α-Ketoglutarate plays an inflammatory inhibitory role by regulating scavenger receptor class a expression through N6-methyladenine methylation during sepsis.

Sepsis arises from an uncontrolled inflammatory response triggered by infection or stress, accompanied by alteration in cellular energy metabolism, and a strong correlation exists between these factors. Alpha-ketoglutarate (α-KG), an intermediate product of the TCA cycle, has the potential to modulate the inflammatory response and is considered a crucial link between energy metabolism and inflammation. The scavenger receptor (SR-A5), a significant pattern recognition receptor, assumes a vital function in anti-inflammatory reactions. In the current investigation, we have successfully illustrated the ability of α-KG to mitigate inflammatory factors in the serum of septic mice and ameliorate tissue damage. Additionally, α-KG has been shown to modulate metabolic reprogramming and macrophage polarization. Moreover, our findings indicate that the regulatory influence of α-KG on sepsis is mediated through SR-A5. We also elucidated the mechanism by which α-KG regulates SR-A5 expression and found that α-KG reduced the N6-methyladenosine level of macrophages by up-regulating the m6A demethylase ALKBH5. α-KG plays a crucial role in inhibiting inflammation by regulating SR-A5 expression through m6A demethylation during sepsis. The outcomes of this research provide valuable insights into the relationship between energy metabolism and inflammation regulation, as well as the underlying molecular regulatory mechanism.

SP1 promotes high glucose-induced lens epithelial cell viability, migration and epithelial-mesenchymal transition via regulating FGF7 and PI3K/AKT pathway.

BACKGROUND: Diabetic cataract (DC) is a common complication of diabetes and its etiology and progression are multi-factorial. In this study, the roles of specific protein 1 (SP1) and fibroblast growth factor 7 (FGF7) in DC development were explored. METHODS: DC cell model was established by treating SRA01/04 cells with high glucose (HG). MTT assay was conducted to evaluate cell viability. Transwell assay and wound-healing assay were performed to assess cell migration and invasion. Western blot assay and qRT-PCR assay were conducted to measure the expression of N-cadherin, E-cadherin, Collagen I, Fibronectin, SP1 and FGF7 expression. CHIP assay and dual-luciferase reporter assay were conducted to analyze the combination between FGF7 and SP1. RESULTS: FGF7 was upregulated in DC patients and HG-induced SRA01/04 cells. HG treatment promoted SRA01/04 cell viability, migration, invasion and epithelial-mesenchymal transition (EMT), while FGF7 knockdown abated the effects. Transcription factor SP1 activated the transcription level of FGF7 and SP1 overexpression aggravated HG-induced SRA01/04 cell injury. SP1 silencing repressed HG-induced SRA01/04 cell viability, migration, invasion and EMT, but these effects were ameliorated by upregulating FGF7. Additionally, SP1 knockdown inhibited the PI3K/AKT pathway by regulating the transcription level of FGF7. CONCLUSION: Transcription factor SP1 activated the transcription level of FGF7 and the PI3K/AKT pathway to regulate HG-induced SRA01/04 cell viability, migration, invasion and EMT.

Factors enhancing and restricting the success of SRA compliance on the FLEGT/VPA initiative in Ghana. Insights from Juaso forest district in Ghana.

This study examines the Voluntary Partnership Agreement (VPA) between Ghana and the European Union (EU) within the Forest Law Enforcement Governance and Trade Initiative (FLEGT). The VPA aims to enhance forest governance, reduce deforestation, combat illegal practices, and improve livelihoods of forest fringe communities. The research focuses on the implementation of social responsibility agreements (SRAs) under the VPA framework and identifies factors contributing to their success or presenting challenges. Data collection involved mixed methods, including literature review and a survey of individuals involved in SRAs. Descriptive and inferential statistical analyses, including exploratory factor analysis, were conducted. Principal component analysis revealed that accountability, monitoring of implementation and progress of SRAs, and documentation of SRA agreements were key factors contributing to the success of negotiated SRAs, explaining about 68.36 % of success variance. Challenges and constraints were categorized into two main factors: weak community capacity to negotiate SRAs and weak community capacity to enforce compliance, explaining about 71.4 % of challenge variance. The study found that the exclusion of the local SRA committee (LSRAC) from certain decision-making processes affected trust and transparency in calculating SRA benefits. Elite capture of benefits was identified as an issue, as the LSRAC did not conduct sufficient consultations with community members before negotiations. The findings emphasize the importance of including local communities in all forest management activities and call for increased awareness of SRAs, particularly for the LSRAC. The study highlights the need for proper representation of community interests during negotiations and their inclusion in forest management plans.

