MEG3 polymorphisms associated with peripheral blood leukocyte mitochondrial DNA copy number in PAHs-exposure workers.

BACKGROUND: Epidemiological studies have shown that exposure to Polycyclic aromatic hydrocarbons (PAHs) is associated with reduced mitochondrial DNA copy number (mtDNA-CN). Long non-coding RNA maternally expressed gene 3 (MEG3) is involved in mitochondrial function regulation. However, it is unknown whether single-nucleotide polymorphisms in the MEG3 can regulate the mtDNAcn in PAHs exposed populations. The aim of this study was to examine the effect of MEG3 genetic polymorphisms on the mtDNA-CN in PAHs exposed populations. MATERIALS AND METHODS: We recruited 544 coke oven workers and 238 controls using random cluster sampling. High-performance liquid chromatography was used to detect the concentrations of four OH-PAHs (1-hydroxypyrene [1-OHPyr], 1-hydroxynathalene [1-OHNap], 2-hydroxynathalene [2-OHNap], and 3-hydroxyphenanthrene [3-OHPhe]) in urine. The mtDNA-CN of peripheral blood leukocytes was measured using the quantitative polymerase chain reaction method. Sequenom Mass ARRAY matrix-assisted laser desorption/ionization-time of flight mass spectrometry platform was used to detect ten polymorphisms in MEG3. RESULTS: The OH-PAHs levels in the exposure group were significantly higher than those in the control group (P < 0.001). The mtDNA-CN in the exposure group was significantly lower than that in the control group (P < 0.001). A linear regression model revealed that PAHs-exposure (ß [95% confidence interval, CI], -0.428 [-0.475, -0.381], P < 0.001), male gender (-0.052 [-0.098, -0.005], P = 0.029), genotype TT for MEG3 rs11859 (-0.088 [-0.142, -0.035], P = 0.001), and genotype GG for MEG3 rs7155428 (-0.114 [-0.210, -0.017], P = 0.021) were associated with decreased mtDNA-CN. CONCLUSION: PAHs-exposure, male gender, genotype TT for rs11859, and genotype GG for rs7155428 were risk factors for mtDNA-CN.

Combined CD25, CD64, and CD69 Biomarker in 3D-Printed Multizone Millifluidic Device for Sepsis Detection in Clinical Samples.

Sepsis is a serious medical condition that arises from a runaway response to an infection, which triggers the immune system to release chemicals into the bloodstream. This immune response can result in widespread inflammation throughout the body, which may cause harm to vital organs and, in more severe cases, lead to organ failure and death. Timely and accurate diagnosis of sepsis remains a challenge in analytical diagnostics. In this work, we have developed and validated a sepsis detection device, utilizing 3D printing technology, which incorporates multiple affinity separation zones. Our device requires minimal operator intervention and utilizes CD64, CD69, and CD25 as the biomarker targets for detecting sepsis in liquid biopsies. We assessed the effectiveness of our 3D-printed multizone cell separation device by testing it on clinical samples obtained from both septic patients (n = 35) and healthy volunteers (n = 8) and validated its performance accordingly. Unlike previous devices using poly(dimethyl siloxane), the 3D-printed device had reduced nonspecific binding for anti-CD25 capture, allowing this biomarker to be assayed for the first time in cell separations. Our results showed a statistically significant difference in cell capture between septic and healthy samples (with p values of 0.0001 for CD64, CD69, and CD25), suggesting that 3D-printed multizone cell capture is a reliable method for distinguishing sepsis. A receiver operator characteristic (ROC) analysis was performed to determine the accuracy of the captured cell counts for each antigen in detecting sepsis. The ROC area under the curve (AUC) values for on-chip detection of CD64+, CD69+, and CD25+ leukocytes were 0.96, 0.92, and 0.88, respectively, indicating our diagnostic test matches clinical outcomes. When combined for sepsis diagnosis, the AUC value for CD64, CD69, and CD25 was 0.99, indicating an improved diagnostic performance due to the use of multiple biomarkers.

The association between triglyceride-rich lipoproteins, circulating leukocytes, and low-grade inflammation: The Brazilian Longitudinal Study of Adult Health (ELSA-Brasil).

