Extracellular Space Research Articles

Effect of Extracellular Vesicles Derived From Tumor Cells on Immune Evasion.

The crosstalk between immunity and cancer in the regulation of tumor growth is considered a hallmark of cancer. Antitumor immunity refers to the innate and adaptive immune responses that regulate cancer development and proliferation. Tumor immune evasion represents a major hindrance to effective anticancer treatment. Extracellular vesicles (EVs) are nano-sized and lipid-bilayer-enclosed particles that are secreted to the extracellular space by all cell types. They are critically involved in numerous biological functions including intercellular communication. Tumor-derived extracellular vesicles (TEVs) can transport a variety of cargo to modulate immune cells in the tumor microenvironment (TME). This review provides the latest update about how tumor cells evade immune surveillance by exploiting TEVs. First, the biogenesis of EVs and the cargo-sorting machinery are discussed. Second, how tumor cells modulate immune cell differentiation, activation, and function via TEVs to evade immune surveillance is illustrated. Last but not least, the novel antitumor strategies that can reverse immune escape are summarized.

MUC5AC levels as predictors of adjuvant therapy outcomes in pancreatic ductal adenocarcinoma: A study of post-surgical gemcitabine-based regimens.

759 Background: The role of tissue mucin 5 AC (MUC5AC) in PDA is not fully understood. In our previous work we showed that mature MUC5AC (MM) detected in the extracellular (EC) space and in apical region has better prognostic value than perinuclear immature MUC5AC (IM). Our previous research demonstrated that mature MUC5AC (MM), present in the extracellular (EC) space and apical region, offers better prognostic value than perinuclear immature MUC5AC (IM). Preclinical studies suggest that MM may confer chemoresistance to gemcitabine (Gem), commonly used in adjuvant therapy (AT) either alone or with capecitabine (Cap) or nab-paclitaxel (NP) in patients not eligible for FOLFIRINOX. This study investigates MM’s effect on outcomes in PDA patients undergoing post-surgical treatment with Gem-only, Gem-NP, and Gem-Cap. Methods: Formalin-fixed, paraffin-embedded tissue samples from resected PDA cases (January 2010 - June 2021) were sourced from the Ohio State University biorepository. Immunohistochemistry using monoclonal antibodies 45M1 (for MM) and CLH2 (for IM) was employed to examine cellular MM and IM expression, with H-scores calculated. Statistical analysis of progression-free survival (PFS) and overall survival (OS) was done using the log-rank test. Results: We had 49 patients available for analysis. The median age of the group was 66 years (range: 38 to 89) with majority Caucasians (90%) and 53% (n=26) females. AT distribution within this group was, 31 Gem-only, 7 Gem-NP, and 11 Gem and capecitabine (Cap). Median expression levels for MM and IM were both 150, and 82% (n=40) were EC-MM positive. Using the median MM H-score as a threshold (< 150 vs. ≥ 150), outcomes were evaluated within each therapy subgroup (Table). Conclusions: This preliminary study suggests that PDA patients with low MM expression may benefit from Gem-Cap therapy. However, larger, prospective studies are needed for definitive conclusions. Comparing outcomes in the sub-groups. Sub-group (n) OS in days (p-value) PFS in days (p-value) MM H-score < 150 vs. ≥ 150 Gem-only (11 vs. 20) 775 vs. 1006 (0.4) 444 vs. 413 (0.4) Gem NP (4 vs. 3) 1409 vs. 551 (0.8) 391 vs. 418 (0.5) Gem-Cap (4 vs. 7) not evaluable (NE) vs. 766 (0.02) NE vs. 415 (0.01)

Intracellular enhancement technique for gadoxetic acid-enhanced hepatobiliary-phase magnetic resonance imaging: evaluation of hepatic function.

