The Pathogenic Role of Anti-Granulocyte-Macrophage Colony-Stimulating Factor Autoantibodies in the Nocardiosis with the Central Nervous System Involvement.

PURPOSE: Anti-granulocyte-macrophage colony-stimulating factor autoantibodies (anti-GM-CSF Abs) are implicated in the pathogenesis of Cryptococcus gattii (C. gattii) infection and pulmonary alveolar proteinosis (PAP). Their presence has also been noted in nocardiosis cases, particularly those with disseminated disease. This study delineates a case series characterizing clinical features and specificity of anti-GM-CSF Abs in nocardiosis patients. METHODS: In this study, eight patients were recruited to determine the presence or absence of anti-GM-CSF Abs. In addition to the detailed description of the clinical course, we thoroughly investigated the autoantibodies regarding the characteristics, isotypes, subclasses, titers, and neutralizing capacities by utilizing the plasma samples from patients. RESULTS: Of eight patients, five tested positive for anti-GM-CSF Abs, all with central nervous system (CNS) involvement; patients negative for these antibodies did not develop CNS nocardiosis. Distinct from previously documented cases, none of our patients with anti-GM-CSF Abs exhibited PAP symptoms. The titer and neutralizing activity of anti-GM-CSF Abs in our cohort did not significantly deviate from those found in C. gattii cryptococcosis and PAP patients. Uniquely, one individual (Patient 3) showed a minimal titer and neutralizing action of anti-GM-CSF Abs, with no relation to disease severity. Moreover, IgM autoantibodies were notably present in all CNS nocardiosis cases investigated. CONCLUSION: The presence of anti-GM-CSF Abs suggests an intrinsic immunodeficiency predisposing individuals toward CNS nocardiosis. The presence of anti-GM-CSF Abs helps to elucidate vulnerability to CNS nocardiosis, even with low titer of autoantibodies. Consequently, systematic screening for anti-GM-CSF Abs should be considered a crucial diagnostic step for nocardiosis patients.

Translational Potential of Baicalein in Mitigating RSL3-Induced Ferroptosis in Fibroblasts: Implications for Therapeutic Interventions.

Background: Ferroptosis is an iron-driven cell-death mechanism that plays a central role in various diseases. Recent studies have suggested that baicalein inhibits ferroptosis, making it a promising therapeutic candidate. Materials and Methods: Fibroblast cultures were treated with different agents to determine the effects of baicalein on ferroptosis. Ferroptosis-related gene expression, lipid peroxidation, and post-treatment cellular structural changes were measured using real-time quantitative polymerase chain reaction, C11-BODIPY dye, and transmission electron microscopy, respectively. Results: Baicalein significantly inhibited rat sarcoma virus selective lethal 3-induced ferroptosis in fibroblasts. Moreover, in baicalein-treated groups, reduced ferroptosis-related gene expression, decreased lipid peroxidation, and maintained cell structure was observed when compared with those of the controls. Discussion: The ability of baicalein to counteract RSL3-induced ferroptosis underscores its potential protective effects, especially in diseases characterized by oxidative stress and iron overload in fibroblasts. Conclusion: Baicalein may serve as a potent therapeutic agent against conditions in which ferroptosis is harmful. The compound's efficacy in halting RSL3-triggered ferroptosis in fibroblasts paves the way for further in vivo experiments and clinical trials.

Cathelicidin Antimicrobial Peptide Acts as a Tumor Suppressor in Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is associated with high rates of metastasis and recurrence, and is one of the most common causes of cancer-associated death worldwide. This study examined the protein changes within circulating exosomes in patients with HCC against those in healthy people using isobaric tags for a relative or absolute quantitation (iTRAQ)-based quantitative proteomics analysis. The protein levels of von Willebrand factor (VWF), cathelicidin antimicrobial peptide (CAMP), and proteasome subunit beta type-2 (PSMB2) were altered in HCC. The increased levels of VWF and PSMB2 but decreased CAMP levels in the serum of patients with HCC were validated by enzyme-linked immunosorbent assays. The level of CAMP (the only cathelicidin found in humans) also decreased in the circulating exosomes and buffy coat of the HCC patients. The serum with reduced levels of CAMP protein in the HCC patients increased the cell proliferation of Huh-7 cells; this effect was reduced following the addition of CAMP protein. The depletion of CAMP proteins in the serum of healthy people enhances the cell proliferation of Huh-7 cells. In addition, supplementation with synthetic CAMP reduces cell proliferation in a dose-dependent manner and significantly delays G1-S transition in Huh-7 cells. This implies that CAMP may act as a tumor suppressor in HCC.

Identification and characterization of hADSC-derived exosome proteins from different isolation methods.

