LAPTM4B counteracts ferroptosis via suppressing the ubiquitin-proteasome degradation of SLC7A11 in non-small cell lung cancer.

Non-small cell lung cancer (NSCLC) is a leading cause of cancer-related deaths worldwide, necessitating the identification of novel therapeutic targets. Lysosome Associated Protein Transmembrane 4B (LAPTM4B) is involved in biological processes critical to cancer progression, such as regulation of solute carrier transporter proteins and metabolic pathways, including mTORC1. However, the metabolic processes governed by LAPTM4B and its role in oncogenesis remain unknown. In this study, we conducted unbiased metabolomic screens to uncover the metabolic landscape regulated by LAPTM4B. We observed common metabolic changes in several knockout cell models suggesting of a role for LAPTM4B in suppressing ferroptosis. Through a series of cell-based assays and animal experiments, we demonstrate that LAPTM4B protects tumor cells from erastin-induced ferroptosis both in vitro and in vivo. Mechanistically, LAPTM4B suppresses ferroptosis by inhibiting NEDD4L/ZRANB1 mediated ubiquitination and subsequent proteasomal degradation of the cystine-glutamate antiporter SLC7A11. Furthermore, metabolomic profiling of cancer cells revealed that LAPTM4B knockout leads to a significant enrichment of ferroptosis and associated metabolic alterations. By integrating results from cellular assays, patient tissue samples, an animal model, and cancer databases, this study highlights the clinical relevance of the LAPTM4B-SLC7A11-ferroptosis signaling axis in NSCLC progression and identifies it as a potential target for the development of cancer therapeutics.

VAMP8 suppresses the metastasis via DDX5/ß-catenin signal pathway in osteosarcoma.

Osteosarcoma is a highly metastatic malignant bone tumor, necessitating the development of new treatments to target its metastasis. Recent studies have revealed the significance of VAMP8 in regulating various signaling pathways in various types of cancer. However, the specific functional role of VAMP8 in osteosarcoma progression remains unclear. In this study, we observed a significant downregulation of VAMP8 in osteosarcoma cells and tissues. Low levels of VAMP8 in osteosarcoma tissues were associated with patients' poor prognosis. VAMP8 inhibited the migration and invasion capability of osteosarcoma cells. Mechanically, we identified DDX5 as a novel interacting partner of VAMP8, and the conjunction of VAMP8 and DDX5 promoted the degradation of DDX5 via the ubiquitin-proteasome system. Moreover, reduced levels of DDX5 led to the downregulation of ß-catenin, thereby suppressing the epithelial-mesenchymal transition (EMT). Additionally, VAMP8 promoted autophagy flux, which may contribute to the suppression of osteosarcoma metastasis. In conclusion, our study anticipated that VAMP8 inhibits osteosarcoma metastasis by promoting the proteasomal degradation of DDX5, consequently inhibiting WNT/ß-catenin signaling and EMT. Dysregulation of autophagy by VAMP8 is also implicated as a potential mechanism. These findings provide new insights into the biological nature driving osteosarcoma metastasis and highlight the modulation of VAMP8 as a potential therapeutic strategy for targeting osteosarcoma metastasis.

Drying Process of HPMC-Based Hard Capsules: Visual Experiment and Mathematical Modeling.

Using plant-based polysaccharide gels to produce hard capsules is a novel application of this technology in the medicinal field, which has garnered significant attention. However, the current manufacturing technology, particularly the drying process, limits its industrialization. The work herein employed an advanced measuring technique and a modified mathematical model to get more insight into the drying process of the capsule. Low field magnetic resonance imaging (LF-MRI) technique is adopted to reveal the distribution of moisture content in the capsule during drying. Furthermore, a modified mathematical model is developed by considering the dynamic variation of the effective moisture diffusivity (Deff) according to Fick's second law, which enables accurate prediction of the moisture content of the capsule with a prediction accuracy of ±15%. The predicted Deff ranges from 3 × 10-10 to 7 × 10-10 m2·s-1, which has an irregular variation with a time extension. Moreover, as temperature increases or relative humidity decreases, there is an increased acceleration of moisture diffusion. The work provides a fundamental understanding of the drying process of the plant-based polysaccharide gel, which is crucial for enhancing the industrial preparation of the HPMC-based hard capsules.