Circ_0000099/miR-223-3p/CTGF Regulates the Growth, Metastasis, and EMT Processes in TGF-ß2-Stimulated Human Lens Epithelial Cells.

PURPOSE: Posterior capsule opacification (PCO) is the major complication of visual impairment after cataract surgery. Circular RNAs (circRNAs) are involved in the development of many diseases. The purpose of this study was to explore the role and molecular mechanism of circ_0000099 in PCO. METHODS: SRA01/04 cells were treated with TGF-ß2 to establish a PCO cell model. The expression of circ_0000099, miR-223-3p, and connective tissue growth factor (CTGF) mRNA was determined by real-time quantitative polymerase chain reaction (qRT-PCR). Western blot assay was used to analyze the protein expression. Cell proliferation, migration, and invasion were analyzed by (4-5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), 5-ethynyl-2 '-Deoxyuridine (EdU), transwell, and wound healing tests. The circ_0000099/miR-223-3p/CTGF relationship was verified by dual luciferase reporter gene and RNA binding protein immunoprecipitation (RIP) assays. RESULTS: TGF-ß2 treatment promoted SRA01/04 cell proliferation invasion, migration, and EMT. Circ_0000099 expression was increased in POC patients and TGF-ß2-treated SRA01/04 cells.Knockdown of circ_0000099 suppressed TGF-ß2-induced proliferation, invasion, migration, and EMT in SRA01/04 cells. miR-223-3p was identified as the target of circ_0000099, and miR-223-3p inhibitor might partly abolish the repression of circ_0000099 silencing on TGF-ß2-triggered SRA01/04 cell disorders. MiR-223-3p directly targeted CTGF. Knockdown of CTGF suppressed TGF-ß2-induced SRA01/04 cell injury. Circ_0000099 can regulate CTGF expression by targeting miR-223-3p. CONCLUSIONS: Circ_0000099 silencing might relieve TGF-2-induced SRA01/04 cell injury by the miR-223-3p/CTGF axis, providing new avenues for the prevention and treatment of PCO.

Detailed role of SR-A1 and SR-E3 in tumor biology, progression, and therapy.

The first host defense systems are the innate immune response and the inflammatory response. Among innate immune cells, macrophages, are crucial because they preserve tissue homeostasis and eradicate infections by phagocytosis, or the ingestion of particles. Macrophages exhibit phenotypic variability contingent on their stimulation state and tissue environment and may be detected in several tissues. Meanwhile, critical inflammatory functions are played by macrophage scavenger receptors, in particular, SR-A1 (CD204) and SR-E3 (CD206), in a variety of pathophysiologic events. Such receptors, which are mainly found on the surface of multiple types of macrophages, have different effects on processes, including atherosclerosis, innate and adaptive immunity, liver and lung diseases, and, more recently, cancer. Although macrophage scavenger receptors have been demonstrated to be active across the disease spectrum, conflicting experimental findings and insufficient signaling pathways have hindered our comprehension of the molecular processes underlying its array of roles. Herein, as SR-A1 and SR-E3 functions are often binary, either protecting the host or impairing the pathophysiology of cancers has been reviewed. We will look into their function in malignancies, with an emphasis on their recently discovered function in macrophages and the possible therapeutic benefits of SR-A1 and SR-E3 targeting.

Rate of atrial fibrillation by Holter-Stroke Risk Analysis in undetermined TIA/rapidly improving stroke symptoms patients.