To investigate the utility of intracellular enhancement (ICE) technique which suppresses signals from the extracellular space for the evaluation of hepatic function on gadoxetic acid-enhanced hepatobiliary-phase (HBP) images. We subjected 67 patients with suspected neoplastic hepatic lesions to gadoxetic acid-enhanced HBP imaging with and without ICE [i-HBP, conventional-HBP (c-HBP)]. A radiologist calculated the liver/spleen contrast (LSC) [LSC = signal intensity (SI) of liver/SI of spleen]. Receiver-operating analysis was used to evaluate the diagnostic value of the LSC on i-HBP- (i-LSC) and c-HBP images (c-LSC) to differentiate between Child-Pugh classes A and B. Of the 67 patients, 57 were in Child-Pugh class A and 10 were in class B. For their differentiation, the area under the curve value of i-LSC was higher than of c-LSC (0.81 vs. 0.68). ICE technique can improve the accuracy of estimating hepatic function on HBP images.

Downmodulation of potassium conductances induces epileptic seizures in cortical network models via multiple synergistic factors.

Voltage-gated potassium conductances [Formula: see text] play a critical role not only in normal neural function, but also in many neurological disorders and related therapeutic interventions. In particular, in an important animal model of epileptic seizures, 4-aminopyridine (4-AP) administration is thought to induce seizures by reducing [Formula: see text] in cortex and other brain areas. Interestingly, 4-AP has also been useful in the treatment of neurological disorders such as multiple sclerosis (MS) and spinal cord injury, where it is thought to improve action potential propagation in axonal fibers. Here, we examined [Formula: see text] downmodulation in bio-physical models of cortical networks that included different neuron types organized in layers, potassium diffusion in interstitial and larger extracellular spaces, and glial buffering. Our findings are fourfold. First, [Formula: see text] downmodulation in pyramidal and fast-spiking inhibitory interneurons led to differential effects, making the latter much more likely to enter depolarization block. Second, both neuron types showed an increase in the duration and amplitude of action potentials, with more pronounced effects in pyramidal neurons. Third, a sufficiently strong [Formula: see text] reduction dramatically increased network synchrony, resulting in seizure like dynamics. Fourth, we hypothesized that broader action potentials were likely to not only improve their propagation, as in 4-AP therapeutic uses, but also to increase synaptic coupling. Notably, graded synapses incorporating this effect further amplified network synchronization and seizure-like dynamics. Overall, our findings elucidate different effects that [Formula: see text] downmodulation may have in cortical networks, explaining its potential role in both pathological neural dynamics and therapeutic applications.Significance Statement The modulation of voltage-gated potassium-conductances [Formula: see text] is thought to play an important role in epileptic seizures and therapeutic interventions in epilepsy, multiple sclerosis and spinal-cord injury. We show that [Formula: see text] downmodulation can lead to a cascade of effects including changes in basal excitability, broadening of action potentials resulting in enhanced robustness to synaptic noise perturbations and strengthening of synaptic coupling; and differential effects in excitatory and fast-spiking inhibitory interneurons, promoting depolarization block in the latter under high downmodulation. All these effects synergistically contribute to the emergence of seizure-like dynamics in the form of almost-periodic synchronized neuronal-population spiking in cortical networks. Under appropriate levels, [Formula: see text] downmodula tion can also have therapeutic effects by improving neuronal communication via the broadening of action potentials.

Neuropilin1-dependent paracrine signaling of cancer cells mediated by miRNA exosomal cargo