Enhancing Pullulan Soft Capsules with a Mixture of Glycerol and Sorbitol Plasticizers: A Multi-Dimensional Study.

The plasticizer is crucial in the plant-based soft capsule. However, meeting the quality requirements of these capsules with a single plasticizer is challenging. To address this issue, this study first investigated the impact of a plasticizer mixture containing sorbitol and glycerol in varying mass ratios and the performance of the pullulan soft film and capsule. The multiscale analysis demonstrates that the plasticizer mixture exhibits superior effectiveness in enhancing the performance of the pullulan film/capsule compared to a single plasticizer. Furthermore, thermogravimetric analysis, Fourier transform infrared spectroscopy, X-ray diffraction, and scanning electron microscopy indicate that the plasticizer mixture enhances the compatibility and thermal stability of the pullulan films without altering their chemical composition. Among the different mass ratios examined, a 15:15 ratio of sorbitol to glycerol (S/G) is identified as the most optimal, leading to superior physicochemical properties and meeting the requirements for brittleness and disintegration time set by the Chinese Pharmacopoeia. This study provides significant insights into the effect of the plasticizer mixture on the performance of pullulan soft capsules and offers a promising application formula for future use.

miR-137-LAPTM4B regulates cytoskeleton organization and cancer metastasis via the RhoA-LIMK-Cofilin pathway in osteosarcoma.

Osteosarcoma (OS) is a rare malignant bone tumor but is one leading cause of cancer mortality in childhood and adolescence. Cancer metastasis accounts for the primary reason for treatment failure in OS patients. The dynamic organization of the cytoskeleton is fundamental for cell motility, migration, and cancer metastasis. Lysosome Associated Protein Transmembrane 4B (LAPTM4B) is an oncogene participating in various biological progress central to cancer biogenesis. However, the potential roles of LAPTM4B in OS and the related mechanisms remain unknown. Here, we established the elevated LAPTM4B expression in OS, and it is essential in regulating stress fiber organization through RhoA-LIMK-cofilin signaling pathway. In terms of mechanism, our data revealed that LAPTM4B promotes RhoA protein stability by suppressing the ubiquitin-mediated proteasome degradation pathway. Moreover, our data show that miR-137, rather than gene copy number and methylation status, contributes to the upregulation of LAPTM4B in OS. We report that miR-137 is capable of regulating stress fiber arrangement, OS cell migration, and metastasis via targeting LAPTM4B. Combining results from cells, patients' tissue samples, the animal model, and cancer databases, this study further suggests that the miR-137-LAPTM4B axis represents a clinically relevant pathway in OS progression and a viable target for novel therapeutics.

LAPTM4B-35 promotes cancer cell migration via stimulating integrin beta1 recycling and focal adhesion dynamics.

Metastasis is the main cause of cancer patients' death despite tremendous efforts invested in developing the related molecular mechanisms. During cancer cell migration, cells undergo dynamic regulation of filopodia, focal adhesion, and endosome trafficking. Cdc42 is imperative for maintaining cell morphology and filopodia, regulating cell movement. Integrin beta1 activates on the endosome, the majority of which distributes itself on the plasma membrane, indicating that endocytic trafficking is essential for this activity. In cancers, high expression of lysosome-associated protein transmembrane 4B (LAPTM4B) is associated with poor prognosis. LAPTM4B-35 has been reported as displaying plasma membrane distribution and being associated with cancer cell migration. However, the detailed mechanism of its isoform-specific distribution and whether it relates to cell migration remain unknown. Here, we first report and quantify the filopodia localization of LAPTM4B-35: mechanically, that specific interaction with Cdc42 promoted its localization to the filopodia. Furthermore, our data show that LAPTM4B-35 stabilized filopodia and regulated integrin beta1 recycling via interaction and cotrafficking on the endosome. In our zebrafish xenograft model, LAPTM4B-35 stimulated the formation and dynamics of focal adhesion, further promoting cancer cell dissemination, whereas in skin cancer patients, LAPTM4B level correlated with poor prognosis. In short, this study establishes an insight into the mechanism of LAPTM4B-35 filopodia distribution, as well as into its biological effects and its clinical significance, providing a novel target for cancer therapeutics development.

