Annals of clinical and laboratory science最新文献

Objective: Biochemical items play a significant role in clinical decision-making, so this study aims to evaluate the performance of different biochemical platforms.

Methods: We collected 1,524 serum samples that were centrifuged, and plasma was analyzed for HDL-C, LDL-C, Apo A1, Apo B, PA, and Fs-CRP with the Mindray BS2000M and Roche Cobas 8000 platforms. The results were evaluated by a non-parametric two-related sample test, Passing-Bablok regression analysis, Weighted Least Square analysis (WLS), and Bland-Altman analysis according to CLSI EP09-A3, EP5-A2, and EP15-A3.

Results: Between the two systems, there were statistically significant differences in the average bias of LDL-C, Apo A1, Apo B, PA, and Fs-CRP (P<0.001). Moreover, the acceptable clinical performance evaluation test results showed that the differences between low medical decision levels of HDL-C, LDL-C, Apo A1, Apo B, and Fs-CRP in the two systems were clinically unacceptable, while the other items were acceptable.

Conclusion: These findings suggest that the two platforms have good correlation and consistency in high-concentration medical decision levels in HDL-C, LDL-C, Apo A1, Apo B, and Fs-CRP, and all levels of PA in the two platforms are interchangeable and can replace each other.

Objective: The extracellular matrix protein Tenascin-C (TNC) is known to promote the development of pancreatic cancer. The current study investigated the effect and mechanism of Triptolide (TP), a monomer component extracted from the traditional Chinese herb Tripterygium wilfordii, on TNC expression and TNC-mediated malignant behaviors in human pancreatic cancer.

Methods: Database analysis showed TNC expression clusters in pancreatic tissue and the prognostic effect. RT-qPCR and ELISA were performed to detect the TNC expression levels after TP treatment. The miR-sequencing results of TP combined with RT-qPCR were used to screen the potential miRNAs of TP that regulate TNC. The migration and apoptosis ability were detected by scratch assay and flow cytometry.

Results: High levels of TNC are associated with poor outcomes in pancreatic cancer patients. TP suppresses both TNC mRNA levels and protein secretion in a dose-dependent manner. Additionally, TNC mediated migration and gemcitabine resistance were remarkably decreased following TP treatment. In addition, TP significantly inhibited the expression of miR-144, miR-654, and miR-520, and the inhibition of TNC expression by TP was reversed by the miR-144 antagomir. Furthermore, TP could suppress migration and gemcitabine resistance by regulating the miR-144/TNC axis, thereby inhibiting the pancreatic cancer progression.

Conclusion: TP may represent a promising therapeutic candidate for pancreatic cancer by targeting the stroma, with its effects mediated through the miR-144/TNC axis.

Objective: The flow cytometric osmotic fragility test (OFT) screens for hereditary spherocytosis (HS) and other red blood cell (RBC) membrane disorders. It has been suggested that the proportion of residual RBCs at the midpoint region of the time/forward scatter plot would be just as informative for HS diagnosis as that at the terminal region. We established reference intervals for both regions in healthy Korean adults using one of the most recently available flow cytometry systems.

Methods: This cross-sectional study included 249 adults (129 men and 120 women) with normal complete blood cell counts. The proportions of residual RBCs after exposure to hypotonic solution were measured at defined intervals using the BD FACSLyric system. Sex-specific differences were determined.

Results: The reference intervals for the midpoint region for men and women were 16.49-19.61% (median: 18.24%) and 16.27-19.22% (18.18%), respectively, while those for the endpoint region were 7.00-15.21% (11.20%) and 5.58-15.16% (10.87%), respectively. The differences between the sexes and age groups were not statistically significant at either time point.

Conclusion: We established reference intervals for the flow cytometric OFT at the midpoint and endpoint time/forward scatter plot regions in a Korean population for the first time.

Objective: Differentiation imbalance of bone marrow mesenchymal stem cells (BMSCs) is a key mechanism of osteoporosis (OP). This study sought to elucidate how kinsenoside (Kin) regulates ferroptosis and enhances BMSCs' osteogenic differentiation.