BACKGROUND: Experimental studies have linked triglyceride-rich lipoproteins (TRLs) to inflammation, but the extent of this phenomenon in vivo has not been completely elucidated. OBJECTIVE: We investigated the association between TRL subparticles and inflammatory markers (circulating leukocytes, plasma high-sensitivity C-reactive protein [hs-CRP], and GlycA) in the general population. METHODS: This was a cross-sectional analysis of the Brazilian Longitudinal Study of Adult Health (ELSA-Brasil). TRLs (number of particles per unit volume) and GlycA were measured by nuclear magnetic resonance spectroscopy. The association between TRLs and inflammatory markers was determined by multiple linear regression models adjusted for demographic data, metabolic conditions, and lifestyle factors. Standardized regression coefficients (beta) with 95% confidence intervals are reported. RESULTS: The study population comprised 4,001 individuals (54% females, age 50 ± 9 years). TRLs, especially medium and large subparticles, were associated with GlycA (beta 0.202 [0.168, 0.235], p<0.001 for total TRLs). There was no association between TRLs and hs-CRP (beta 0.022 [-0.011, 0.056], p = 0.190). Medium, large, and very large TRLs were associated with leukocytes, with stronger connections with neutrophils and lymphocytes than monocytes. When TRL subclasses were analyzed as the proportion of the total pool of TRL particles, medium and large TRLs were positively related to leukocytes and GlycA, whereas smaller particles were inversely associated. CONCLUSIONS: There are different patterns of association between TRL subparticles and inflammatory markers. The findings support the hypothesis that TRLs (especially medium and larger subparticles) may induce a low-grade inflammatory environment that involves leukocyte activation and is captured by GlycA, but not hs-CRP.

Investigation of Serum Interleukin 6, High-Sensitivity C-Reactive Protein and White Blood Cell Levels during the Diagnosis and Treatment of Paediatric Appendicitis Patients Before and during the COVID-19 Pandemic.

Introduction: In this study, we prospectively investigated changes in serum interleukin-6 (IL-6), high-sensitivity C-reactive protein (hsCRP) and full white blood cell (WBC) counts during the diagnosis and treatment of paediatric patients with appendicitis. We also investigated the effects of the COVID-19 pandemic on the diagnosis and treatment processes of paediatric appendicitis patients. Materials and Methods: A non-perforated appendicitis group (n = 110), a perforated appendicitis group (n = 35) and an appendicitis + COVID-19 group (n = 8) were formed. Blood samples were taken upon admission and every day until the three studied parameters returned to normal values. To investigate the effects of the COVID-19 pandemic on paediatric appendicitis patients, the perforated appendicitis rates and the times from the onset of the first symptoms to the operation before and during the pandemic were compared. Results: WBC, IL-6, and hsCRP dropped below the upper limits on the second postoperative day in the non-perforated appendicitis group, four to six days postoperatively in the perforated appendicitis group, and three to six days postoperatively in the appendicitis + COVID-19 group. These parameters were not within normal range in patients who developed complications during follow-up. The time from the onset of abdominal pain to the surgery was significantly longer during than before the pandemic in both the non-perforated appendicitis group and the perforated appendicitis group. Conclusions: Our results show that WBC, IL-6, and hsCRP are useful laboratory parameters that can complete clinical examinations in the diagnosis of appendicitis in paediatric patients and the identification of complications that may develop postoperatively.

Cytokine expression profiles in white blood cells of patients with small fiber neuropathy.

BACKGROUND: The role of cytokines in the pathophysiology, diagnosis, and prognosis of small fiber neuropathy (SFN) is incompletely understood. We studied expression profiles of selected pro- and anti-inflammatory cytokines in RNA from white blood cells (WBC) of patients with a medical history and a clinical phenotype suggestive for SFN and compared data with healthy controls. METHODS: We prospectively recruited 52 patients and 21 age- and sex-matched healthy controls. Study participants were characterized in detail and underwent complete neurological examination. Venous blood was drawn for routine and extended laboratory tests, and for WBC isolation. Systemic RNA expression profiles of the pro-inflammatory cytokines interleukin (IL)-1ß, IL-2, IL-8, tumor necrosis factor-alpha (TNF) and the anti-inflammatory cytokines IL-4, IL-10, transforming growth factor beta-1 (TGF) were analyzed. Protein levels of IL-2, IL-8, and TNF were measured in serum of patients and controls. Receiver operating characteristic (ROC)-curve analysis was used to determine the accuracy of IL-2, IL-8, and TNF in differentiating patients and controls. To compare the potential discriminatory efficacy of single versus combined cytokines, equality of different AUCs was tested. RESULTS: WBC gene expression of IL-2, IL-8, and TNF was higher in patients compared to healthy controls (IL-2: p = 0.02; IL-8: p = 0.009; TNF: p = 0.03) and discriminated between the groups (area under the curve (AUC) ≥ 0.68 for each cytokine) with highest diagnostic accuracy reached by combining the three cytokines (AUC = 0.81, sensitivity = 70%, specificity = 86%). Subgroup analysis revealed the following differences: IL-8 and TNF gene expression levels were higher in female patients compared to female controls (IL-8: p = 0.01; TNF: p = 0.03). The combination of TNF with IL-2 and TNF with IL-2 and IL-8 discriminated best between the study groups. IL-2 was higher expressed in patients with moderate pain compared to those with severe pain (p = 0.02). Patients with acral pain showed higher IL-10 gene expression compared to patients with generalized pain (p = 0.004). We further found a negative correlation between the relative gene expression of IL-2 and current pain intensity (p = 0.02). Serum protein levels of IL-2, IL-8, and TNF did not differ between patients and controls. CONCLUSIONS: We identified higher systemic gene expression of IL-2, IL-8, and TNF in SFN patients than in controls, which may be of potential relevance for diagnostics and patient stratification.