BackgroundNeuropilin-1 (NRP1) is a transmembrane protein involved in surface receptor complexes for a variety of extracellular signals. NRP1 expression in human cancers is associated with prominent angiogenesis and advanced progression stage. However, the molecular mechanisms underlying NRP1 activity in the tumor microenvironment remain unclear. Notably, diffusible forms of NRP1 in the extracellular space have been reported, but their functional role is poorly understood.MethodsExtracellular vesicles (EV) were isolated from conditioned media of diverse cancer cells. The quality of exosome-enriched preparations was validated by the presence of specific markers in western blotting, as well as by light scattering and nanoparticle tracking analysis. Wound healing, transwell, and digital real-time migration assays were carried out to assess the activity of cancer cell-derived exosomes in the regulation of endothelial cells. RNA interference was applied to obtain NRP1 knock-down, and cDNA transfer to achieve its overexpression, in exosome-releasing cells. The micro-RNA profile carried by exosomes was investigated by Next Generation Sequencing. miRNA-Scope in situ hybridization was used to assess the transfer of miRNA exosome cargo to target cells, and immunofluorescence analysis revealed expression regulation of targeted proteins. miRNA activity was blocked by the use of specific antago-miRs.ResultsIn this study, we show that diverse human cancer cells release NRP1 embedded in exosome-like small extracellular vesicles, which mediate a previously unknown NRP1-dependent paracrine signaling mechanism regulating endothelial cell migration. By transcriptomic analysis of the cargo of NRP1-loaded exosomes, we found a significant enrichment of miR-210-3p, known to promote tumor angiogenesis. Gene knock-down and overexpression experiments demonstrated that the loading of miR-210-3p into exosomes is dependent on NRP1. Data furthermore indicate that the exosomes released through this NRP1-driven mechanism effectively transfer miR-210-3p to human endothelial cells, causing paracrine downregulation of the regulatory cue ephrin-A3 and promotion of cell migration. The mechanistic involvement of miR-210-3p in this pathway was confirmed by applying a specific antago-miR.ConclusionsIn sum, we unveiled a previously unknown NRP1-dependent paracrine signaling mechanism, mediated by the loading of pro-angiogenic miR-210-3p in exosomes released by cancer cells, which underscores the relevance of NRP1 in controlling the tumor microenvironment.

The secreted host-cell protein clusterin interacts with PmpD and promotes Chlamydia trachomatis infection

Attachment and uptake into host cells are pivotal steps in the life cycle of the Chlamydiaceae, a family of obligate intracellular pathogens. Chlamydia trachomatis (Ctr) possesses a family of nine polymorphic membrane proteins (Pmps), which have been shown to be crucial for adhesion and internalization. However, the host-cell molecules involved have so far remained unknown. Here, we show that a fragment of Ctr PmpD, which forms high-molecular-weight oligomers in solution and adheres to epithelial cells, also binds to secreted clusterin (sCLU), a chaperone-like protein that is secreted into the extracellular space by the host cell, and forms part of the chaperone- and receptor-mediated extracellular protein degradation (CRED) pathway. Using in vitro assays, we demonstrate that sCLU interacts directly with soluble rPmpD. In infection experiments, depletion of sCLU from the culture medium leads to a significant decrease in Ctr infection. Thus, sCLU is the first host-cell interaction partner identified for a Ctr Pmp and the first case in which sCLU has been shown to be a vital component for the establishment of a bacterial infection.

Whole-transcriptome analysis reveals the profiles and roles of coding and non-coding RNAs during hair follicle cycling in Rex rabbits