Introduction: Holter-SRA (Stroke Risk Analysis) is an automated analysis of ECG monitoring for Atrial Fibrillation (AF) detection. The aim of this study was to evaluate the rate of AF in undetermined TIA/Rapidly improving stroke symptoms (RISS) patients. Methods: Prospective study of undetermined TIA/RISS patients who presented to the emergency department. Early vascular studies (angio CT, transthoracic echocardiography and ECG) were performed in emergency department. The Holter-SRA device was placed for 2 h and the patients were classified into: confirmed AF, high risk of AF or low risk of AF. Prolonged ambulatory monitoring (7 days) was carried out every month for patients with a high-risk pattern. The results were evaluated until definitive detection of AF or low-risk pattern. The endpoints were rate of AF and vascular recurrence at 90 days. Results: Over a period of 24 months, 83 undetermined TIA/RISS patients were enrolled. The mean age was 70 ± 10 years and 61% were men. The median ABCD2 score was 4 points (1-7). After 2 h of monitoring in the emergency department, AF was detected in one patient (1.2%), 51 patients with a low-risk pattern and 31 patients (37.3%) showed a high-risk pattern of AF. During the ambulatory monitoring, of the 31 patients high risk pattern patients, AF was diagnosed to 17 cases and of the 51 patients with a low-risk pattern, one case experienced a recurrent vascular due to undetected AF (1.9% false negative). Three patients (3.6%) suffered a vascular recurrence within the first 90 days, before AF diagnosis. Conclusions: In our study, AF was detected in 22.9% of the 83 patients with indeterminate TIA/RISS. Holter-SRA has allowed us to increase the detection of AF, especially those patients with a high-risk pattern in the first 3 months.

Macrophage scavenger receptor-A1 promotes skeletal muscle regeneration after hindlimb ischemia.

Macrophages mediated inflammatory response is crucial for the recovery of skeletal muscle following ischemia. Thus, it's necessary to exploit macrophages based therapeutic targets for ischemic disease. Here, we found mRNA level of SR-A1 was elevated in patients with critical limb ischemia by analysis of gene expression omnibus (GEO) database. Then we investigated the role and the underlined mechanisms of macrophage SR-A1 in a mouse HLI model. Compared with the SR-A1 fl/fl mice, the Lyz Cre/+/SR-A1 flox/flox (SR-A1 ΔMΦ) mice showed significantly lower laser doppler blood flow in the ischemic limb at day 7 after HLI. Consistently, histological analysis exhibited that ischemic limb of SR-A1 ΔMΦ mice displayed more sever and sustained necrotic morphology, inflammation and fibrosis, decreased vessel density and regeneration rate, compared with which of control SR-A1 fl/fl mice. Furthermore, restoration of wild-type myeloid cells to SR-A1 knock-out mice effectively relieved the doppler perfusion in the ischemic limb and restrained skeletal muscle damage 7 days post HLI. In line with in vivo findings, when co-cultivating macrophages with the mouse myoblast line C2C12, SR-A1 -/- bone marrow macrophage significantly inhibited myoblast differentiation in vitro. Mechanically, SR-A1 enhanced skeletal muscle regeneration response to HLI by inhibiting the oncostatin M (OSM) production via suppressed NF-κB signaling activation. These results indicates that SR-A1 is a promising candidate molecule to improve tissue repair and regeneration in peripheral ischemic arterial disease.

Medical ozone alleviates acute lung injury by enhancing phagocytosis targeting NETs via AMPK/SR-A1 axis.

Acute lung injury (ALI) linked to sepsis has a high mortality rate, with limited treatment options available. In recent studies, medical ozone has shown promising results in alleviating inflammation and infection. Here, we aimed to evaluate the therapeutic potential of medical ozone in sepsis-induced ALI using a mouse model, measuring behavioral assessments, lung function, and blood flow. Western blot was used to quantify the levels of protein. In vitro, experiments on BMDM cells examine the impact of AMPK inhibitors and agonists on phagocytic activity. Results indicate that medical ozone can enhance the survival rate, ameliorate lung injury, and improve lung function and limb microcirculation in mice with ALI. Notably, it inhibits NETs formation, a crucial player in ALI development. Medical ozone also counteracts elevated TF, MMP-9, and IL-1ß levels. In ALI mice, the effects of ozone are nullified and BMDMs exhibit impaired engulfment of NETs following Sr-a1 knockout. Under normal physiological conditions, the use of an AMPK antagonist produces similar effects to Sr-a1 knockout, significantly inhibiting the phagocytosis of NETs by BMDMs. On the contrary, AMPK agonists enhance this phagocytic process. In conclusion, medical ozone can alleviate sepsis-induced lung injury via the AMPK/SR-A1 pathway, thereby enhancing phagocytosis of NETs by macrophages.

