liver inflammation, liver fibrosis, and liver cancer. It is believed to have hepatoprotective effects, promoting liver cell regeneration and protecting against liver damage. In addition, Bupleurum has also been used as a Jie Yu (depression-relieving) medicine in China, Japan, Republic of Korea, and other Asian countries for centuries. This review article aims to summarize the research conducted on the antidepressant properties and mechanisms of Bupleurum, as well as discuss the potential of TCM formulas containing Bupleurum. This review highlights various antidepressant ingredients isolated from Bupleurum, including saikosaponin A, saikosaponin D, rutin, puerarin, and quercetin, each with distinct mechanisms of action. Additionally, Chinese herb prescriptions and extracts containing Bupleurum, such as Chaihu Shugansan, Xiaoyaosan, and Sinisan, are also included due to their demonstrated antidepressant effects. This review reveals that these Bupleurum compounds exhibit antidepressant effects through the regulation of neurotransmitter mechanisms (such as 5-HT and DA), the NMDA (N-methyl-D-aspartate) system, brain-derived neurotrophic factor (BDNF), and other intracellular signaling pathways. Collectively, this comprehensive review provides insights into the multiple applications of Bupleurum in the treatment of depression and highlights its potential as an alternative or complementary approach to traditional therapies. However, it is essential to consider the potential adverse effects and clinical restrictions of Bupleurum despite its promising potential. Further research is needed to elucidate its specific mechanisms of action and evaluate its effectiveness in human subjects.

The E3 Ubiquitin Protein Ligase LINCR Amplifies the TLR-Mediated Signals through Direct Degradation of MKP1.

Toll-like receptors (TLRs) induce innate immune responses through activation of intracellular signaling pathways, such as MAP kinase and NF-κB signaling pathways, and play an important role in host defense against bacterial or viral infections. Meanwhile, excessive activation of TLR signaling leads to a variety of inflammatory disorders, including autoimmune diseases. TLR signaling is therefore strictly controlled to balance optimal immune response and inflammation. However, its balancing mechanisms are not fully understood. In this study, we identified the E3 ubiquitin ligase LINCR/ NEURL3 as a critical regulator of TLR signaling. In LINCR-deficient cells, the sustained activation of JNK and p38 MAPKs induced by the agonists for TLR3, TLR4, and TLR5, was clearly attenuated. Consistent with these observations, TLR-induced production of a series of inflammatory cytokines was significantly attenuated, suggesting that LINCR positively regulates innate immune responses by promoting the activation of JNK and p38. Interestingly, our further mechanistic study identified MAPK phosphatase-1 (MKP1), a negative regulator of MAP kinases, as a ubiquitination target of LINCR. Thus, our results demonstrate that TLRs fine-tune the activation of MAP kinase pathways by balancing LINCR (the positive regulator) and MKP1 (the negative regulator), which may contribute to the induction of optimal immune responses.

Low-Molecular Weight Compounds that Extend the Chronological Lifespan of Yeasts, Saccharomyces cerevisiae, and Schizosaccharomyces pombe.

Yeast is an excellent model organism for research for regulating aging and lifespan, and the studies have made many contributions to date, including identifying various factors and signaling pathways related to aging and lifespan. More than 20 years have passed since molecular biological perspectives are adopted in this research field, and intracellular factors and signal pathways that control aging and lifespan have evolutionarily conserved from yeast to mammals. Furthermore, these findings have been applied to control the aging and lifespan of various model organisms by adjustment of the nutritional environment, genetic manipulation, and drug treatment using low-molecular weight compounds. Among these, drug treatment is easier than the other methods, and research into drugs that regulate aging and lifespan is consequently expected to become more active. Chronological lifespan, a definition of yeast lifespan, refers to the survival period of a cell population under nondividing conditions. Herein, low-molecular weight compounds are summarized that extend the chronological lifespan of Saccharomyces cerevisiae and Schizosaccharomyces pombe, along with their intracellular functions. The low-molecular weight compounds are also discussed that extend the lifespan of other model organisms. Compounds that have so far only been studied in yeast may soon extend lifespan in other organisms.

