Experimental Biology and Medicine最新文献

Accurate identifying internal tandem duplication (ITD) mutation is indispensable for diagnosis and prognosis of acute myeloid leukemia (AML) patients, but specialized full-size detection tools are lacking. Therefore, we aimed to develop a reliable system for accurate assessment of ITD mutations of various size ranges and improve prognosis for AML. Bone marrow samples from AML patients from December 2021 to March 2022 were collected for methodology establishment. After a large-scale sample testing by next-generation sequencing (NGS), a short-read tandem duplication recognition system based on soft-clip was established. During performance validation, the lower detection limit was set to a parameter close to capillary electrophoresis ("gold standard") by adjusting reference values (sensitivity 3-5%). Data simulation was performed using the FLT3 gene CDS as wild-type data. Methodological concordance of this system with capillary electrophoresis was analyzed. The applicability to other pathogenic tandem duplication mutations was validated. We have developed an innovative NGS-based system named "ITDFinder" for accurate detection of ITD mutations, with the lower detection limit of 4%, corresponding to a sequencing depth of 1000X. Compared to capillary electrophoresis, ITDFinder exhibited good consistency (mean difference: -0.0085) in mutation detection and correlation across various length of ITD. Clinical case validation (n = 1,032) showed an overall agreement rate of 96.5% between the two approaches used for characterization. In addition, data simulation results suggested that the new system could observe BCOR-ITD and KMT2A-PTD mutations (depths, 500-1300X; mutation rates, 0.04-0.8). The innovative mutation detection system is appropriate to small-to large-sized ITDs and other pathogenic tandem duplication mutations, expected to save 96.3% of the workload. This offers significant potential for accurate clinical assessment of ITD mutations and subsequent prognosis in AML patients.

As an inflammatory disease, atherosclerosis is associated with acute ischemic stroke (AIS), but its early identification and intervention efficacy remain suboptimal. A new research direction may be to explore peripheral atherosclerotic biomarkers from the perspective of mitochondrial dysfunction, which can induce inflammatory cell activation. Moreover, the degree of overall cervicocephalic atherosclerosis (namely, atherosclerotic burden) is more closely related to AIS prognosis than local atherosclerotic lesions. Therefore, this study investigated the relationship between mitochondrial dysfunction in peripheral blood mononuclear cells (PBMCs), including monocytes and lymphocytes, and overall cervicocephalic atherosclerotic burden and AIS outcome. Patients with AIS and cervicocephalic atherosclerosis were enrolled and followed up for 90 days. The reactive oxygen species (ROS) and the mitochondrial deoxyribonucleic acid copy number (mtDNA-CN) in PBMCs were measured respectively through a fluorescence probe and a droplet digital polymerase chain reaction to evaluate mitochondrial function. The overall intracranial and cervical atherosclerotic burden (ICAB) was quantified by summing up the atherosclerosis degree points in each arterial segment as assessed by computed tomography angiography. A modified Rankin Scale (mRS) score >2 was considered a 90-day unfavorable functional outcome. Five (4.9%) of the 103 patients with AIS were lost to follow-up. mtDNA-CN [adjusted β = -0.099, 95% confidence intervals (CIs) = -0.153 ∼ -0.044, p < 0.001] and ROS content (adjusted β = 1.275, 95%CI = 0.885 ∼ 1.665, p < 0.001) were correlated with ICAB. The risk of a 90-day unfavorable functional outcome increased with higher ROS content [adjusted odds ratio (OR) = 1.523, 95%CI = 1.172 ∼ 1.981, p = 0.002] and decreased with higher mtDNA-CN (adjusted OR = 0.911, 95%CI = 0.850 ∼ 0.976, p = 0.008). PBMC mitochondrial dysfunction was found to be independently associated with extensive and severe cervicocephalic atherosclerosis and a 90-day unfavorable functional outcome in patients with AIS, which may provide a novel approach to improving the early identification and risk stratification of cervicocephalic atherosclerosis, along with the prediction of the outcome of atherosclerotic AIS.