BackgroundRex rabbit is famous for its silky and soft fur coat, a characteristic predominantly attributed to its hair follicles. Numerous studies have confirmed the crucial roles of mRNAs and non-coding RNAs (ncRNAs) in regulating key cellular processes such as cell proliferation, differentiation, apoptosis and immunity. However, their involvement in the regulation of the hair cycle in Rex rabbits remains unknown.ResultsIn this study, we identified the hair follicle stages of Rex rabbits aged 3 to 5.5 months. Skin samples collected at 4, 5 and 5.5 months, representing the morphological features of the anagen, catagen and telogen stage separately, were finally selected for whole-transcriptome analysis. 25,736 mRNA, 8280 lncRNA, 24,885 circRNA and 1138 miRNA transcripts were identified. 6027 differently expressed mRNAs (DEGs), 2381 differently expressed lncRNAs (DELs), 438 differently expressed circRNAs (DECs) and 167 differently expressed miRNAs (DEMs) were detected in the anagen vs. catagen (AvC) comparison. 4092 DEGs, 1540 DELs, 356 DECs and 141 DEMs were detected in the anagen vs. telogen (AvT) comparison. 2290 DEGs, 779 DELs, 249 DECs and 92 DEMs were detected in the catagen vs. telogen (CvT) comparison. DEGs were primarily enriched in GO items including plasma membrane, integral component of plasma membrane and extracellular space. KEGG enrichment analysis revealed that DEGs were mainly enriched in PI3K-Akt signaling pathway, cell cycle and Wnt signaling pathway (p < 0.05). KEGG analysis showed trans-acting genes of DELs were significantly enriched in Hippo signaling pathway, PI3K-Akt signaling pathway and Melanogenesis. Target genes of DEMs were mainly enriched in MAPK signaling pathway, Wnt signaling pathway, ECM-receptor interaction and Signaling pathways regulating pluripotency of stem cells. Based on the ceRNA mechanism, lncRNA/circRNA-miRNA-mRNA networks were constructed involving 9 DECs, 437 DELs, 50 DEMs and 416 DEGs.ConclusionsTotally, this study provides comprehensive insights into the expression patterns of protein-coding genes and non-coding transcripts throughout the HF cycle, and enhancing the understanding of the regulatory mechanisms underlying mammalian hair fiber development.

Secretome and Proteome of Extracellular Vesicles Provide Protein Markers of Lung and Colorectal Cancer

Colorectal cancer (CRC) and lung cancer (LC) are leading causes of cancer-related mortality, highlighting the need for minimally invasive diagnostic, prognostic, and predictive markers for these cancers. Proteins secreted by a tumor into the extracellular space directly, known as the tumor secretome, as well as proteins in the extra-cellular vesicles (EVs), represent an attractive source of biomarkers for CRC and LC. We performed proteomic analyses on secretome and EV samples from LC (A549, NCI-H23, NCI-H460) and CRC (Caco2, HCT116, HT-29) cell lines and targeted mass spectrometry on EVs from plasma samples of 20 patients with CRC and 19 healthy controls. A total of 782 proteins were identified across the CRC and LC secretome and EV samples. Of these, 22 and 44 protein markers were significantly elevated in the CRC and LC samples, respectively. Functional annotation revealed enrichment in proteins linked to metastasis and tumor progression for both cancer types. In EVs isolated from the plasma of patients with CRC, ITGB3, HSPA8, TUBA4A, and TLN1 were reduced, whereas FN1, SERPINA1, and CST3 were elevated, compared to healthy controls. These findings support the development of minimally invasive liquid biopsy methods for the detection, prognosis, and treatment monitoring of LC and CRC.

Extracellular calreticulin regulates fibrogenic and immunogenic properties of hepatic stellate cells.

Liver fibrosis is a persistent damage repair response triggered by various etiological factors, resulting in an excessive accumulation of extracellular matrix (ECM). Activated hepatic stellate cells (HpSCs) are the primary source of ECM proteins. Therefore, specifically targeting HpSCs has become a crucial approach for treating liver fibrosis. Calreticulin (CRT) is a molecular chaperone mainly located in the endoplasmic reticulum (ER), regulating protein folding and calcium homeostasis. Recently, CRT has gained much attention for its role outside the ER, particularly at the cell surface and extracellular space, acting as an immunomodulatory protein. The current study investigates the role of extracellular CRT in hepatic injury and its effects on HpSCs. Elevated levels of circulating CRT were observed in mouse models of liver injury, suggesting that hepatic injury may trigger CRT release. Extracellular CRT was found to moderately inhibit HpSC viability and induce morphological changes. Additionally, CRT treatment led to a decrease in α-smooth muscle actin and an upregulation of matrix metalloproteinase-2 and -9, indicating a potential fibrolytic effect. Immunomodulatory activities of CRT were also noted, as it increased cytokine expression in both macrophages and HpSCs. These effects were partially mediated through low-density lipoprotein receptor-related protein 1 (LRP1), as evidenced by altered cytokine expression upon co-treatment with a known LRP1 ligand receptor-associated protein (RAP). Overall, this study elucidates the complex role of extracellular CRT in liver injury and its potential impact on HpSC behavior and immune responses.