Effect of nicotinamide mononucleotide on osteogenesis in MC3T3-E1 cells against inflammation-induced by lipopolysaccharide.

Nicotinamide mononucleotide (NMN) is extensively utilized as an anti-aging agent and possesses anti-inflammatory properties. Lipopolysaccharide (LPS) activates Toll-like receptor 4, a process modulated by intracellular signaling pathways such as the Wnt/β-catenin pathway. This study investigated the impact of NMN on osteogenesis in the presence of LPS. To elucidate the role of NMN in osteogenesis in the context of Gram-negative bacterial infection after LPS treatment, we cultured a mouse pre-osteoblast cell line (MC3T3-E1) and subsequently incubated it with NMN and/or LPS. We then evaluated osteogenic activity by measuring alkaline phosphatase activity, assessing gene expression and protein levels, and performing Alizarin Red S staining and immunocytochemistry. MC3T3-E1 cells underwent successful differentiation into osteoblasts following treatment with osteogenic induction medium. LPS diminished features related to osteogenic differentiation, which were subsequently partially reversed by treatment with NMN. The restorative effects of NMN on LPS-exposed MC3T3-E1 cells were further substantiated by elucidating the role of Wnt/β-catenin signaling, as confirmed through immunocytochemistry. This study showed that infection with Gram-negative bacteria disrupted the osteogenic differentiation of MC3T3-E1 cells. This adverse effect was partially reversed by administering a high-dose of NMN. Drawing on these results, we propose that NMN could serve as a viable therapeutic strategy to preserve bone homeostasis in elderly and immunocompromised patients.

Genistein Prevents Apoptosis and Oxidative Stress Induced by Methylglyoxal in Endothelial Cells.

Glycolytic overload promotes accumulation of the highly reactive dicarbonyl compounds, resulting in harmful conditions called dicarbonyl stress. Methylglyoxal (MG) is a highly reactive dicarbonyl species and its accumulation plays a crucial pathophysiological role in diabetes and its vascular complications. MG cytotoxicity is mediated by reactive oxygen species (ROS) generation, a key event underlying the intracellular signaling pathways leading to inflammation and apoptosis. The identification of compounds able to inhibit ROS signaling pathways and counteract the MG-induced toxicity is a crucial step for developing new therapeutic strategies in the treatment of diabetic vascular complications. In this study, the effect of genistein, a natural soybean isoflavone, has been evaluated on MG-induced cytotoxicity in human endothelial cells. Our results show that genistein is able to counteract the MG-induced apoptosis by restraining ROS production, thus inhibiting the MAPK signaling pathways and caspase-3 activation. These findings identify a beneficial role for genistein, providing new insights for its potential clinical applications in preserving endothelial function in diabetic vascular complications.

Cytokine inhibitory activity of eurycomanone in RAW 264.7 cells stimulated with viral-mimicking poly (I:C)

Poly (I:C) (Polyinosinic:polycytidylic acid) is a synthetic analog of double-stranded RNA binding toll-like receptor 3 (TLR3) and mimicking a virus infection in macrophage. Here, for the first time, we showed the inhibitory effects of eurycomanone on the production of IL-6, TNF-alpha and IL-10 of RAW 264.7 cells stimulated with poly (I:C). Eurycomanone inhibited production of pro-inflammatory cytokines including IL-6 and TNF-alpha of RAW 264.7 cells stimulated with poly (I:C) with the IC50 value being 5.14 +/- 0.60 uM and 2.32 +/- 0.40 uM, respectively. Interestingly, this compound also inhibited the production of anti-inflammatory cytokine IL-10 of RAW 264.7 cells stimulated with poly (I:C) with the IC50 value being 14.60 +/- 0.32 uM. These results suggested that eurycomanone has a potentially regulatory role on production of both pro-inflammatory and anti-inflammatory cytokines in macrophage in the mimicking context of virus infection. Whether eurycomanone inhibits the binding of poly (I:C)-TLR3 or intracellular signaling pathways needs further investigation.