Analyses of Nuclear Reads Obtained Using Genome Skimming.

In this protocol paper, we review a set of methods developed in recent years for analyzing nuclear reads obtained from genome skimming. As the cost of sequencing drops, genome skimming (low-coverage shotgun sequencing of a sample) becomes increasingly a cost-effective method of measuring biodiversity at high resolution. While most practitioners only use assembled over-represented organelle reads from a genome skim, the vast majority of the reads are nuclear. Using assembly-free and alignment-free methods described in this protocol, we can compare samples to each other and reference genomes to compute distances, characterize underlying genomes, and infer evolutionary relationships.

Long non-coding RNA SRA1 suppresses radiotherapy resistance in esophageal squamous cell carcinoma by modulating glycolytic reprogramming.

Esophageal squamous cell carcinoma (ESCC), a highly aggressive subtype of esophageal cancer, is characterized by late-stage diagnosis and limited treatment options. Recent advancements in transcriptome sequencing technologies have illuminated the molecular intricacies of ESCC tumors, revealing metabolic reprogramming as a prominent feature. Specifically, the Warburg effect, marked by enhanced glycolysis, has emerged as a hallmark of cancer, offering potential therapeutic targets. In this study, we comprehensively analyzed bulk RNA-seq data from ESCC patients, uncovering elevated SRA1 expression in ESCC development and a poorer prognosis. Silencing of SRA1 led to a modulation of glycolysis-related products and a shift in PKM2 expression. Our findings shed light on the intricate molecular landscape of ESCC, highlighting SRA1 as a potential therapeutic target to disrupt glycolysis-dependent energy production. This metabolic reprogramming may hold the key to innovative treatment strategies for ESCC, ultimately improving patient outcomes.

The Hsa_circ_0105558/miR-182-5p/ATF6 Cascade Affects H2O2-Triggered Oxidative Damage and Apoptosis of Human Lens Epithelial Cells.

Age-related cataract (ARC) is the prevalent cause of useful vision loss. Circular RNAs are related to ARC pathogenesis partly through their competing endogenous RNA (ceRNA) activity. Herein, we defined the action of hsa_circ_0105558 in hydrogen peroxide (H2O2)-driven apoptosis and oxidative damage in human lens epithelial SRA01/04 cells. Hsa_circ_0105558, microRNA (miR)-182-5p and activating transcription factor 6 (ATF6) were evaluated by a qRT-PCR or immunoblotting method. The hsa_circ_0105558/miR-182-5p and miR-182-5p/ATF6 relationships were predicted by bioinformatics analysis and confirmed by dual-luciferase reporter assay. Reactive oxygen species level, glutathione peroxidase level, superoxide dismutase activity, and malondialdehyde level were measured using the matched assay kits. Hsa_circ_0105558 was upregulated in human ARC lens and H2O2-exposed SRA01/04 cells. Suppression of hsa_circ_0105558 attenuated H2O2-driven SRA01/04 cell apoptosis and oxidative damage. Hsa_circ_0105558 targeted miR-182-5p, and reduced miR-182-5p expression reversed the influence of hsa_circ_0105558 depletion on H2O2-driven oxidative damage and apoptosis. ATF6 was a target of miR-182-5p, and miR-182-5p-driven downregulation of ATF6 regulated cell oxidative damage and apoptosis under H2O2 insult. Moreover, hsa_circ_0105558 functioned as a ceRNA to post-transcriptionally control ATF6 expression through miR-182-5p competition. Our study demonstrates that hsa_circ_0105558 modulates SRA01/04 cell oxidative damage and apoptosis under H2O2 insult through the miR-182-5p/ATF6 cascade.