A new weapon: the application of tumor vaccines based on extracellular exosomal heat shock proteins in immunotherapy

Despite the significant advancements in cancer research, innovative approaches are still needed to reduce tumor incidence, progression, and dissemination, as well as for prolonging patient survival. Currently, the development of cancer vaccines is gaining attention as a novel preventative and therapeutic strategy. Although the concept of cancer vaccination is not new, a limited number of vaccines have received approval for tumor therapy. Heat shock protein (HSP)-based vaccination represents a promising strategy that harnesses specific tumor antigens to activate immune responses. Exosomes (Exs) are highly heterogeneous bilayer vesicles capable of transporting various types of molecules through extracellular space. Compared with conventional anticancer drugs, exosomes exhibit low toxicity and good biocompatibility, and they can stimulate the immune system either directly or indirectly. Ex-based vaccines may elicit an antitumor immune response that generates memory cells capable of recognizing cancer antigens, thereby inhibiting disease progression. This paper reviews the potential applications of HSPs and exosomes in the prevention and treatment of solid tumors. Finally, we discuss the advantages of the extracellular exosomal heat shock protein (HSP-Ex1) vaccine and future research directions aimed at optimizing heat shock protein-based cancer immunotherapy strategies.

Genome-Wide Identification and Analysis of Carbohydrate-Binding Modules in Colletotrichum graminicola

Colletotrichum graminicola is the causative agent of both maize stem rot and leaf blight, which are among the most damaging diseases affecting maize. Carbohydrate-binding modules (CBMs) are protein domains that lack catalytic activity and are commonly found alongside carbohydrate-hydrolyzing enzymes in fungi. A comprehensive examination of the C. graminicola TZ-3 genome resulted in the identification of 83 C. graminicola CBM (CgCBM) genes, which are characterized by distinct gene structures and protein motifs. Subcellular localization analysis revealed that the majority of CgCBM proteins were localized in the extracellular space. Investigation of the promoter regions of CgCBM genes uncovered a variety of responsive elements associated with plant hormones, including abscisic acid and methyl jasmonate response elements, as well as various stress-related response elements for drought, cold, defense, and other stress factors. Gene ontology analysis identified the primary functions of CgCBM genes as being linked to polysaccharide metabolism processes. Furthermore, the 83 CgCBM genes exhibited varying responses at different time points during C. graminicola infection, indicating their contribution to the fungus–maize interaction and their potential roles in the fungal pathogenic process. This study provides essential insights into CgCBMs, establishing a crucial foundation for further exploration of their functions in the mechanisms of fungal pathogenicity.

Transcriptome analysis provides insights into the role of TLP16 in Musa acuminata Resistance to Fusarium oxysporum f. sp. cubense wilt

BackgroundThaumatin-like proteins (TLPs) are crucial pathogenesis-related proteins that significantly contribute to plant defense rection. Fusarium oxysporum f. sp. cubense (Foc) causes Fusarium wilt of bananas, a serious threat to global production. However, the role of TLPs in disease resistance remains unclear.ResultsThis study identified 49 TLP genes in banana, predominantly localized in the extracellular space, and distributed across 11 chromosomes. The ancestor–descendant relationship was explained, six genes remained remarkably conserved across species could represent the ancestral genes of the TLP gene family. Promoter regions, transcriptome and qRT-PCR analysis suggested that MaTLP16 might be involved in disease resistance. Furthermore, transcriptional silencing of MaTLP16 resulted in more severe leaf damage compared to the control, indicating that MaTLP16 is an important Foc resistance-related gene.ConclusionThis study conducted a comprehensive genome-wide identification and systematic analysis of the TLP gene family in bananas. Our findings establish a foundation for further functional studies of MaTLP genes and highlight MaTLP16 as a strong candidate for use in breeding programs aimed at enhancing resistance to Musa diseases.