Exosomes are secreted into the extracellular space by most cell types and contain various molecular constituents, which play roles in many biological processes. Adipose-derived mesenchymal stem cells (ADSCs) can differentiate into a variety of cell types and secrete a series of paracrine factors through exosomes. ADSC-derived exosomes have shown diagnostic and therapeutic potential in many clinical diseases. The molecular components are critical for their mechanisms. Several methods have been developed for exosome purification, including ultracentrifugation, ultrafiltration, density gradient purification, size-based isolation, polymer precipitation and immuno-affinity purification. Thus, we employed four methods to isolate exosomes from the hADSC culture medium, including ultracentrifugation, size exclusion chromatography, ExoQuick-TC precipitation and ExoQuick-TC ULTRA isolation. Following exosome isolation, we performed quantitative proteomic analysis of the exosome proteins using isobaric tags for relative and absolute quantification (iTRAQ) labelling, combined with 2D-LC-MS/MS. There were 599 universal and 138 stably expressed proteins in hADSC-derived exosomes. We proved that these proteins were potential hADSC-derived exosomes markers, including CD109, CD166, HSPA4, TRAP1, RAB2A, RAB11B and RAB14. From the quantitative proteomic analysis, we demonstrated that hADSC-derived exosome protein expression varied, with lipopolysaccharide (LPS) treatment, in the different isolation methods. Pathway analysis and proliferation, migration and endothelial tube formation assays showed varying effects in cells stimulated with hADSC-derived exosomes from different isolation methods. Our study revealed that different isolation methods might introduce variations in the protein composition in exosomes, which reflects their effects on biological function. The pros and cons of these methods are important points to consider for downstream research applications.

Subpopulations of exosomes purified via different exosomal markers carry different microRNA contents.

The heterogeneity of exosome populations presents a great challenge to their study. The current study was designed to investigate the potential heterogeneity miRNA contents in circulating exosomes purified via different exosomal markers. In this study, exosomes from the serum of C57BL/6 mice after cecum ligation and perforation (CLP) or sham operation were isolated by precipitation using ExoQuick-TC and affinity purified with anti-Rab5b, anti-CD9, anti-CD31, and anti-CD44 antibodies using the Exo-Flow Exosome Capture kit to collect exosome subpopulations. RNA extracted from the exosomes isolated by ExoQuick-TC were profiled by next-generation sequencing (NGS). Real-time quantitative reverse transcription polymerase chain reaction (RT-qPCR) was also employed to determine the expression profiles of four representative exosomal miRNAs (mmu-miR-486-5p, mmu-miR-10a-5p, mmu-miR-143-3p, and mmu-miR-25-3p) selected from the NGS analysis. The results revealed that the expression patterns of these miRNAs in exosomes isolated by ExoQuick-TC as determined by RT-qPCR and NGS were similar, showing upregulation of mmu-miR-10a-5p and mmu-miR-143-3p but downregulation of mmu-miR-25-3p and mmu-miR-486-5p following CLP when compared to the levels in exosomes from sham control mice. However, their expression levels in the antibody-captured exosome subpopulations varied. The miRNAs in the exosomes captured by anti-Rab5b or anti-CD9 antibodies were more similar to those isolated by ExoQuick-TC than to those captured by anti-CD44 antibodies. However, there were no significant differences in these four miRNAs in CD31-captured exosomes. This study demonstrated that purification with different exosomal markers allows the collection of different exosome subpopulations with various miRNA contents. The results of this study demonstrate the heterogeneity of circulating exosomes and suggest the importance of stratifying exosome subpopulations when using circulating exosomes as biomarkers or investigating exosome function. In addition, this study also emphasized the necessity of using a consistent exosome marker across different samples as detecting biomarkers.

Exosomes Secreted by Adipose-Derived Stem Cells Following FK506 Stimulation Reduce Autophagy of Macrophages in Spine after Nerve Crush Injury.

Macrophages emerge in the milieu around innervated neurons after nerve injuries. Following nerve injury, autophagy is induced in macrophages and affects the regulation of inflammatory responses. It is closely linked to neuroinflammation, while the immunosuppressive drug tacrolimus (FK506) enhances nerve regeneration following nerve crush injury and nerve allotransplantation with additional neuroprotective and neurotrophic functions. The combined use of FK506 and adipose-derived stem cells (ADSCs) was employed in cell therapy for organ transplantation and vascularized composite allotransplantation. This study aimed to investigate the topical application of exosomes secreted by ADSCs following FK506 treatment (ADSC-F-exo) to the injured nerve in a mouse model of sciatic nerve crush injury. Furthermore, isobaric tags for relative and absolute quantitation (iTRAQ) were used to profile the potential exosomal proteins involved in autophagy. Immunohistochemical analysis revealed that nerve crush injuries significantly induced autophagy in the dorsal root ganglia and dorsal horn of the spinal segments. Locally applied ADSC-F-exo significantly reduced autophagy of macrophages in the spinal segments after nerve crush injury. Proteomic analysis showed that of the 22 abundant exosomal proteins detected in ADSC-F-exo, heat shock protein family A member 8 (HSPA8) and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) are involved in exosome-mediated autophagy reduction.