Water Treadmill Training Ameliorates Neurite Outgrowth Inhibition Associated with NGR/RhoA/ROCK by Inhibiting Astrocyte Activation following Spinal Cord Injury.

Spinal cord injury (SCI) often results in damage to or degeneration of axons. Crosstalk between astrocytes and neurons plays a pivotal role in neurite outgrowth following SCI. Rehabilitative training is a recognized method for the treatment of SCI, but the specific mechanism underlying its effect on axonal outgrowth in the central nervous system (CNS) has not yet been determined. A total of 190 adult male SD rats weighing 200-250 g were randomly divided into eight groups for use as animal models of SCI. Rats were subjected to water treadmill training (TT) for 7 or 14 d. The Basso-Beattie-Bresnahan (BBB) motor function scale, hematoxylin-eosin (HE) staining, Nissl staining, Western blotting, and immunofluorescence were used to measure tissue morphology and the degree of neurological deficit and to determine quantitative expression and accurate localization of the corresponding proteins. We found that TT decreased tissue structure damage and improved functional recovery. TT also promoted the regeneration of neurons and reduced SCI-induced apoptosis SCI around the lesion, as well as significantly increasing the expression of GAP43 and NF200 after SCI. In addition, TT significantly inhibited the injury-induced increase in the expression of proinflammatory factors. Moreover, TT reduced the activation of astrocytes and microglia, accompanied by the reduced expression of C3d and increased expression of S100A10. Finally, TT effectively reduced the level of chondroitin sulfate proteoglycan (CSPG) surrounding the lesion and inhibited the NGR/RhoA/ROCK signaling pathway in neurons after SCI. Overall, we found that TT played a novel role in recovery from SCI by promoting axonal outgrowth associated with NGR/RhoA/ROCK signaling by inhibiting astrocyte activation after SCI.

Experimental and Numerical Study on Friction and Wear Performance of Hot Extrusion Die Materials.

For the aluminium alloys produced by the hot extrusion process, the profile is shaped according to the bearing at the exit of the extrusion die. The tribological process has significant effects on the die service life, profile dimensional tolerances, and profile surface finish. Recently, new technologies have been introduced to the hot extrusion die, such as cemented carbide insert die and surface coating. However, under hot extrusion working conditions, quantitative studies on their friction and wear performances are lacking. In this work, the friction and wear performances of three typical extrusion die materials, traditional hot tool steel (H13), cemented carbide (YG8), and chemical vapour deposition (CVD) coating, were studied. Macro and nano hardness tests, Pin-on-disk friction and wear tests, optical profiler and SEM observations, and experiments and simulations of hot extrusion were conducted. The results show that the coefficients of friction of CVD coatings and H13 hot work tool steel specimens were smaller under the hot extrusion condition than at room temperature. The wear mechanisms of H13, YG8, and CVD coatings at 500 °C are adhesion, abrasive, and fatigue, respectively. Moreover, the tribology results were validated by the extrusion experiments and the finite element analysis of hot extrusion. The conclusion of this manuscript is useful not only for the numerical simulation of the hot extrusion process but also for the surface finishing of the extrusion profile.

Electroacupuncture suppresses spinal nerve ligation-induced neuropathic pain via regulation of synaptic plasticity through upregulation of basic fibroblast growth factor expression.