Understanding the Role of Scavenger Receptor A1 in Nanoparticle Uptake by Murine Macrophages.

Nanoparticles can be cleared from the circulation and taken up by tissue-resident macrophages. This property can be beneficial when drug or antigen delivery to macrophages is desired; however, rapid clearance of nanoparticles not intended for delivery to immune cells may reduce nanoparticle circulation time and affect the efficacy of nanoparticle-formulated drug products. Therefore, understanding nanoparticles' uptake by macrophages is an essential step in the preclinical development of nanotechnology-based drug products. Understanding the route of nanoparticle uptake by macrophages may also provide mechanistic insights into the immunotoxicity of nanomaterials. The protocol described herein can be used to assess the nanoparticles' uptake by macrophages and understand the involvement of scavenger receptor A1 to inform mechanistic studies.

Aggregation of recount3 RNA-seq data improves inference of consensus and tissue-specific gene co-expression networks.

Background: Gene co-expression networks (GCNs) describe relationships among expressed genes key to maintaining cellular identity and homeostasis. However, the small sample size of typical RNA-seq experiments which is several orders of magnitude fewer than the number of genes is too low to infer GCNs reliably. recount3, a publicly available dataset comprised of 316,443 uniformly processed human RNA-seq samples, provides an opportunity to improve power for accurate network reconstruction and obtain biological insight from the resulting networks. Results: We compared alternate aggregation strategies to identify an optimal workflow for GCN inference by data aggregation and inferred three consensus networks: a universal network, a non-cancer network, and a cancer network in addition to 27 tissue context-specific networks. Central network genes from our consensus networks were enriched for evolutionarily constrained genes and ubiquitous biological pathways, whereas central context-specific network genes included tissue-specific transcription factors and factorization based on the hubs led to clustering of related tissue contexts. We discovered that annotations corresponding to context-specific networks inferred from aggregated data were enriched for trait heritability beyond known functional genomic annotations and were significantly more enriched when we aggregated over a larger number of samples. Conclusion: This study outlines best practices for network GCN inference and evaluation by data aggregation. We recommend estimating and regressing confounders in each data set before aggregation and prioritizing large sample size studies for GCN reconstruction. Increased statistical power in inferring context-specific networks enabled the derivation of variant annotations that were enriched for concordant trait heritability independent of functional genomic annotations that are context-agnostic. While we observed strictly increasing held-out log-likelihood with data aggregation, we noted diminishing marginal improvements. Future directions aimed at alternate methods for estimating confounders and integrating orthogonal information from modalities such as Hi-C and ChIP-seq can further improve GCN inference.

Optimization of laboratory diagnosis of heparin-induced thrombocytopenia using HemosIL-AcuStar-HIT-IgG assay.

OBJECTIVE: The aim of this study was to determine an optimal cutoff value for the newly available HemosIL-AcuStar-HIT-IgG assay (AcuStar) for the diagnosis of heparin-induced thrombocytopenia (HIT). METHOD: We evaluated the performance of AcuStar using serotonin release assay (SRA) as the gold standard and incorporated 4T score calculation in a cohort of suspected HIT cases. Statistical analysis was performed to determine optimal cutoff value for the diagnosis of HIT. RESULT: A diagnosis of HIT can be excluded with a platelet factor 4 (PF4) value of <0.4 U/mL by AcuStar and 4T score in the low-risk category (≤3). All other cases will require confirmation with a functional test. CONCLUSION: Our study resulted in the implementation of a diagnostic algorithm for laboratory diagnosis of HIT, which incorporates pretest calculation of 4T score and AcuStar as a screening test, with reflex confirmation by SRA. This new algorithm resulted in extended hours of test availability and a more rapid turnaround time in reporting PF4 results.

Identification of potential biomarkers for melanoma cancer (black tumor) using bioinformatics strategy: a study based on GEO and SRA datasets.