Enhanced Nerve Regeneration by Exosomes Secreted by Adipose-Derived Stem Cells with or without FK506 Stimulation.

Exosomes secreted by adipose-derived stem cells (ADSC-exo) reportedly improve nerve regeneration after peripheral nerve injury. Herein, we investigated whether pretreatment of ADSCs with FK506, an immunosuppressive drug that enhances nerve regeneration, could secret exosomes (ADSC-F-exo) that further augment nerve regeneration. Designed exosomes were topically applied to injured nerve in a mouse model of sciatic nerve crush injury to assess the nerve regeneration efficacy. Outcomes were determined by histomorphometric analysis of semi-thin nerve sections stained with toluidine blue, mouse neurogenesis PCR array, and neurotrophin expression in distal nerve segments. Isobaric tags for relative and absolute quantitation (iTRAQ) were used to profile potential exosomal proteins facilitating nerve regeneration. We observed that locally applied ADSC-exo and ADSC-F-exo significantly enhanced nerve regeneration after nerve crush injury. Pretreatment of ADSCs with FK506 failed to produce exosomes possessing more potent molecules for enhanced nerve regeneration. Proteomic analysis revealed that of 192 exosomal proteins detected in both ADSC-exo and ADSC-F-exo, histone deacetylases (HDACs), amyloid-beta A4 protein (APP), and integrin beta-1 (ITGB1) might be involved in enhancing nerve regeneration.

Increased Angiogenesis by Exosomes Secreted by Adipose-Derived Stem Cells upon Lipopolysaccharide Stimulation.

Exosomes secreted by adipose-derived stem cells (ADSCs) enhance angiogenesis and wound healing. However, in clinical settings, wounds may be infected by various bacteria or pathogens. We investigated whether human ADSCs stimulated with lipopolysaccharide (LPS) secrete exosomes (ADSC-LPS-exo) that augment the angiogenesis of human umbilical vein endothelial cells (HUVECs). ExoQuick-TC exosome precipitation solution was used to purify exosomes from human ADSC culture media in the presence or absence of 1 µg/mL LPS treatment for 24 h. The uptake of ADSC-LPS-exo significantly induced the activation of cAMP response element binding protein (CREB), activating protein 1 (AP-1), and nuclear factor-κB (NF-κB) signaling pathways and increased the migration of and tube formation in HUVECs. RNA interference with CREB, AP-1, or NF-κB1 significantly reduced the migration of and tube formation in HUVECs treated with ADSC-LPS-exo. An experiment with an antibody array for 25 angiogenesis-related proteins revealed that only interleukin-8 expression was significantly upregulated in HUVECs treated with ADSC-LPS-exo. In addition, proteomic analysis revealed that eukaryotic translation initiation factor 4E, amyloid beta A4 protein, integrin beta-1, and ras-related C3 botulinum toxin substrate 1 may be potential candidates involved in ADSC-LPS-exo-mediated enhanced angiogenesis.

The influence of ageing on the incidence and site of trauma femoral fractures: a cross-sectional analysis.

BACKGROUND: This study aimed to determine the influence of ageing on the incidence and site of femoral fractures in trauma patients, by taking the sex, body weight, and trauma mechanisms into account. METHODS: This retrospective study reviewed data from adult trauma patients aged ≥20 years who were admitted into a Level I trauma center, between January 1, 2009 and December 31, 2016. According to the femoral fracture locations, 3859 adult patients with 4011 fracture sites were grouped into five subgroups: proximal type A (n = 1359), proximal type B (n = 1487), proximal type C (n = 59), femoral shaft (n = 640), and distal femur (n = 466) groups. A multivariate logistic regression analysis was applied to identify independent effects of the univariate predictive variables on the occurrence of fracture at a specific site. A two-dimensional plot was presented visually with age and the propensity score accounts for the risk of a fracture at a specific femoral site. RESULTS: This analysis revealed that older age was an independent variable that could positively predict the occurrence of proximal type A (OR [95%CI]: 1.03 [1.03-1.04], p < 0.001) and B fractures (1.02 [1.01-1.02], p < 0.001), and negatively predict the occurrence of proximal type C (0.96 [0.94-0.98], p < 0.001), shaft (0.95 [0.95-0.96], p < 0.001), and distal fractures (0.98 [0.98-0.99], p < 0.001). DISCUSSION: Using the propensity scores which account for the risk of a fracture in a specific femoral site, this study revealed that the older patients were at a higher risk of developing proximal type A and type B fractures, while a lower risk of developing fractures in the shaft and distal femur. This incidence of fracture site can largely be explained by age-related factors, including a decrease in bone strength and falling being the most common mechanism of trauma in older patients. CONCLUSIONS: This study revealed a difference in the involvement of age in the incidence of femoral fracture sites in the trauma patients.