C-reactive protein "at first sight": Standard postoperative trend in gynecological surgery and a comparative analysis with white blood cell levels.

OBJECTIVE: There is a lack of information about the normal trend of C-reactive protein (CRP) blood levels in the postoperative days after gynecological benign surgery. We investigated the impact of different surgical techniques on CRP trend. We performed a comparative analysis between a CRP and white blood cell (WBC) trend in postoperative monitoring. METHODS: We studied 207 surgical patients for benign gynecological pathology. We analyzed CRP and WBC levels after surgery in the total number of women and separately by approaches. RESULTS: CRP mean log scores showed a typical behavior. Moreover, results from chi-square test underline that the proportion of women with this result is independent from the type of surgery they underwent. Log score mean values of CPR differed between all groups and between times. No difference in the mean number of white cells between the second and the third day was found, as observed for CRP. CONCLUSIONS: Our study shows a trend reference model in postoperative monitoring of patients with benign gynecological surgery. The comparative analysis between the CRP and WBC trend in the postoperative days provided us data demonstrating the superiority of CRP in postsurgical patient outcomes monitoring.

A Microfluidic Platform Revealing Interactions between Leukocytes and Cancer Cells on Topographic Micropatterns.

Immunoassay for detailed analysis of immune-cancer intercellular interactions can achieve more promising diagnosis and treatment strategies for cancers including nasopharyngeal cancer (NPC). In this study, we report a microfluidic live-cell immunoassay integrated with a microtopographic environment to meet the rising demand for monitoring intercellular interactions in different tumor microenvironments. The developed assay allows: (1) coculture of immune cells and cancer cells on tunable (flat or micrograting) substrates, (2) simultaneous detection of different cytokines in a wide working range of 5-5000 pg/mL, and (3) investigation of migration behaviors of mono- and co-cultured cells on flat/grating platforms for revealing the topography-induced intercellular and cytokine responses. Cytokine monitoring was achieved on-chip by implementing a sensitive and selective microbead-based sandwich assay with an antibody on microbeads, target cytokines, and the matching fluorescent-conjugated detection antibody in an array of active peristaltic mixer-assisted cytokine detection microchambers. Moreover, this immunoassay requires a low sample volume down to 0.5 µL and short assay time (30 min) for on-chip cytokine quantifications. We validated the biocompatibility of the co-culture strategy between immune cells and NPC cells and compared the different immunological states of undifferentiated THP-1 monocytic cells or PMA-differentiated THP-1 macrophages co-culturing with NP460 and NPC43 on topographical and planar substrates, respectively. Hence, the integrated microfluidic platform provides an efficient, broad-range and precise on-chip cytokine detection approach, eliminates the manual sampling procedures and allows on-chip continuous cytokine monitoring without perturbing intercellular microenvironments on different topographical ECM substrates, which has the potential of providing clinical significance in early immune diagnosis, personalized immunotherapy, and precision medicine.

A model for sepsis prediction after retrograde intrarenal surgery and the use of the preoperative/postoperative white blood cell ratio to predict progression from sepsis to septic shock.