Melanoma, a highly invasive type of skin cancer that penetrates the entire dermis layer, is associated with increased mortality rates. Excessive exposure of the skin to sunlight, specifically ultraviolet radiation, is the underlying cause of this malignant condition. The appearance of unique skin moles represents a visible clue, referred to as the "ugly duckling" sign, indicating the presence of melanoma and its association with cellular DNA damage. This research aims to explore potential biomarkers derived from microarray data, employing bioinformatics techniques and methodologies, for a thorough investigation of melanoma skin cancer. The microarray dataset for melanoma skin cancer was obtained from the GEO database, and thorough data analysis and quality control measures were performed to identify differentially expressed genes (DEGs). The top 14 highly expressed DEGs were identified, and their gene information and protein sequences were retrieved from the NCBI gene and protein database. These proteins were further analyzed for domain identification and network analysis. Gene expression analysis was conducted to visualize the upregulated and downregulated genes. Additionally, gene metabolite network analysis was carried out to understand the interactions between highly interconnected genes and regulatory transcripts. Molecular docking was employed to investigate the ligand-binding sites and visualize the three-dimensional structure of proteins. Our research unveiled a collection of genes with varying expression levels, some elevated and others reduced, which could function as promising biomarkers closely linked to the development and advancement of melanoma skin cancer. Through molecular docking analysis of the GINS2 protein, we identified two natural compounds (PubChem-156021169 and PubChem-60700) with potential as inhibitors against melanoma. This research has implications for early detection, treatment, and understanding the molecular basis of melanoma.

A SNARE protective pool antagonizes APOL1 renal toxicity in Drosophila nephrocytes.

BACKGROUND: People of Sub-Saharan African ancestry are at higher risk of developing chronic kidney disease (CKD), attributed to the Apolipoprotein L1 (APOL1) gene risk alleles (RA) G1 and G2. The underlying mechanisms by which the APOL1-RA precipitate CKD remain elusive, hindering the development of potential treatments. RESULTS: Using a Drosophila genetic modifier screen, we found that SNARE proteins (Syx7, Ykt6, and Syb) play an important role in preventing APOL1 cytotoxicity. Reducing the expression of these SNARE proteins significantly increased APOL1 cytotoxicity in fly nephrocytes, the equivalent of mammalian podocytes, whereas overexpression of Syx7, Ykt6, or Syb attenuated their toxicity in nephrocytes. These SNARE proteins bound to APOL1-G0 with higher affinity than APOL1-G1/G2, and attenuated APOL1-G0 cytotoxicity to a greater extent than either APOL1-RA. CONCLUSIONS: Using a Drosophila screen, we identified SNARE proteins (Syx7, Ykt6, and Syb) as antagonists of APOL1-induced cytotoxicity by directly binding APOL1. These data uncovered a new potential protective role for certain SNARE proteins in the pathogenesis of APOL1-CKD and provide novel therapeutic targets for APOL1-associated nephropathies.

Genomic data of two chickpea populations sharing a potential Ascochyta blight resistance region.

Chickpea (Cicer arietinum L.) is one of the most important crops worldwide and a valuable nutritional source. The availability of data from different genotypes and populations is important for the comprehension of the biology and trait control of chickpea. Tissue from young leaves was collected from adult plants and sequenced using an Illumina HiSeq X platform, which provided sequencing data for a total of 169 individuals from two different populations. Furthermore, functional annotation was performed with BLAST2GO software in a candidate region for resistance to Ascochyta blight, a devastating disease that produces huge yield reductions if the growth conditions are optimal for the fungus. A total of 273 different genes in a region spanning ∼4.67 Mb in chromosome 4 were fully annotated. The raw DNA sequences and functional annotation data can be reused by the scientific community for the analysis of different agronomic traits of interest in chickpea.

Role of Albumin as a Targeted Drug Carrier in the Management of Rheumatoid Arthritis: A Comprehensive Review.