The protective effect of helmet use in motorcycle and bicycle accidents: a propensity score-matched study based on a trauma registry system.

BACKGROUND: Transportation by motorcycle and bicycle has become popular in Taiwan, this study was designed to investigate the protective effect of helmet use during motorcycle and bicycle accidents by using a propensity score-matched study based on trauma registry system data. METHODS: Data of adult patients hospitalized for motorcycle or bicycle accidents between January 1, 2009 and December 31, 2015 were retrieved from the Trauma Registry System. These included 7735 motorcyclists with helmet use, 863 motorcyclists without helmet use, 76 bicyclists with helmet use, and 647 bicyclists without helmet use. The primary outcome measurement was in-hospital mortality. Secondary outcomes were the hospital length of stay (LOS), intensive care unit (ICU) admission rate, and ICU LOS. Normally distributed continuous data were analyzed by the unpaired Student t-test, and non-normally distributed data were compared using the Mann-Whitney U-test. Two-sided Fisher exact or Pearson chi-square tests were used to compare categorical data. Propensity score matching (1:1 ratio using optimal method with a 0.2 caliper width) was performed using NCSS software, adjusting for the following covariates: sex, age, and comorbidities. Further logistic regression was used to evaluate the effect of helmet use on mortality rates of motorcyclists and bicyclists, respectively. RESULTS: The mortality rate for motorcyclists with helmet use (1.1%) was significantly lower than for motorcyclists without helmet use (4.2%; odds ratio [OR] 0.2; 95% confidence interval [CI]: 0.17-0.37; p < 0.001). Among bicyclists, there was no significant difference in mortality rates between the patients with helmet use (5.3%) and those without helmet use (3.7%; OR 1.4; 95% CI: 0.49-4.27; p = 0.524). After propensity-score matching for covariates, including sex, age, and comorbidities, 856 well-balanced pairs of motorcyclists and 76 pairs of bicyclists were identified for outcome comparison, showing that helmet use among motorcyclists was associated with lower mortality rates (OR 0.2; 95% CI: 0.09-0.44; p < 0.001). In contrast, helmet use among bicyclists was not associated with a decrease in mortality (OR 1.3; 95% CI: 0.30-5.96; p = 0.706). The hospital LOS was also significantly shorter for motorcyclists with helmet use than for those without (9.5 days vs. 12.0 days, respectively, p < 0.001) although for bicyclists, helmet use was not associated with hospital LOS. Fewer motorcyclists with helmet use were admitted to the ICU, regardless of the severity of injury; however, no significant difference of ICU admission rates was found between bicyclists with and without helmets. CONCLUSIONS: Motorcycle helmets provide protection to adult motorcyclists involved in traffic accidents and their use is associated with a decrease in mortality rates and the risk of head injuries. However, no such protective effect of helmet use was observed for bicyclists involved in collisions.

Traumatic injuries among adult obese patients in southern Taiwan: a cross-sectional study based on a trauma registry system.

BACKGROUND: The adverse impact of obesity has been extensively studied in the general population; however, the added risk of obesity on trauma-related mortality remains controversial. This study investigated and compared mortality as well injury patterns and length of stay (LOS) in obese and normal-weight patients hospitalized for trauma in the hospital and intensive care unit (ICU) of a Level I trauma center in southern Taiwan. METHODS: Detailed data of 880 obese adult patients with body mass index (BMI) ≥ 30 kg/m(2) and 5391 normal-weight adult patients (25 > BMI ≥ 18.5 kg/m(2)) who had sustained a trauma injury between January 1, 2009 and December 31, 2013 were retrieved from the Trauma Registry System. Pearson's chi-squared, Fisher's exact, and independent Student's t-tests were used to compare differences between groups. Propensity score matching with logistic regression was used to evaluate the effect of obesity on mortality. RESULTS: In this study, obese patients were more often men, motorcycle riders and pedestrians, and had a lower proportion of alcohol intoxication compared to normal-weight patients. Analysis of Abbreviated Injury Scale scores revealed that obese trauma patients presented with a higher rate of injury to the thorax, but a lower rate of facial injuries than normal-weight patients. No significant differences were found between obese and normal-weight patients regarding Injury Severity Score (ISS), Trauma-Injury Severity Score (TRISS), mortality, the proportion of patients admitted to the ICU, or LOS in ICU. After propensity score matching, logistic regression of 66 well-matched pairs did not show a significant influence of obesity on mortality (odds ratio: 1.51, 95% confidence interval: 0.54-4.23 p = 0.438). However, significantly longer hospital LOS (10.6 vs. 9.5 days, respectively, p = 0.044) was observed in obese patients than in normal-weight patients, particularly obese patients with pelvic, tibial, or fibular fractures. CONCLUSION: Compared to normal-weight patients, obese patients presented with different injury characteristics and bodily injury patterns but no difference in mortality.