Nanoparticles containing intracellular proteins modulate neutrophil functional and phenotypic heterogeneity

Neutrophils are rapidly recruited to sites of infection, injury, or to immune complexes. Upon arrival, they initiate degranulation, release reactive oxygen species (ROS), and/or nuclear extracellular traps (NETs) to eliminate invading microorganisms, clear debris, or remove abnormal immunoglobulins. While these processes ideally trigger healing and a return to balance, overshooting neutrophil function can lead to life-threatening infections such as sepsis or persistent inflammation observed in various autoimmune diseases. However, recent evidence highlights a phenotypic and functional heterogeneity of neutrophils that extends well beyond their traditional - potentially harmful- role as first responders. For example, neutrophils regulate ongoing inflammation by modulating macrophage function through efferocytosis, T cell responses by antigen presentation and the release of cytokines. The factors that induce neutrophil differentiation into activating or regulatory phenotypes remain poorly defined. Here, we hypothesize that intracellular components that have been released into the extracellular space could contribute to the phenotypic heterogeneity of neutrophils. To find out, we used nanoparticles composed of intracellular proteins (cell-derived nanoparticles, CDNPs) and analyzed their effects on cultured murine bone marrow neutrophils (BMN). We observed that CDNPs activate BMN transiently with an increase in the expression of CD11b without triggering classical effector functions. Additionally, CDNPs induce the secretion of IL-10, shift PMA-induced cell death toward apoptosis, and increase the expression of CD80. Mechanistically, our findings indicate that 26% of BMNs ingest CDNPs. These BMNs preferentially express CD54+, fail to migrate toward CXCL12, exhibit diminished responses to LPS, and undergo apoptosis. These data identify CDNPs as biomaterials that modulate neutrophil behavior by fine-tuning the expression of CD11b and CD80.

A novel transgenic reporter of extracellular acidification in zebrafish elucidates skeletal muscle T-tubule pH regulation.

Disruption of extracellular pH and proton-sensing can profoundly impact cellular and protein functions, leading to developmental defects. To visualize changes in extracellular pH in the developing embryo, we generated a zebrafish transgenic line that ubiquitously expresses the ratiometric pH-sensitive fluorescent protein pHluorin2, tethered to the extracellular face of the plasma membrane using a glycosylphosphatidylinositol (GPI) anchor. Monitoring of pHluorin2 with ratiometric fluorescence revealed dynamic and discrete domains of extracellular acidification over the first 72 h of embryonic development. These included acidification of the notochord intercalations, transient acidification of the otic placode, and persistent acidification of the extracellular space of the myotome at distinctly different pH from that within the T-tubules. Knockdown of centronuclear myopathy genes Bin1b (OMIM: 255200) and MTM1 (OMIM: 310400), which disrupt T-tubule formation, also disrupted myotome acidification. In this study we visualize extracellular acidic microdomains in the tissues of whole live animals. This real-time reporter line for directly measuring changes in extracellular pH can be used to illuminate the role of extracellular pH in normal physiological development and disease states.

Extracellular ATP regulates phagocytic activity, mitochondrial respiration, and cytokine secretion of human astrocytic cells.