Peripheral nerve trauma commonly results in chronic neuropathic pain by up-regulating the synthesis and release of pro-inflammatory mediators from local and invading cells and inducing hyperexcitability of nociceptive neurons and spontaneous electrical activity. The pain decreases when these cells down-regulate genes supporting the pro-inflammatory state, up-regulate genes for expressing anti-inflammatory factors, and modulate genes that reduce nociceptive neuron spontaneous electrical activity. Pharmacological agents, the primary technique for reducing pain, do not eliminate pain, and <50% of patients achieve benefits because they do not address the underlying causes of pain. Alternative techniques providing longer lasting, but not complete or long-term pain relief include surgical interventions, electrical stimulation, and antibody treatment. Anti-inflammatory mediators can reduce pain, but the effect is not complete or long-lasting. Platelet-rich plasma (PRP) contains a readably available evolutionarily developed cocktail of factors that induce longer-lasting and more significant, but not complete, pain relief than other techniques. However, a novel study shows that unique formulations of PRP can induce long-term pain elimination. This review examines (1) the efficacy of drugs, regenerative peripheral nerve interface (RPNI), targeted muscle reinnervation (TMR), and PRP in reducing chronic neuropathic pain, (2) recent clinical data showing that a novel PRP application technique induces long-term chronic neuropathic pain reduction/elimination, and (3) discusses why the novel PRP may be more effective in reducing/eliminating chronic neuropathic pain.

Fentanyl is a potent and short-acting opioid that is often given to pediatric patients during surgery to relieve pain and as an adjunct to anesthesia. Its effects on the developing brain are yet to be determined. In the present study, commercially available human neural stem cells (NSCs) were used to model the effects of fentanyl on the developing human brain. We determined the dose dependent effects and temporal relationships between fentanyl exposures and NSC health, viability, and differentiation. Markers of mitochondrial health [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetra-zolium bromide (MTT)] and cell death/damage [lactate dehydrogenase (LDH)] were monitored to determine the dose response effects of fentanyl on NSC viability. In addition, lipidomics analysis was conducted to investigate lipid profile changes in differentiated neural cells treated with fentanyl. Fentanyl did not cause a significant increase in LDH release, nor MTT reduction after 24-h exposure at concentrations of 0.5, 1.0, 3.0, 10, or 100 μM, for both NSCs and differentiated neural cells. Lipidomics data showed the top 15 most variable important in projection (VIP) lipid species (the higher the VIP scores, the bigger changes in treated groups vs. controls), including lysophosphatidylcholines (LPCs), lysophosphatidylethanolamines (LPEs), ceramides (CER), cholesterol esters (ChEs) and sphingosine (SPH). The lipidomic data indicate that LPC (16:0), LPC (16:1), LPC (18:1), CER (d18:0_22:0), CER (d18:2_18:0), CER(d18:2_24:1) were significantly increased, and only ChE (24:5) and SPH (d18:1) were significantly decreased in the highest dose group versus control. These data indicated that fentanyl exposure (24-h) did not induce detectable cell death. However, a lipidomic analysis indicated that fentanyl may affect immature neural cell functions through modifying lipid composition and lipid metabolism. These data indicated that despite the absence of clear neurodegeneration, fentanyl may still have a negative impact on the developing brain.

Ceramides and other sphingolipids are associated with diabetes risk. Here, we examined the association of plasma sphingolipids with insulin sensitivity and secretion in people without diabetes. We enrolled adults without diabetes based on 75-g oral glucose tolerance test. Assessments included clinical examination, insulin sensitivity (hyperinsulinemic euglycemic clamp), and insulin secretion (intravenous glucose tolerance test). Plasma levels of 58 sphingolipid species (including ceramides, monohexosylceramides, sphingomyelins, and sphingosine) were assayed using liquid chromatography tandem mass spectrometry. The study participants (N = 240; 129 Black, 111 White) had a mean age of 43.1 ± 12.0 y, body mass index (BMI) 29.4 ± 6.23 kg/m², fasting plasma glucose 91.4 ± 6.91 mg/dL, and 2-h plasma glucose 123 ± 26.3 mg/dL. Several of the 58 SPLs species assayed showed variable associations with insulin sensitivity (r = 0.17-0.35, P = 0.039 - <0.0001) and secretion (r = 0.14-0.27; P = 0.038 - <0.0001). After correction for multiple testing, plasma levels of very-long-chain (VLC) monohexosylceramide C34:0 (r = 0.31 - 0.43, P < 0.0001) and VLC sphingomyelins C28-C34 (r = 0.31-0.35, P = 0.0004 - <0.0001) were significantly associated with insulin sensitivity. Plasma VLC sphingomyelin level were inversely associated with insulin secretion, plasma glucose, BMI, and waist circumference. We conclude that circulating VLC sphingomyelins are associated positively with insulin action and inversely with insulin secretion and adiposity in normoglycemic adults, indicating a possible link to glucoregulation that precedes the development of dysglycemia.