Weight-reduction through a low-fat diet causes differential expression of circulating microRNAs in obese C57BL/6 mice.

BACKGROUND: To examine the circulating microRNA (miRNA) expression profile in a mouse model of diet-induced obesity (DIO) with subsequent weight reduction achieved via low-fat diet (LFD) feeding. RESULTS: Eighteen C57BL/6NCrl male mice were divided into three subgroups: (1) control, mice were fed a standard AIN-76A (fat: 11.5 kcal %) diet for 12 weeks; (2) DIO, mice were fed a 58 kcal % high-fat diet (HFD) for 12 weeks; and (3) DIO + LFD, mice were fed a HFD for 8 weeks to induce obesity and then switched to a 10.5 kcal % LFD for 4 weeks. A switch to LFD feeding led to decreases in body weight, adiposity, and blood glucose levels in DIO mice. Microarray analysis of miRNA using The Mouse & Rat miRNA OneArray® v4 system revealed significant alterations in the expression of miRNAs in DIO and DIO + LFD mice. Notably, 23 circulating miRNAs (mmu-miR-16, mmu-let-7i, mmu-miR-26a, mmu-miR-17, mmu-miR-107, mmu-miR-195, mmu-miR-20a, mmu-miR-25, mmu-miR-15b, mmu-miR-15a, mmu-let-7b, mmu-let-7a, mmu-let-7c, mmu-miR-103, mmu-let-7f, mmu-miR-106a, mmu-miR-106b, mmu-miR-93, mmu-miR-23b, mmu-miR-21, mmu-miR-30b, mmu-miR-221, and mmu-miR-19b) were significantly downregulated in DIO mice but upregulated in DIO + LFD mice. Target prediction and function annotation of associated genes revealed that these genes were predominantly involved in metabolic, insulin signaling, and adipocytokine signaling pathways that directly link the pathophysiological changes associated with obesity and weight reduction. CONCLUSIONS: These results imply that obesity-related reductions in the expression of circulating miRNAs could be reversed through changes in metabolism associated with weight reduction achieved through LFD feeding.

Profiling the circulating miRNAs in mice exposed to gram-positive and gram-negative bacteria by Illumina small RNA deep sequencing.

BACKGROUND: We profiled the expression of circulating microRNAs (miRNAs) in mice using Illumina small RNA deep sequencing in order to identify the miRNAs that may potentially be used as biomarkers to distinguish between gram-negative and gram-positive bacterial infections. RESULTS: Recombinant-specific gram-negative pathogen Escherichia coli (Xen14) and gram-positive pathogen Staphylococcus aureus (Xen29) were used to induce bacterial infection in mice at a concentration of 1 × 10(8) bacteria/100 µL of phosphate buffered saline (PBS). Small RNA libraries generated from the serum of mice after exposure to PBS, Xen14, Xen29, and Xen14 + Xen29 via the routes of subcutaneous injection (I), cut wound (C), or under grafted skin (S) were analyzed using an Illumina HiSeq2000 Sequencer. Following exposure to gram-negative bacteria alone, no differentially expressed miRNA was found in the injection, cut, or skin graft models. Exposure to mixed bacteria induced a similar expression pattern of the circulating miRNAs to that induced by gram-positive bacterial infection. Upon gram-positive bacterial infection, 9 miRNAs (mir-193b-3p, mir-133a-1-3p, mir-133a-2-3p, mir-133a-1-5p, mir-133b-3p, mir-434-3p, mir-127-3p, mir-676-3p, mir-215-5p) showed upregulation greater than 4-fold with a p-value < 0.01. Among them, mir-193b-3p, mir-133a-1-3p, and mir-133a-2-3p presented the most common miRNA targets expressed in the mice exposed to gram-positive bacterial infection. CONCLUSIONS: This study identified mir-193b-3p, mir-133a-1-3p, and mir-133a-2-3p as potential circulating miRNAs for gram-positive bacterial infections.