The two main glial cell types of the central nervous system (CNS), astrocytes and microglia, are responsible for neuroimmune homeostasis. Recent evidence indicates astrocytes can participate in removal of pathological structures by becoming phagocytic under conditions of neurodegenerative disease when microglia, the professional phagocytes, are impaired. We hypothesized that adenosine triphosphate (ATP), which acts as damage-associated molecular pattern (DAMP), when released at high concentrations into extracellular space, upregulates phagocytic activity of human astrocytes. This study is the first to measure changes in phagocytic activity and mitochondrial respiration of human astrocytic cells in response to extracellular ATP. We demonstrate that ATP-induced phagocytic activity of U118 MG astrocytic cells is accompanied by upregulated mitochondrial oxidative phosphorylation, which likely supports this energy-dependent process. Application of a selective antagonist A438079 provides evidence identifying astrocytic purinergic P2X7 receptor (P2X7R) as the potential regulator of their phagocytic function. We also report a rapid ATP-induced increase in intracellular calcium ([Ca2+]i), which could serve as regulator of both the phagocytic activity and mitochondrial metabolism, but this hypothesis will need to be tested in future studies. Since ATP upregulates interleukin (IL)-8 secretion by astrocytes but has no effect on their cytotoxicity towards neuronal cells, we conclude that extracellular ATP affects only specific functions of astrocytes. The selectivity of P2X7R-dependent regulation of astrocyte functions by extracellular ATP could allow targeting this receptor-ligand interaction to upregulate their phagocytic function. This could have beneficial outcomes in neurodegenerative disorders, such as Alzheimer's disease, that are characterized by reactive astrocytes and defective phagocytic processes.

Glioblastoma-derived migrasomes promote migration and invasion by releasing PAK4 and LAMA4

Almost all high-grade gliomas, particularly glioblastoma (GBM), are highly migratory and aggressive. Migrasomes are organelles produced by highly migratory cells capable of mediating intercellular communication. Thus, GBM cells may produce migrasomes during migration. However, it remains unclear whether migrasomes can influence GBM migration and invasion. In this study, we observed the presence and formation of migrasomes in GBM cells. We found that expression levels of key migrasome formation factor, tetraspanin 4 (TSPAN4), correlated positively with pathological grade and poor prognosis of GBM based on the databases and clinical samples analysis. Subsequently, we knocked down TSPAN4 and found that GBM cell migration and invasion were significantly inhibited due to the reduced formation of migrasomes. We further confirmed that migrasomes are enriched in extracellular matrix (ECM)-related proteins such as p21-activating kinase 4 (PAK4) and laminin alpha 4 (LAMA4). Our experimental results suggest that migrasomes promote GBM cells migration by releasing such proteins into the extracellular space. Overall, we identified migrasomes in GBM and the molecular mechanisms by which they regulate them, providing potential targets for treating GBM.

KIF9 Ameliorates Neuropathology and Cognitive Dysfunction by Promoting Macroautophagy in a Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is a prevalent neurodegenerative disorder affecting the elderly. The imbalance of protein production and degradation processes leads to the accumulation of misfolded and abnormally aggregated amyloid-beta (Aβ) in the extracellular space and forms senile plaques, which constitute one of the most critical pathological hallmarks of AD. KIF9, a member of the kinesin protein superfamily, mediates the anterograde transport of intracellular cargo along microtubules. However, the exact role of KIF9 in AD pathogenesis remains largely elusive. In this study, we reported that the expression of kinesin family member 9 (KIF9) in the hippocampus of APP23/PS45 double-transgenic AD model mice declined in an age-dependent manner, concurrent with macroautophagy dysfunction. Furthermore, we found that KIF9 mediated the transport of lysosomes through kinesin light chain 1 (KLC1), thereby participating in the degradation of amyloidogenic pathway-related proteins of Aβ precursor protein (APP) in AD model cells through promoting the macroautophagy pathway. Importantly, genetic upregulation of KIF9 via adeno-associated virus (AAV) diminished Aβ deposition and alleviated cognitive impairments in AD model mice by enhancing macroautophagy function. Collectively, our findings underscore the ability of KIF9 to promote macroautophagy through KLC1-mediated anterograde transport of lysosomes, effectively ameliorating cognitive dysfunction in AD model mice. These discoveries suggest that KIF9 may represent a novel therapeutic target for the treatment of AD.