An endogenous transporter protein called albumin interacts with the Fc receptor to provide it with multiple substrate-binding domains, cell membrane receptor activation, and an extended circulating half-life. Albumin has the remarkable ability to bind with receptors viz. secreted protein acidic and rich in cysteine (SPARC) and scavenger protein-A (SR-A) that are overexpressed during rheumatoid arthritis (RA), enabling active targeting of the disease site instead of requiring specialized substrates to be added to the nanocarrier. RA, a chronic autoimmune illness, is characterized by the presence of a severe inflammatory response. RA patients have low serum albumin concentration, which signifies the high uptake of albumin at the inflammatory sites, giving a rationale to use albumin as a drug carrier for RA therapy. Albumin has the capacity for both passive and active targeting. It is an abundantly available protein in the bloodstream showing excellent cellular compatibility, degradability in biological tissues, nonantigenicity, and safety. There are three strategies of albumin mediated drug delivery as encapsulating therapeutics in albumin nanoparticles, chemically conjugating drugs with functional proteins, and albumin itself which is used as a targeting ligand to deliver drugs specifically to cells or tissues that express albumin-binding receptors. In the current review, an attempt has been made to highlight the significant evidence of albumin as a drug delivery carrier for the safe and effective management of RA. Evidence has been provided in the form of recent research advances, clinical trials, and patents. Additionally, this review will outline the prospective for the potential utilization of albumin as a drug vehicle for RA and suggest possible future avenues to provide the perspective for subsequent studies.

A Functional Assay for the Determination of Heparin-Induced Thrombocytopenia via Flow Cytometry.

Heparin-induced thrombocytopenia (HIT) is a life-threatening complication of heparin therapy (both unfractionated heparin and low-molecular-weight heparin). In our study, we examined a group of 122 patients with suspected HIT. The samples of all patients were analyzed in the first step using an immunoassay (ID-PaGIA Heparin/PF4, Hemos1L-Acustar HIT IgG, ZYMUTEST HIA Monostrip IgG) to detect the presence of antibodies against heparin-PF4 complexes (platelet factor 4). When the immunoassay was positive, the sample was subsequently analyzed for HIT with a functional flow cytometry assay, the HITAlert kit, the purpose of which was to demonstrate the ability of the antibodies present to activate platelets. A diagnosis of HIT can be made only after a positive functional test result. In this article, we present an overview of our practical experience with the use of the new functional method of analysis, HIT, with flow cytometry. In this work, we compared the mutual sensitivity of two functional tests, SRA and the flow cytometry HITAlert kit, in patients perceived as being at risk for HIT. This work aims to delineate the principle, procedure, advantages, pitfalls, and possibilities of the application of the functional test HITAlert using flow cytometry.

Promoting AMPK/SR-A1-mediated clearance of HMGB1 attenuates chemotherapy-induced peripheral neuropathy.

BACKGROUND: Chemotherapy-induced peripheral neuropathy (CIPN) is a serious side effect of chemotherapy with poorly understood mechanisms and few treatments. High-mobility group box 1 (HMGB1)-induced neuroinflammation is the main cause of CIPN. Here, we aimed to illustrate the role of the macrophage scavenger receptor A1 (SR-A1) in HMGB1 clearance and CIPN resolution. METHODS: Oxaliplatin (L-OHP) was used to establish a CIPN model. Recombinant HMGB1 (rHMGB1) (his tag) was used to evaluate the phagocytosis of HMGB1 by macrophages. RESULTS: In the clinic, HMGB1 expression and MMP-9 activity were increased in the plasma of patients with CIPN. Plasma HMGB1 expression was positively correlated with the cumulative dose of L-OHP and the visual analog scale. In vitro, engulfment and degradation of rHMGB1 increased and inflammatory factor expression decreased after AMP-activated protein kinase (AMPK) activation. Neutralizing antibodies, inhibitors, or knockout of SR-A1 abolished the effects of AMPK activation on rHMGB1 engulfment. In vivo, AMPK activation increased SR-A1 expression in the dorsal root ganglion, decreased plasma HMGB1 expression and MMP-9 activity, and attenuated CIPN, which was abolished by AMPK inhibition or SR-A1 knockout in the CIPN mice model. CONCLUSION: Activation of the AMPK/SR-A1 axis alleviated CIPN by increasing macrophage-mediated HMGB1 engulfment and degradation. Therefore, promoting HMGB1 clearance may be a potential treatment strategy for CIPN. Video abstract.