Altered exosomal protein expression in the serum of NF-κB knockout mice following skeletal muscle ischemia-reperfusion injury.

BACKGROUND: The NF-κB signaling pathway plays a role in local and remote tissue damage following ischemia-reperfusion (I/R) injury to skeletal muscles. Evidence suggests that exosomes can act as intercellular communicators by transporting active proteins to remote cells and may play a role in regulating inflammatory processes. This study aimed to profile the exosomal protein expression in the serum of NF-κB knockout mice following skeletal muscle ischemia-reperfusion injury. RESULTS: To investigate the potential changes in protein expression mediated by NF-κB in secreted exosomes in the serum following I/R injury, the levels of circulating exosomal proteomes in C57BL/6 and NF-κB(-/-) mice were compared using two dimensional differential in-gel electrophoresis (2-DE), liquid chromatography tandem mass spectrometry (LC-MS/MS), and proteomic analysis. In C57BL/6 mice, the levels of circulating exosomal proteins, including complement component C3 prepropeptide, PK-120 precursor, alpha-amylase one precursor, beta-enolase isoform 1, and adenylosuccinate synthetase isozyme 1, increased following I/R injury. However, in the NF-κB(-/-) mice, the expression of the following was upregulated in the exosomes: protease, serine 1; glyceraldehyde-3-phosphate dehydrogenase-like isoform 1; glyceraldehyde-3-phosphate dehydrogenase; and pregnancy zone protein. In contrast, the expression of apolipoprotein B, complement component C3 prepropeptide, and immunoglobulin kappa light chain variable region was downregulated in NF-κB(-/-) mice. Bioinformatic annotation using the Protein Analysis Through Evolutionary Relationships (PANTHER) database revealed that the expression of the exosomal proteins that participate in metabolic processes and in biological regulation was lower in NF-κB(-/-) mice than in C57BL/6 mice, whereas the expression of proteins that participate in the response to stimuli, in cellular processes, and in the immune system was higher. CONCLUSIONS: The data presented in this study suggest that NF-κB might regulate exosomal protein expression at a remote site via circulation following I/R injury.

Regulatory and Effector Helper T-Cell Profile after Nerve Xenografting in the Toll-Like Receptor-Deficient Mice.

INTRODUCTION: The balance between regulatory T cells (Tregs) and effector T help cells (Th cells) is critical for the control of adaptive immune response during nerve transplantation. However, whether the homeostasis of immune regulation between Tregs and Th cells requires toll-like receptor (TLR) signaling is unclear. The aim of this study is to profile the distribution of spleen Tregs and Th cells in a mouse model of nerve xenografting in the TLR2 and NF-κB gene knockout mice. METHODS: The sciatic nerve was taken from a SD rat or an allogeneic mouse and transplanted to a right back leg of recipient C57BL/6, TLR2(-/-), or NF-κB(-/-) mice by subcutaneous transplantation. After 7 days, the T lymphocytes were then isolated from spleen, stained with phenotyping kits, and analyzed by flow cytometry. RESULTS: The results showed that Tregs were decreased after nerve xenografting in the recipient C57BL/6 mouse. In addition, nerve xenografting also increased the Th1 and Th17 but not the Th2 cell populations. In contrast, amelioration of the Tregs elimination was found in TLR2(-/-) and NF-κB(-/-) mice after transplantation of the nerve xenograft. Moreover, the mice lacking TLR2 or NF-κB showed attenuation of the increase in Th1 and Th17 cells after nerve xenografting. CONCLUSIONS: TLR signaling is involved in T cell population regulation during tissue transplantation. Knock-out of TLR2 and NF-κB prevented Tregs elimination and inhibited Th1- and Th17-driven immune response after nerve xenografting. This study highlighted the potential of inhibiting TLR signaling to modulate T cell-mediated immune regulation to facilitate tolerance to nerve transplantation.

Bicycle-related hospitalizations at a Taiwanese level I Trauma Center.