In-ovo imaging using ostrich eggs has been described as an alternative to animal testing using rodents. This approach is not considered an animal experiment and it does not require small-animal imaging devices as ostrich eggs provide good image quality on regular CT, MRI or PET used in humans. The aims of this study were 1) to describe methods of radiopharmaceutical injection, 2) to explore normal biodistribution of F-18-FDG during a 60-min list-mode-PET/CT examination and 3) to compare biodistribution in-ovo to existing literature considering chicken and rodents. Vessel access was successful in 54/78 ostrich eggs. Highest FDG-uptake was observed in epiphyseal plates (0.36 ± 0.06 IA%/g; range 0.29-0.48 IA%/g) and brain (0.25 ± 0.05 IA%/g; range 0.21-0.36 IA%/g). In-vivo activity distribution on PET and ex-vivo activity distribution (well counter) showed comparable results (Spearman's Rho range 0.795-0.882). No significant differences were observed regarding previous isoflurane exposure. Normal biodistribution of F-18-FDG in ostrich embryos using a standard PET/CT system for humans was mainly found as expected with highest uptake in epiphyseal plates and brain which is comparable to results on rodents and chicken embryos. Isoflurane anesthesia did not reveal significant differences regarding organ uptake. The results of this normal distribution study allow for interpretation of future disease models (inflammation, tumor) in ostrich embryos using F-18-FDG as radiopharmaceutical.

This study explored the association between inflammatory biomarkers-C-reactive protein to albumin ratio (CAR), platelet to lymphocyte ratio (PLR), and neutrophil to lymphocyte ratio (NLR)-and the prognosis of patients with cardiogenic cerebral embolism (CCE). We retrospectively analyzed data from 80 CCE patients diagnosed between June 2020 and June 2024, categorizing them into favorable and unfavorable prognosis groups based on outcomes such as death, recurrence, and disability. The CAR, PLR, and NLR values were calculated from routine blood tests, and statistical analyses, including Spearman correlation, multivariate logistic regression, and ROC curve analysis, were performed to examine their prognostic significance. Results showed that the unfavorable prognosis group had significantly higher CAR, PLR, and NLR values compared to the favorable group (P < 0.05). Spearman correlation analysis revealed positive associations between these biomarkers and prognosis (r = 0.319 for CAR, 0.238 for PLR, 0.251 for NLR, all P < 0.05). Multivariate analysis identified CAR and NLR as independent risk factors for unfavorable prognosis (OR = 1.034 for CAR, OR = 3.887 for NLR). ROC analysis determined optimal cutoff values for CAR (>0.74), PLR (>160.00), and NLR (>3.53) to predict unfavorable prognosis with AUCs of 0.796, 0.694, and 0.705, respectively. The combined biomarker test yielded an AUC of 0.899. Kaplan-Meier survival analysis indicated significantly lower survival rates for patients with higher levels of CAR, PLR, and NLR (P < 0.05). In conclusion, elevated CAR, PLR, and NLR are reliable indicators of a poor prognosis in CCE patients.

Cannabidiol (CBD) has been used for different purposes by different populations in recent years. When consumed by pregnant women, CBD can pass through the placenta and enter the fetal blood stream. There is concern over adverse effects of fetal exposure to CBD and its major metabolites (7-OH-CBD and 7-COOH-CBD). In the present study, human neural stem cells (NSCs) were treated with CBD and its metabolites at different concentrations for various durations to understand how the drug may affect fetal brain development. NSCs were also treated with delta-9 tetrahydrocannabinol (THC) for comparison purposes. CBD, 7-OH-CBD and 7-COOH-CBD dose-dependently reduced NSC viability. CBD and 7-OH-CBD reduced NSC number at the G1 phase. A 24 h exposure did not cause significant change in NSC proliferation. At concentrations comparable to those detected in human blood, longer exposures to CBD, 7-OH-CBD and 7-COOH-CBD caused more obvious cell death. After NSCs differentiation, CBD treatment reduced GFAP and cannabinoid receptor 2 (CB2) expression. THC treatment reduced the GFAP expression, but the change in CB2 expression did not reach statistical significance. The expression of cannabinoid receptor 1 (CB1) and beta-tubulin III were not significantly altered by drug exposures. The study demonstrated that clinically relevant concentrations of CBD, 7-OH-CBD and 7-COOH-CBD affect basic physiological features of human NSCs. After NSC differentiation, the reduced expression of CB2 receptors and GFAP on differentiated cells further indicated the vulnerability of developing central nervous system to CBD and THC. These data will help to contextualize in vivo neurodevelopmental studies that may not accurately model human metabolite profiles of CBD.