Methods: Network pharmacology, Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis, as well as molecular docking were employed for the identification and analysis of the intersection targets of Kin and OP. Differential gene expression analysis was conducted to validate the identified core targets. The culture of BMSCs was carried out in high glucose (HG) cell culture medium containing Kin, the ferroptosis agonist erastin, or the AMP-activated protein kinase (AMPK) inhibitor dorsomorphin (DM). The viability of cells was evaluated using the cell counting kit-8 assay. BMSCs' osteogenic differentiation was assessed through alkaline phosphatase and alizarin red S staining. Additionally, the expression of OP-related proteins, ferroptosis-related proteins, p-AMPK, AMPK, and sirtuin 1 (SIRT1) were detected via Western blot. The ferroptosis of BMSCs was evaluated using 2,7-dichlorofluorescein diacetate staining, along with quantification of malondialdehyde, glutathione, and ferroptosis-related proteins.

Results: HG suppressed BMSCs' osteogenic differentiation and enhanced ferroptosis, while Kin promoted BMSCs' osteogenic differentiation and inhibited ferroptosis. Kin and erastin co-treatment reduced osteogenic differentiation and enhanced ferroptosis of BMSCs compared to Kin treatment alone. Network pharmacology, molecular docking, differential gene expression, and KEGG enrichment analyses, suggested AMPK/SIRT1 as a potential key pathway underlying Kin's therapeutic effect on OP. Kin activated AMPK/SIRT1 signaling pathway, while co-treatment with DM reversed this activation, accompanied by decreased osteogenic differentiation and increased ferroptosis in BMSCs.

Conclusion: Kin enhances BMSCs' osteogenic differentiation under HG conditions by suppressing ferroptosis via activation of the AMPK/SIRT1 signaling pathway.

Objective: Acute lung injury (ALI) is a severe and potentially life-threatening inflammatory disorder of the lungs. Given the limited efficacy and significant side effects of current treatments for ALI, there is an urgency for novel therapeutic strategies.

Methods: Potential pathogenic factors related to the mTOR complex 1 (mTORC1) pathway in ALI were identified through public database mining. In vivo and in vitro models of ALI were constructed through lipopolysaccharide induction. A549 cells were transfected with the constructed si-Hsp90β1, and AMPK activity in cells was inhibited using dorsomorphin. Hematoxylin-eosin (HE) staining, ELISA, lung wet/dry weight (W/D) ratio, and protein content in bronchoalveolar lavage fluid were employed to verify the successful establishment of the ALI animal model. The percentage of Caspase-1 positive cells was determined by the flow cytometry. The expression of Caspase-1 p20 in cells was quantified by immunofluorescence. Expression levels of Hsp90β1, epithelial barrier-related proteins (E-cadherin, ZO-1, and Occludin), pyroptosis-related proteins (Caspase-1, Caspase-1 p20, NLRP3), and proteins related to the AMPK/mTORC1 pathway were detected by western blot.

Results: Through bioinformatics analysis, seven hub genes were screened out, among which Hsp90β1 was selected for further investigation. Hsp90β1 was observed to be up-regulated in the ALI rat model, accompanied by significant histopathological changes, elevated lung W/D ratio, and increased levels of inflammatory factors observed in ALI rats. Both in vitro and in vivo models exhibited enhanced pyroptosis and impaired barrier. In addition, AMPK activity was inhibited and mTORC1 was activated in the ALI rat model. Inhibiting Hsp90β1 reduced inflammation and pyroptosis while restoring barrier function. However, further inhibition of AMPK activity reversed the effect induced by Hsp90β1 inhibition.

Conclusion: Low expression of Hsp90β1 alleviates inflammation, suppresses pyroptosis, and restores epithelial barrier function in ALI through the AMPK/mTORC1 pathway.

Objective: To investigate the effect of exosomal miR-98-5p derived from bone marrow mesenchymal stem cells (BMSCs) on cardiomyocyte injury in acute myocardial infarction (AMI) and analyze its mechanism of action.