BACKGROUND: This study aimed to investigate differences in injury severity and mortality between patients who met with bicycle or motorcycle accidents and were hospitalized at a Level I trauma center in Taiwan. METHODS: We performed a retrospective analysis of bicycle-related injuries that have been reported in the Trauma Registry System in order to identify and compare 699 bicyclists to 7,300 motorcyclists who were hospitalized for treatment between January 1, 2009 and December 31, 2013. Statistical analyses of the injury severity, associated complications, and length of stay in the hospital and intensive care unit (ICU) were performed to compare the risk of injury of bicyclists to that of motorcyclists with the corresponding unadjusted odds ratios and 95 % confidence intervals (CIs). Adjusted odds ratios (AORs) and 95 % CIs for mortality were calculated by controlling for confounding variables that included age, and an Injury Severity Score (ISS) was calculated. RESULTS: More of the cyclists were under 19 years of age or over 70 than were the motorcyclists. In contrast, fewer bicyclists than motorcyclists wore helmets, arrived at the emergency department between 11 p.m. and 7 a.m., and had a positive blood alcohol concentration test. The bicyclists sustained significantly higher rates of injuries to the extremities, while motorcyclists sustained significantly higher rates of injuries to the head and neck, face, and thorax. Compared to motorcyclists, the bicyclists had significantly lower ISSs and New Injury Severity Scores, shorter length hospital stays, and a smaller proportion of admittance into the ICU. However, the bicyclists had higher AORs for in-hospital mortality (AOR: 1.2, 95 % CI: 1.16-1.20). In terms of critical injury severity (ISS ≥ 25), the bicyclists had 4.4 times (95 % CI: 1.95-9.82) the odds of mortality than motorcyclists with the same ISSs. CONCLUSIONS: Data analysis indicated that the bicyclists had unique injury characteristics including bodily injury patterns and lower ISSs, but had higher in-hospital mortality compared to motorcycle riders. In this study, given that only 9 % of bicyclists reported wearing helmets and considering the high mortality associated with head injury, it is possible that some bicycle riders underestimated the gravity of cycling accidents.

Motorcycle-related hospitalization of adolescents in a Level I trauma center in southern Taiwan: a cross-sectional study.

BACKGROUND: The aim of this study was to investigate and compare the injury pattern, mechanisms, severity, and mortality of adolescents and adults hospitalized for treatment of trauma following motorcycle accidents in a Level I trauma center. METHODS: Detailed data regarding patients aged 13-19 years (adolescents) and aged 30-50 years (adults) who had sustained trauma due to a motorcycle accident were retrieved from the Trauma Registry System between January 1, 2009 and December 31, 2012. The Pearson's chi-squared test, Fisher's exact test, or the independent Student's t-test were performed to compare the adolescent and adult motorcyclists and to compare the motorcycle drivers and motorcycle pillion. RESULTS: Analysis of Abbreviated Injury Scale (AIS) scores revealed that the adolescent patients had sustained higher rates of facial, abdominal, and hepatic injury and of cranial, mandibular, and femoral fracture but lower rates of thorax and extremity injury; hemothorax; and rib, scapular, clavicle, and humeral fracture compared to the adults. No significant differences were found between the adolescents and adults regarding Injury Severity Score (ISS), New Injury Severity Score (NISS), Trauma-Injury Severity Score (TRISS), mortality, length of hospital stay, or intensive care unit (ICU) admission rate. A significantly greater percentage of adolescents compared to adults were found not to have worn a helmet. Motorcycle riders who had not worn a helmet were found to have a significantly lower first Glasgow Coma Scale (GCS) score, and a significantly higher percentage was found to present with unconscious status, head and neck injury, and cranial fracture compared to those who had worn a helmet. CONCLUSION: Adolescent motorcycle riders comprise a major population of patients hospitalized for treatment of trauma. This population tends to present with a higher injury severity compared to other hospitalized trauma patients and a bodily injury pattern differing from that of adult motorcycle riders, indicating the need to emphasize use of protective equipment, especially helmets, to reduce their rate and severity of injury.

Effect of Weight-Reduction in Obese Mice Lacking Toll-Like Receptor 5 and C57BL/6 Mice Fed a Low-Fat Diet.

BACKGROUND: This study aims to investigate the effect of feeding low-fat diet (LFD) to diet-induced obesity (DIO) mice lacking TLR5 (TLR5(-/-)), which have a tendency to develop glucose intolerance with increased adiposity, compared to that in C57BL/6 mice. RESULTS: TLR5(-/-) and C57BL/6 male mice were divided into three subgroups: (1) control, mice were fed a standard AIN-76A (fat: 11.5 kcal%) diet for 12 weeks; (2) DIO, mice were fed a 58 kcal% high-fat diet (HFD) for 12 weeks; and (3) diet, mice were fed a HFD for 8 weeks to induce obesity and then switched to a 10.5 kcal% LFD for 4 weeks. The glucose intolerance in DIO TLR5(-/-) mice was more significant than that in DIO C57BL/6 mice and was not attenuated by a switch to the LFD. Weight-reduction with LFD had significantly decreased the epididymal fat mass in C57BL/6 mice but not in TLR5(-/-) mice. In addition, the LFD-fed TLR5(-/-) mice showed significantly higher expression of ghrelin in the serum and resistin in the epididymal fat than that in C57BL/6 mice. CONCLUSIONS: This study demonstrated that TLR5 gene knockout impairs some effects of weight-reduction in DIO.