BACKGROUND: Improving synaptic plasticity is a good way to alleviate neuropathic pain. Electroacupuncture (EA) is currently used worldwide to treat this disease, but its specific mechanisms of action need further investigation. Evidence has suggested that basic fibroblast growth factor (bFGF) plays an important role in promoting nerve regeneration and can promote the expression of vascular endothelial growth factor (VEGF). OBJECTIVE: In this study, we examined the effects of EA on synaptic plasticity and its underlying mechanism. METHODS: A spinal nerve ligation (SNL) rat model was established. NSC37204 (a specific inhibitor of bFGF) was used to determine the relationship between bFGF and putative EA-mediated improvements in synaptic plasticity. Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were assessed to evaluate hyperalgesia in rats with SNL. Tissue morphology was detected by hematoxylin-eosin (HE) and Nissl staining, while neural plasticity and its molecular mechanisms were examined by Western blotting, quantitative real-time polymerase chain reaction (qPCR), dual-label immunohistochemistry and transmission electron microscopy. RESULTS: We found that EA improved synaptic plasticity, consistent with higher levels of expression of bFGF and VEGF. Contrary to the beneficial effects of EA, NSC37204 promoted synaptic reconstruction. Furthermore, EA-induced improvements in the neurobehavioral state and improved synaptic plasticity were blocked by NSC37204, consistent with lower expression levels of bFGF and VEGF. CONCLUSION: These findings indicate that EA suppresses SNL-induced neuropathic pain by improving synaptic plasticity via upregulation of bFGF expression.

Prohibitin plays a role in the functional plasticity of macrophages.

Immunometabolism plays a crucial role in the activation and functional plasticity of immune cells, which in large determines a variety of health and disease states. Factors that integrate immunometabolism in immune cell signaling and functions are beginning to be identified. Previously, we have reported that two transgenic mouse models, Mito-Ob and mutant Mito-Ob (m-Mito-Ob), overexpressing a pleiotropic protein, prohibitin (PHB) or a mutant form of PHB (Tyr114Phe-PHB or m-PHB), respectively, developed distinct immunometabolic phenotypes. Specifically, the immune phenotype appears to be driven by the monocytic cell lineage. Based on immunophenotyping of their splenocytes, we focused our attention on macrophages and hypothesized that PHB may play a role in regulating the two functionally polarized states, M1 and M2. Here, we report that macrophage polarization to the M1 and M2 phenotypes did not alter PHB protein level, but overexpression of PHB in macrophages differentially affected cytokine production in the two polarized states. Furthermore, we found that mutation of the Tyr114 phosphorylation site in PHB affects ERK and STAT6 signaling, arginase synthesis and activity, and mitochondrial respiration in macrophages indicating an important role of PHB in integrating cell signaling events with cell metabolism. In summary, we have discovered that PHB is a crucial regulator in the functional plasticity of macrophages. These initial studies expect to lay the foundation for future research into the relationship between cell signaling events pertaining to immunometabolism in immune cell functions, which are integral components of immune-related health and disease.

The Predictive Role of Systemic Inflammation Response Index (SIRI) in the Prognosis of Stroke Patients.

PURPOSE: Stroke is a disease associated with high mortality. Many inflammatory indicators such as neutrophil-to-lymphocyte ratio (NLR), platelet-to-lymphocyte ratio (PLR), lymphocyte to monocyte ratio (LMR) and red blood cell distribution width (RDW) have been documented to predict stroke prognosis, their predictive power is limited. A novel inflammatory indicator called systemic inflammatory response index (SIRI) has been advocated to have an essential role in the prognostic assessment of cancer and infectious diseases. In this study, we attempted to assess the prognosis of stroke by SIRI. Moreover, we compared SIRI with other clinical parameters, including NLR, PLR, LMR and RDW. METHODS: This was a retrospective cohort study. We obtained data of 2450 stroke patients from the Multiparametric Intelligent Monitoring in Intensive Care III database. We used the Cox proportional hazards models to evaluate the relationship between SIRI and all-cause mortality and sepsis. Receiver operating curve (ROC) analysis was used to assess the predictive power of SIRI compared to NLR, PLR, LMR and RDW for the prognosis of stroke. We collected data of 180 patients from the First Affiliated Hospital of Wenzhou Medical University, which used the Pearson's correlation coefficient to assess the relationship between SIRI and the National Institute of Health stroke scale (NIHSS). RESULTS: After adjusting multiple covariates, we found that SIRI was associated with all-cause mortality in stroke patients. Rising SIRI accompanied by rising mortality. Besides, ROC analysis showed that the area under the curve of SIRI was significantly greater than for NLR, PLR, LMR and RDW. Besides, Pearson's correlation test confirmed a significant positive correlation between SIRI and NIHSS. CONCLUSION: Elevated SIRI was associated with higher risk of mortality and sepsis and higher stroke severity. Therefore, SIRI is a promising low-grade inflammatory factor for predicting stroke prognosis that outperformed NLR, PLR, LMR, and RDW in predictive power.