Methods: An AMI model in rats was established via ligation of the left anterior descending (LAD) coronary artery. Cardiac systolic function was assessed via echocardiography. Histopathological alterations in myocardial tissue were evaluated by hematoxylin and eosin (HE) staining, while the extent of myocardial infarction was determined through triphenyltetrazolium chloride (TTC) staining. Serum levels of CK-MB, cTnT, and LDH were quantified through ELISA. An AMI cell model was established by subjecting H9C2 cardiomyoblasts to hypoxic conditions. Exosomes were isolated from BMSCs, and their effects on cardiomyocyte injury were investigated. Cellular autophagy levels were examined via western blot (WB). The regulatory interplay between miR-98-5p and E2F6 was validated via a dual-luciferase reporter (DLR) assay.

Results: In comparison to the Sham group, myocardial tissue in rats with AMI exhibited significant structural damage, accompanied by reduced autophagic activity and reduced expression of miR-98-5p. In comparison to the AMI + phosphate-buffered saline (PBS) group, treatment with BMSCs-derived exosomes (BMSCs-exo) markedly improved cardiac function and further enhanced autophagy in AMI rats. In vitro, cells subjected to hypoxic conditions displayed diminished viability and proliferative capacity, increased apoptosis, impaired autophagy, and decreased miR-98-5p expression relative to the control group. However, administration of BMSCs-exo restored miR-98-5p expression, mitigated cellular injury, and promoted autophagic activity. Notably, these protective influences were reversed by the addition of the autophagy inhibitor 3-methyladenine (3-MA). DLR assays confirmed a direct regulatory interaction between miR-98-5p and E2F6. Suppression of miR-98-5p resulted in the upregulation of E2F6, thereby attenuating the reparative effects of BMSCs-exo on myocardial tissue and inhibiting autophagy.

Conclusion: BMSCs-exo miR-98-5p ameliorates AMI-induced myocardial injury by regulating cardiomyocyte autophagy through targeting E2F6.

Objective: To determine the molecular mechanism of pulmonary ischemia-reperfusion (I/R) injury and seek effective therapeutic targets to reduce the incidence and mortality of pulmonary I/R injury.

Methods: Two models were established to explore the expression of the HMGB1-TLR4 pathway in a pulmonary I/R injury, its correlation with downstream inflammatory factors, and the effects of HMGB1-neutralizing antibodies on inflammation.

Results: IL-6 and TNF-α levels in the three mouse models showed a rapid increase, IL-1β, IL-6, and TNF-α were up-regulated in alveolar macrophages after LPS stimulation, TNF-α and HMGB1 were up-regulated in TLR4+/+ cells and peaked at 48 h but was not up-regulated in TLR4-/- cells. Western blot assays revealed that in TLR4+/+ cells, TLR4 was up-regulated after stimulation by LPS and was rapidly down-regulated after treatment with the HMGB1-neutralizing antibody. In contrast, TLR4-/- cells did not respond to LPS stimulation, and the HMGB1-neutralizing antibody did not significantly alter the TLR4 concentration.

Conclusions: HMGB1-TLR4 pathway plays an important role in the regulation of inflammation in pulmonary I/R injury. Furthermore, HMGB1 up-regulated downstream inflammatory factors via TLR4. HMGB1-neutralizing antibodies had a protective effect against lung injury by down-regulating the inflammatory response.

Objective: Accurate measurement of thyroglobulin (Tg) is crucial for managing patients with differentiated thyroid cancer (DTC). This study aims to verify the analytical performance of the Abbott Tg assay and establish Tg reference intervals in an apparently healthy Chinese population.

Methods: Analytical performance was evaluated in terms of precision, detection capabilities, and linearity. A method comparison with the Roche Tg assay was conducted using Passing-Bablok regression and Bland-Altman analysis. Reference intervals were established using serum Tg levels measured by the Abbott Alinity system in 1,081 healthy volunteers, categorized by age and gender.