Inhibition of the phosphoinositide 3-kinase pathway decreases innate resistance to lipopolysaccharide toxicity in TLR4 deficient mice.

BACKGROUND: Upon lipopolysaccharide (LPS) stimulation, activation of both the Toll-like receptor 4 (TLR4) and phosphoinositide 3-kinase (PI3K) pathways serves to balance proinflammatory and anti-inflammatory responses. Although the antagonist to TLR4 represents an emerging promising target for the treatment of sepsis; however, the role of the PI3K pathway under TLR4-null conditions is not well understood. This goal of this study was to investigate the effect of inhibition of PI3K on innate resistance to LPS toxicity in a murine model. RESULTS: The overall survival of the cohorts receiving intraperitoneal injections of 100, 500, or 1000 µg LPS from Escherichia coli serotype 026:B6 after 7 d was 100%, 10%, and 10%, respectively. In contrast, no mortality was noted after 500-µg LPS injection in Tlr4-/- mice. When the PI3K inhibitor LY294002 was injected (1 mg/25 g body weight) 1 h prior to the administration of LPS, the overall survival of the Tlr4-/- mice was 30%. In the Tlr4-/- mice, the LPS injection induced no NF-κB activation but an increased Akt phosphorylation in the lung and liver, when compared to that of the C57BL/6 mice. Injection of 500 µg LPS led to a significant induction in O2⁻ detected by electron paramagnetic resonance (EPR) spin trapping spectroscopy in the lung and liver at 3 and 6 h in C57BL/6 but not Tlr4-/- mice. Addition of LY294002 only significantly increased the O2⁻ level in the lung and liver of the Tlr4-/- mice but not in the C57BL/6 mice following 500-µg LPS injection. In addition, the serum IL-1ß and IL-2 levels were more elevated in C57BL/6 mice than in Tlr4-/- mice. Notably, IL-1ß and IL-2 were significantly increased in Tlr4-/- mice but not in the C57BL/6 mice when the PI3K pathway was inhibited by LY294002 prior to LPS injection. CONCLUSIONS: In this study, we demonstrate that innate resistance to LPS toxicity in Tlr4-/- mice is impaired by inhibition of the PI3K pathway, with a corresponding increase in mortality and production of tissue O2⁻ and inflammatory cytokines.

Prognostic Value of Dynamic Segmented Neutrophil to Monocyte (SeMo) Ratio Changes in Patients with Moderate to Severe Traumatic Brain Injury.

BACKGROUND: Traumatic brain injury (TBI) is a leading cause of morbidity and mortality in trauma patients, necessitating reliable prognostic tools. The segmented neutrophil-to-monocyte (SeMo) ratio, indicative of the inflammatory response, has emerged as a valuable biomarker. This study evaluates the prognostic value of dynamic changes in the SeMo ratio in predicting outcomes for patients with moderate to severe TBI. METHODS: A retrospective analysis was conducted on data from 1118 TBI patients admitted to the surgical intensive care unit at a level I trauma center between January 2009 and December 2020. Patients were selected based on an Abbreviated Injury Scale (AIS) score ≥ 3 in the head region. Initial and follow-up SeMo ratios were calculated upon admission and 48-72 h later, respectively. The dynamic SeMo ratio was defined as the difference between the second and initial SeMo ratios. Statistical analyses included receiver operating characteristic (ROC) curve analysis to determine the optimal threshold for mortality prediction, and comparative analysis of clinical outcomes. RESULTS: The study cohort included 121 deceased and 997 surviving patients. Deceased patients had significantly higher second SeMo ratios (20.9 ± 16.1 vs. 15.8 ± 17.2, p = 0.001) and dynamic SeMo ratios (2.4 ± 19.8 vs. -2.1 ± 19.5, p = 0.019) than those survival patients. In the multivariate analysis, the dynamic SeMo is a significant independent risk factor for in-hospital mortality (OR 1.01, 95%CI: 1.01-1.03, p = 0.031). The optimal cut-off for the dynamic SeMo ratio was 5.96, above which patients exhibited higher mortality (21.4% vs. 8.5%, p < 0.001), higher adjusted mortality (adjusted odds ratio: 2.98; 95% confidence interval: 1.95-4.56; p = 0.005), and longer hospital stays (23.6 days vs. 19.7 days, p = 0.005). DISCUSSION: Dynamic SeMo ratio changes serve as a prognostic marker for in-hospital mortality and hospital stay duration in moderate to severe TBI patients. A higher dynamic SeMo ratio indicates increased risk, highlighting the importance of early monitoring and intervention. Future prospective studies should validate these findings and explore integration with other biomarkers for enhanced prognostication.