Ischemic stroke induces cardiac dysfunction and alters transcriptome profile in mice.

BACKGROUND: Stroke can induce cardiac dysfunction in the absence of primary cardiac disease; however, the mechanisms underlying the interaction between the neurological deficits and the heart are poorly understood. The objective of this study was to investigate the effects of stroke on cardiac function and to identify the transcriptome characteristics of the heart. RESULTS: Stroke significantly decreased heart weight/tibia length ratio and cardiomyocyte cross-sectional areas and increased atrogin-1 and the E3 ubiquitin ligase MuRF-1, indicating myocardial atrophy in MCAO-induced mouse hearts. RNA sequencing of mRNA revealed 383 differentially expressed genes (DEGs) in MCAO myocardium, of which 221 were downregulated and 162 upregulated. Grouping of DEGs based on biological function and quantitative PCR validation indicated that suppressed immune response and collagen synthesis and altered activity of oxidoreductase, peptidase, and endopeptidase may be involved in MCAO-induced cardiomyopathy. The DEGs were mainly distributed in the membrane or extracellular region of cardiomyocytes and acted as potential mediators of stroke-induced cardiac dysregulation involved in cardiac atrophy. CONCLUSION: Stroke induced a unique transcriptome response in the myocardium and resulted in immediate cardiac atrophy and dysfunction.

[Effect of electroacupuncture on angiopoietin-1 in spinal cord of rats with neuropathic pain].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on the behavior, histomorphology and the expression of angiopoietin-1 (Angpt-1) in rats with spinal nerve injury, so as to explore its mechanism on neuropathic pain. METHODS: Forty-five male SD rats were randomly divided into sham, model and EA groups (n=15 rats in each group). Spinal nerve ligation (SNL) of the L5 lumbar vertebra was performed to establish a rat model of neuropathic pain. The rats in the EA group were given EA at "Zusanli" (ST36) and "Kunlun" (BL60) of the operation side with continuous wave at a frequency of 2 Hz and an intensity of 1.5 mA once a day, 30 minutes each time for 7 days. The sham group only exposed L5 spinal nerves without ligation. Mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) were observed and recorded before modeling and on days 3,5,7,10,12 and 14 after modeling. L4-L6 segments of spinal cord were taken and the morphological changes of spinal dorsal horn were observed by HE staining. The changes of spinal dorsal horn nerve fiber structure were observed by silver plating staining. Angpt-1 expression was detected by Western blot and immunohistochemistry. RESULTS: Compared with the sham group, the model group had significant reductions in MWT and TWL at each time point (P<0.01); compared with the model group, the EA group had significant increases in MWT and TWL on days 10,12 and 14 after intervention (P<0.05, P<0.01). HE staining showed that in the model group, the spinal dorsal horn showed degeneration and necrosis of neurons, nuclear fixation and shrinkage, and loose surrounding tissues. The degree of tissue damage of the EA group was milder than that of the model group. The silver staining results showed the model group had obvious neuronal fibrillary tangles, while there were fewer neuronal fibrillary tangles in the EA group. Compared with the sham group, the Angpt-1 expression in the model group was significantly decreased (P<0.01), and compared with the model group, the EA group had a significant increase in the expression of Angpt-1 (P<0.01). CONCLUSION: EA can promote the recovery of nerve function in SNL rats by up-regulating Angpt-1 expression.

Electroacupuncture improves neuronal plasticity through the A2AR/cAMP/PKA signaling pathway in SNL rats.

