More than 75% of traumatic brain injuries (TBIs) are mild (mTBI) and military service members often experience repeated combat-related mTBI. The chronic comorbidities concomitant with repetitive mTBI (rmTBI) include depression, post-traumatic stress disorder or neurological dysfunction. This study sought to determine a long noncoding RNA (lncRNA) expression signature in serum samples that correlated with rmTBI years after the incidences. Serum samples were obtained from Long-Term Impact of Military-Relevant Brain-Injury Consortium Chronic Effects of Neurotrauma Consortium (LIMBIC CENC) repository, from participants unexposed to TBI or who had rmTBI. Four lncRNAs were identified as consistently present in all samples, as detected via droplet digital PCR and packaged in exosomes enriched for CNS origin. The results, using qPCR, demonstrated that the lncRNA VLDLR-AS1 levels were significantly lower among individuals with rmTBI compared to those with no lifetime TBI. ROC analysis determined an AUC of 0.74 (95% CI: 0.6124 to 0.8741; p = 0.0012). The optimal cutoff for VLDLR-AS1 was ≤153.8 ng. A secondary analysis of clinical data from LIMBIC CENC was conducted to evaluate the psychological symptom burden, and the results show that lncRNAs VLDLR-AS1 and MALAT1 are correlated with symptoms of depression. In conclusion, lncRNA VLDLR-AS1 may serve as a blood biomarker for identifying chronic rmTBI and depression in patients.

Background: Traumatic brain injury (TBI) occurs in individuals of all ages, predominantly during sports, accidents, and in active military service members. Chronic consequences of TBI include declined cognitive and motor function, dementia, and emotional distress. Small extracellular vesicles (sEVs), previously referred to as exosomes, are nano-sized lipid vesicles that play a role in intercellular communication. Our prior research established the efficacy of sEVs derived from human adipose stem cells (hASC sEVs) in accelerating the healing of brain injuries. The hASC sEVs are a biologic therapeutic and need to be stored at −20 °C or −80 °C. This limits their use in translating to everyday use in clinics or their inclusion in first-aid kits for application immediately after injury. To address this, here we demonstrate that hASC sEVs can be stored at room temperature (RT) for two months post lyophilization. Methods: A transmission electron microscope (TEM) and nanoparticle tracking analysis (NTA) were used to validate the morphology of lyophilized RT sEVs. Using in vitro models of neuronal injury mimicking physical injury, inflammation, and oxidative stress, we demonstrate that lyophilized RT hASC sEVs are viable and promote the healing of neuronal injuries. Results: The lyophilized sEVs maintain their purity, size, and morphology upon rehydration. Lyophilized, RT stored sEVs showed better efficacy after two months compared with −80 °C stored sEVs. Conclusions: RT storage of lyophilized hASC sEVs maintains their efficacy to accelerate the healing of injuries in neuronal cells. This will advance the use of hASC sEVs, bringing them closer to use in clinics, home first-aid kits, and on battlefields by active service members.

Wound healing progresses through four phases: hemostasis, inflammation, proliferation, and remodeling. Wounds may become chronic if this process is disrupted. The use of small extracellular vesicle (sEV; EVs < 200 nm) exosomes (exo; ~40–120 nm) derived from human adipose stem cells (hASCs) as a treatment for wounds is well studied. The cargo of these exosomes is of great interest as this accelerates wound healing. Our previous studies identified lncRNAs GAS5 and MALAT1 as packaged and enriched in hASC exosomes. In this study, we use a rat model to examine the effects on wound healing when hASC exosomes are depleted of GAS5 and MALAT1. Rats were wounded and wounds were treated with 100 μg hASC_exo or hASC_exo-G-M every 2 days for 1 week. qPCR was completed to evaluate the molecular effects of depletion of GAS5 and MALAT1 from hASC_exo. RNAseq was performed on wound tissue to evaluate the molecular mechanisms changed by hASC_exo-G-M in wound healing. While hASC_exo-G-M significantly improved wound healing rate compared to control wounds, healing occurred slower than in wounds treated with hASC_exo that were not depleted of GAS5 and MALAT1. Overall, this study reveals that molecular functions associated with healing are reduced in the absence of GAS5 and MALAT1, highlighting the importance of these lncRNAs.