Results: Repeatability (CV%) for Tg at 0.96 and 7.53 ng/mL was 1.7% and 1.44%, respectively. Within-laboratory imprecision (CV%) at these concentrations were 2.17% and 1.61%. All test results (100%) were at or below the claimed limits of blank (LoB) and detection (LoD), exceeding the 85% threshold for the critical value observation ratio. The CV for the limit of quantification (LoQ) was 6.94%. Passing-Bablok and Bland-Altman analyses for Abbott vs. Roche Tg assays revealed a y-intercept of 0.025, a slope of 0.981, and a Spearman rank correlation coefficient of 0.993 (P<0.0001), with a negative bias of -3.7%. Reference intervals were set between 0.22 and 53.30 ng/mL (2.5^th-97.5^th percentile). Gender had no impact on Tg levels while significant differences were found across different age groups (P=0.008).

Conclusion: The Abbott Tg assay demonstrates verified analytical performance as per the manufacturer's claims. Reference intervals were established for an apparently healthy Chinese population, highlighting the significant influence of age on serum Tg levels.

Objective: Gestational diabetes mellitus (GDM) significantly increases the risk of type 2 diabetes mellitus (T2DM) in both mothers and offspring. Although a two-hour oral glucose tolerance test (OGTT) is advised postpartum, adherence remains poor. Our institution has reported suboptimal adherence (<30%) when testing is deferred to 1-3 months postpartum. In May 2024, the American College of Obstetricians and Gynecologists (ACOG) released updated guidance stating that offering a 75g OGTT during delivery hospitalization in the immediate postpartum period is a reasonable alternative to improve screening. This study evaluates the implementation of the updated guideline and examines early outcomes of inpatient postpartum OGTT over a six-month period.

Methods: Postpartum patients with GDM received a 75g two-hour OGTT during delivery hospitalization, with samples drawn at zero and 120 minutes. Nursing and phlebotomy teams coordinated logistics to ensure test completion. Data were retrospectively reviewed.

Results: Among 196 eligible patients, 184 (93.9%) completed testing. Of those tested, 24 (13.0%) had abnormal glucose values. Reported barriers included patient refusal, discharge timing, and existing hyperglycemia.

Conclusion: Offering inpatient OGTT led to improved screening adherence and detection of abnormal glucose tolerance. This model may enhance postpartum diabetes prevention efforts in high-risk populations.

Objective: Chlamydia pneumoniae is a gram-negative intracellular bacterium that causes respiratory infections, and may contribute to inflammation in asthma. Studies in our laboratory demonstrated that T lymphocyte/cytokine responses to C. pneumoniae in children with asthma were significantly higher, compared with non-asthma, which may indicate the presence of T effector memory (TEM) lymphocytes. In the present study, C. pneumoniae -specific TEMs and their intracellular cytokines were compared in asthmatic and non-asthmatic adults.

Methods: Peripheral blood mononuclear cells (PBMC) (1×10⁶/mL) from asthmatic (N=6) and non-asthmatic (N=14) adults were infected for 24hr +/- C. pneumoniae TW-183 at a multiplicity of infection (MOI)=0.1 and cultured (48 hrs). Distributions of lymphocytes (CD3+, CD4+, CD8+) and TEM cells (CD4+CCR7-CD45RA+CD154+, CD8+CCR7-CD45RA+CD154+) were determined. Levels of intracellular Interleukin (IL)-2, IL-4, and Interferon (IFN)-gamma were measured (flow microfluorimetry).

Results: Numbers of C. pneumoniae-stimulated CD3+CD4+CD45RO+CCR7-TEM (unstimulated, 1:10, 1:100) were higher in asthma compared with non-asthma (mean differences: unstimulated-stimulated) (-21±15, -17±15, -19±15; P=0.02, 0.04, 0.03, respectively) (Wilcoxon-signed rank test). However, CD3+CD4+IFN-gamma+TEMS (1:10, 1:100) were lower in asthma compared with non-asthma (mean differences: 2.2±5, 0.9±1; P=0.03, 0.04, respectively). When stratified according to C. pneumoniae IgG status, numbers of CD3+CD4+IL-2 (1:10) and CD3+CD4+IL-4+ (1:100) cells were higher in C. pneumoniae IgG+ compared with IgG- (mean differences: -0.2±0.2, -1.2±2.4; P=0.02. 0.05, respectively).

Conclusion: Increased numbers of C. pneumoniae -stimulated TEM cells in asthma may indicate decreased effectiveness in clearing infection, suggesting an impaired IFN-gamma response.