Improvements in neuronal plasticity are considered to be conducive to recovery from neuropathic pain. Electroacupuncture (EA) is regarded as an effective rehabilitation method for neuropathic pain. However, the effects and potential mechanism associated with EA-induced repair of hyperesthesia are not fully understood. Evidence has suggested that the adenosine A2A receptor (A2AR) and the cyclic adenosine monophosphate (cAMP)/protein kinase A (PKA) pathway play an important role in improving neuropathic pain. Here, we examined the function of EA in promoting neuronal plasticity in spinal nerve ligation (SNL) rats. The A2AR antagonist SCH58261, A2AR agonist 2-p-(2-carboxyethyl)phenethylamino-50-N-ethylcarboxamido adenosine HCl (CGS21680) and A2AR siRNA were used to confirm the relationship between A2AR and the cAMP/PKA pathway as well as the effects of A2AR on EA-induced improvements in neurobehavioral state and neuronal plasticity. Mechanical withdrawal threshold (MWT), thermal withdrawal latency (TWL), HE staining, Western blotting, RT-PCR, immunofluorescence, enzyme-linked immunosorbent assay, Nissl staining, silver staining, Golgi-Cox staining and transmission electron microscopy were used to evaluate the changes in neurobehavioral performance, protein expression, neuronal structure and dendrites/synapses. The results showed that EA and CGS21680 improved the behavioral performance, neuronal structure and dendritic/synaptic morphology of SNL rats, consistent with higher expression levels of A2AR, cAMP and PKA. In contrast to the positive effects of EA, SCH58261 inhibited dendritic growth and promoted dendritic spine/synaptic remodeling. In addition, the EA-induced improvement in neuronal plasticity was inhibited by SCH58261 and A2AR siRNA, consistent with lower expression levels of A2AR, cAMP and PKA, and worse behavioral performance. These results indicate that EA suppresses SNL-induced neuropathic pain by improving neuronal plasticity via upregulating the A2AR/cAMP/PKA signaling pathway.

Electroacupuncture may alleviate neuropathic pain via suppressing P2X7R expression.

Neuropathic pain is a severe problem that is difficult to treat clinically. Reducing abnormal remodeling of dendritic spines/synapses and increasing the anti-inflammatory effects in the spinal cord dorsal horn are potential methods to treat this disease. Previous studies have reported that electroacupuncture (EA) could increase the pain threshold after peripheral nerve injury. However, the underlying mechanism is unclear. P2X7 receptors (P2X7R) mediate the activation of microglia and participate in the occurrence and development of neuropathic pain. We hypothesized that the effects of EA on relieving pain may be related to the downregulation of the P2X7R. Spinal nerve ligation (SNL) rats were used as a model in this experiment, and 2'(3')-O-(4-benzoyl)benzoyl ATP (BzATP) was used as a P2X7R agonist. We found that EA treatment decreased dendritic spine density, inhibited synaptic reconstruction and reduced inflammatory response, which is consistent with the decrease in P2X7R expression as well as the improved neurobehavioral performance. In contrast to the beneficial effects of EA, BzATP enhanced abnormal remodeling of dendritic spines/synapses and inflammation. Furthermore, the EA-mediated positive effects were reversed by BzATP, which is consistent with the increased P2X7R expression. These findings indicated that EA improves neuropathic pain by reducing abnormal dendritic spine/synaptic reconstruction and inflammation via suppressing P2X7R expression.

Electroacupuncture effects on the P2X4R pathway in microglia regulating the excitability of neurons in the substantia gelatinosa region of rats with spinal nerve ligation.