Traumatic brain injury (TBI) is a major cause of long-term neurological impairment, with aging amplifying vulnerability and worsening recovery. Older individuals face greater cognitive and motor deficits post-TBI and respond less effectively to treatments, as both aging and TBI independently elevate neuroinflammation and cognitive decline. This study evaluated the therapeutic effects of human adipose-derived stem cell small extracellular vesicles (hASC-sEVs) on neurological recovery and neuroinflammation in a mouse model of TBI. Male C57BL/6 mice (3, 15, and 20 months old) underwent controlled cortical impact (CCI) and received intranasal hASC-sEVs 48 h post-injury; control groups received PBS. A dose–response study at 7 days post injury (dpi) identified 20 µg as the optimal therapeutic dose, improving motor function, reducing neuroinflammation, and enhancing neurogenesis. This was followed by a 30-dpi study assessing cognitive function, neuroinflammation, neurogenesis, and proteomic changes in microglia and astrocytes via mass spectrometry. hASC-sEV treatment significantly improved behavioral outcomes and reduced neuroinflammatory markers (GFAP, IBA-1, and MHC-II), with reduced efficacy observed in older mice. Proteomics revealed that hASC-sEVs reduce inflammatory proteins (TNF-α, IL-1β, IFNG, CCL2) and modulated mitochondrial dysfunction and reactive oxygen species. These results highlight hASC-sEVs as a promising cell-free therapy for improving TBI outcomes, especially in aging populations.

Type 2 diabetes mellitus is a metabolic disorder defined by systemic insulin resistance. Insulin resistance in adipocytes, an important regulator of glucose metabolism, results in impaired glucose uptake. The trafficking protein, sortilin, regulates major glucose transporter 4 (Glut4) movement, thereby promoting glucose uptake in adipocytes. Here, we demonstrate the presence of an alternatively spliced sortilin variant (Sort^17b), whose levels increase with insulin resistance in mouse 3T3L1 adipocytes. Using a splicing minigene, we show that inclusion of alternative exon 17b results in the expression of Sort^17b splice variant. Bioinformatic analysis indicated a novel intrinsic disorder region (IDR) encoded by exon 17b of Sort^17b. Root mean square deviation (RMSD) and root mean square fluctuation (RMSF) measurements using molecular dynamics demonstrated increased flexibility of the protein backbone within the IDR. Using protein–protein docking and co-immunoprecipitation assays, we show robust binding of Glut4 to Sort^17b. Further, results demonstrate that over-expression of Sort^17b correlates with reduced Glut4 translocation and decreased glucose uptake in adipocytes. The study demonstrates that insulin resistance in 3T3L1 adipocytes promotes expression of a novel sortilin splice variant with thus far unknown implications in glucose metabolism. This knowledge may be used to develop therapeutics targeting sortilin variants in the management of type 2 diabetes and metabolic syndrome.

In the United States, an estimated 38 million individuals (10% of the population) have type 2 diabetes mellitus (T2D), while approximately 97.6 million adults (38%) have prediabetes. Long noncoding RNAs (lncRNAs) are critical regulators of gene expression and metabolism. We were the first to demonstrate that lncRNA Growth Arrest-Specific Transcript 5 (GAS5 (human)/gas5 (mouse)) is decreased in the serum of T2D patients and established GAS5 as a biomarker for T2D diagnosis and onset prediction, now validated by other groups. We further demonstrated that GAS5 depletion impaired glucose uptake, decreased insulin receptor levels, and inhibited insulin signaling in human adipocytes, highlighting its potential as a therapeutic target in T2D. To address this, we developed NPC86, a small-molecule compound that stabilizes GAS5 by disrupting its interaction with UPF-1, an RNA helicase involved in nonsense-mediated decay (NMD) that regulates RNA stability. NPC86 increased GAS5 and insulin receptor (IR) levels, enhanced insulin signaling, and improved glucose uptake in vitro. In this study, we tested the efficacy of NPC86 in vivo in a diet-induced obese diabetic (DIOD) mouse model, and NPC86 treatment elevated gas5 levels, improved glucose tolerance, and enhanced insulin sensitivity, with no observed toxicity or weight changes. A transcriptomics analysis of adipose tissue revealed the upregulation of insulin signaling and metabolic pathways, including oxidative phosphorylation and glycolysis, while inflammatory pathways were downregulated. These findings highlight NPC86’s therapeutic potential in T2D.