Detection of Deaths Caused by Hyperkalemia.

Under normal conditions, potassium is predominantly found within cells. The concentration gradient of sodium and potassium ions between intracellular and extracellular spaces enables signal transmission through membrane depolarization. The disruption of this transcellular process leads to elevated potassium ion levels in the extracellular space, and thus in the blood, a condition known as hyperkalemia. Clinically, hyperkalemia may present as cardiac arrhythmias, muscle weakness, and palpitations. The post-mortem accumulation of potassium ions in various human tissues and organs, such as the heart, liver, kidneys, lungs, and vitreous body, particularly in cases of overdose, has been an area of research interest for years. Unfortunately, deaths caused by hyperkalemia are difficult to identify due to their non-specific symptoms and are often misinterpreted as cardiovascular-related. Furthermore, most potassium ion concentration tests developed in recent years are non-specific, have limitations, or are based on outdated techniques. Consequently, alternative methods, such as histopathological tissue analysis, potassium concentration assessment in the vitreous body, and aldosterone level measurement, show promise for improving the post-mortem detection of exogenous hyperkalemia.

Propagation of neuronal micronuclei regulates microglial characteristics.

Microglia-resident immune cells in the central nervous system-undergo morphological and functional changes in response to signals from the local environment and mature into various homeostatic states. However, niche signals underlying microglial differentiation and maturation remain unknown. Here, we show that neuronal micronuclei (MN) transfer to microglia, which is followed by changing microglial characteristics during the postnatal period. Neurons passing through a dense region of the developing neocortex give rise to MN and release them into the extracellular space, before being incorporated into microglia and inducing morphological changes. Two-photon imaging analyses have revealed that microglia incorporating MN tend to slowly retract their processes. Loss of the cGAS gene alleviates effects on micronucleus-dependent morphological changes. Neuronal MN-harboring microglia also exhibit unique transcriptome signatures. These results demonstrate that neuronal MN serve as niche signals that transform microglia, and provide a potential mechanism for regulation of microglial characteristics in the early postnatal neocortex.

Decoding the MMP14 Integrin Link: Key Player in the Secretome Landscape.

Rapid progress has been made in the exciting field of secretome research in health and disease. The tumor secretome, which is a significant proportion of the tumor proteome, is secreted into the extracellular space to promote intercellular communication and thus tumor progression. Among the many molecules of the secretome, integrins and matrix metalloproteinase 14 (MMP14) stand out as the interplay of adhesion and proteolysis drives invasion. Integrins serve as mechanosensors that mediate the contact of cells with the scaffold of the extracellular matrix and are significantly involved in the precise positioning and activity control of the membrane-bound collagenase MMP14. As a secretome proteinase, MMP14 influences and modifies the secretome itself. While integrins and MT-MMPs are membrane bound, but can be released and are therefore border crossers between the cell surface and the secretome, the extracellular matrix is not constitutively cell-bound, but its binding to integrins and other cell receptors is a stringently regulated process. To understand the mutual interactions in detail, we first summarize the structure and function of MMP14 and how it is regulated at the enzymatic and cellular level. In particular, the mutual interactions between integrins and MMP14 include the proteolytic cleavage of integrins themselves by MMP14. We then review the biochemical, cell biological and physiological effects of MMP14 on the composition and associated functions in the tumor secretome when either bound to the cell membrane, or located on extracellular microvesicles, or as a proteolytically shed non-membrane-bound ectodomain. Novel methods of proteomics, including the analysis of extravesicular vesicles, and new methods for the quantification of MMP14 will provide new research and diagnostic tools. The proteolytic modification of the tumor secretome, especially by MMP14, may bring an additional aspect to tumor secretome studies and will have an impact on the diagnosis and most likely also on the therapy of cancer patients.