Electroacupuncture (EA) has been used to treat neuropathic pain induced by peripheral nerve injury (PNI) by applying an electrical current to acupoints with acupuncture needles. However, the mechanisms by which EA treats pain remain indistinct. High P2X4 receptor (P2X4R) expression levels demonstrate a notable increase in hyperactive microglia in the ipsilateral spinal dorsal horn following PNI. In order to demonstrate the possibility that EA analgesia is mediated in part by P2X4R in hyperactive microglia, the present study performed mechanical withdrawal threshold (MWT) and thermal withdrawal latency (TWL) tests in male Sprague­Dawley rats that had undergone spinal nerve ligation (SNL). The expression levels of spinal P2X4R were determined using reverse transcription­quantitative PCR, western blotting analysis and immunofluorescence staining. Furthermore, spontaneous excitatory postsynaptic currents (sEPSCs) were recorded using whole­cell patch clamp to demonstrate the effect of EA on synaptic transmission in rat spinal substantia gelatinosa (SG) neurons. The results of the present study demonstrated that EA increased the MWT and TWL and decreased overexpression of P2X4R in hyperactive microglia in SNL rats. Moreover, EA attenuated the frequency of sEPSCs in SG neurons in SNL rats. The results of the present study indicate that EA may mediate P2X4R in hyperactive spinal microglia to inhibit nociceptive transmission of SG neurons, thus relieving pain in SNL rats.

Water treadmill training protects the integrity of the blood-spinal cord barrier following SCI via the BDNF/TrkB-CREB signalling pathway.

Following spinal cord injury (SCI), destruction of the blood-spinal cord barrier (BSCB) leads to increased microvascular permeability and tissue oedema. The BSCB, formed by a dense network of tight junctions (TJs) and adhesion junctions (AJs) is considered a therapeutic target. Most studies have focused on the effect of drug therapy on the neurovascular system after SCI, ignoring the protection and functional recovery of the vascular system by exercise training. Previously, we indicated that water treadmill training (TT) has a protective effect on the BSCB after SCI, but the specific molecular mechanism of the effect of TT on BSCB is still not clear. In this study, we used a specific inhibitor of TrkB (ANA-12) to explore whether the BDNF/TrkB-CREB signalling pathway is involved in TT-mediated BSCB protection after SCI. A New York University (NYU) impactor was used to establish the SCI model. Rats in the SI (Sham + ANA-12), IM (SCI + ANA-12) and ITM (SCI + TT + ANA-12) groups were injected with ANA-12 (0.5 mg/kg) daily, and rats in TM (SCI + TT) and ITM (SCI + TT + ANA-12) groups were treated with water TT for 7 or 14 d. The degree of neurological deficit, water content, BSCB permeability, protein expression and ultrastructure of vascular endothelial cells were assessed by the Basso-Beattie-Bresnahan (BBB) motor rating scale, Evans blue (EB), Western blot (WB) experiments, immunofluorescence and transmission electron microscopy (TEM). Our results suggest that TT upregulates the BDNF/TrkB-CREB signalling pathway following SCI. The BDNF/TrkB-CREB signalling pathway is involved in the protection of the BSCB. Application of the inhibitor blocked the protective effect of TT on the BSCB. We concluded that TT ameliorated SCI-induced BSCB impairment by upregulating the BDNF/TrkB-CREB signalling pathways.

LAPTM4B controls the sphingolipid and ether lipid signature of small extracellular vesicles.

Lysosome Associated Protein Transmembrane 4B (LAPTM4B) is a four-membrane spanning ceramide interacting protein that regulates mTORC1 signaling. Here, we show that LAPTM4B is sorted into intraluminal vesicles (ILVs) of multivesicular endosomes (MVEs) and released in small extracellular vesicles (sEVs) into conditioned cell culture medium and human urine. Efficient sorting of LAPTM4B into ILV membranes depends on its third transmembrane domain containing a sphingolipid interaction motif (SLim). Unbiased lipidomic analysis reveals a strong enrichment of glycosphingolipids in sEVs secreted from LAPTM4B knockout cells and from cells expressing a SLim-deficient LAPTM4B mutant. The altered sphingolipid profile is accompanied by a distinct SLim-dependent co-modulation of ether lipid species. The changes in the lipid composition of sEVs derived from LAPTM4B knockout cells is reflected by an increased stability of membrane nanodomains of sEVs. These results identify LAPTM4B as a determinant of the glycosphingolipid profile and membrane properties of sEVs.