OBJECTIVES: To study the predictors of sepsis and progression to septic shock after RIRS; to establish and validate predictive models accordingly. METHODS: In total, 1220 patients were included in the study during. Eight hundred forty-eight patients were assigned to the development cohort and 372 to the validation cohort according to medical record. Univariate and multivariate logistic regression analyses were used to screen independent risk factors for post-RIRS (Retrograde intrarenal surgery) sepsis and progression to septic shock. Nomogram prediction models were established according to the related independent risk factors. Areas under the receiver operating characteristic curves, calibration plots, and DCA (Decision curve analysis) were used to estimate the discrimination, calibration and clinical usefulness of the prediction model, respectively. RESULTS: In the development cohort, sepsis occurred in 59 patients, 16 of whom developed septic shock. Multivariate logistic regression analyses showed that the independent risk factors for sepsis after RIRS were preoperative D-J stent implantation, hydronephrosis > 6.25 HU (Hounsfield units), AGR (Albumin/globulin ratio) < 1.95, hs-CRP/Alb (High-sensitivity C-reactive protein/albumin ratio) > 0.060, operating time > 67.5 min, and urinary nitrite positivity. The preoperative/postoperative WBC ratio > 1.5 was an independent risk factor for progression from sepsis to septic shock. In the development cohort, the AUC (Area under curve) for predicting sepsis risk was 0.845, and the AUC for predicting septic shock risk was 0.896; in the validation cohort, the corresponding values were 0.896 and 0.974, respectively. In the development cohort, the calibration test P values in the sepsis and septic shock cohorts, respectively, were 0.921 and 0.817; in the validation cohort, these values were 0.882 and 0.859. DCA of the model in the sepsis and septic shock cohorts showed threshold probabilities of 10-90% in the development cohort and 10-50% and 10-20% in the validation cohort. CONCLUSION: These individualized nomogram prediction models can improve the early identification of patients at risk for developing sepsis after RIRS or progressing from sepsis to septic shock.

Morphology and cytochemical patterns of peripheral blood cells of tiger frog (Rana rugulosa).

Background: Tiger frog (Rana rugulosa) is a national second-class protected amphibian species in China with an important ecological and economic value. In recent years, due to excessive human hunting, pollution and habitat loss, the wild population of tiger frog has declined sharply. To protect wildlife resources, the artificial breeding of tiger frogs has rapidly developed in China. Diseases are increasing and spreading among tiger frogs due to the increasing scale of artificial farming. The blood examination is the most straightforward and less invasive technique to evaluate the animal health condition. Thus, it is essential to obtain the normal hematological indicators of tiger frogs. The objective of this study was to investigate the morphometry, microstructure and cytochemical patterns of peripheral blood cells in tiger frogs. Methods: The number of blood cells in tiger frogs was counted on a blood count board, and the cell sizes were measured by a micrometer under light microscope. The morphology and classification of blood cells were studied by Wright-Giemsa staining, and the cytochemical pateerns was investigated by various cytochemical staining including periodic acid-Schiff (PAS), Sudan black B (SBB), peroxidase (POX), alkaline phosphatase (AKP), acid phosphatase (ACP), chloroacetic acid AS-D naphthol esterase (CAE) and α-naphthol acetate esterase (ANAE) staining. Results: Besides erythrocytes and thrombocytes, five types of leukocytes were identified in tiger frogs: neutrophils, eosinophils, basophils, lymphocytes and monocytes. The mean erythrocyte, leukocyte and thrombocyte counts were 1.33 ± 0.15 million/mm3, 3.73 ± 0.04 × 104/mm3 and 1.7 ± 0.01 × 104/mm3, respectively. Small lymphocytes were the most abundant leukocytes, followed by large lymphocytes, Neutrophils, eosinophils and monocytes, basophils were the fewest. Eosinophils were strongly positive for PAS, positive for SBB, POX, ACP, CAE, ANAE, while weakly positive for AKP staining; basophils were strongly positive for PAS, ACP, positive for SBB, CAE, weakly positive for ANAE, negative for AKP, POX staining; neutrophils were strongly positive for ACP, SBB, positive for PAS, POX, weakly positive for AKP, CAE and ANAE staining; monocytes were positive for PAS, SBB, ANAE, weakly positive for ACP, AKP, POX, CAE staining; large lymphocytes and thrombocytes were positive for PAS, ACP, weakly positive for ANAE, while negative for SBB, POX, AKP, CAE; small lymphocytes were similar to large lymphocytes, except for strongly positive for PAS and ACP staining. Conclusions: The blood cell types and morphology of tiger frogs were generally similar to those of other amphibians, while their cytochemical patterns had some notable species specificity.Our study could enrich the knowledge of peripheral blood cell morphology and cytochemistry in amphibians, and provide baseline data for health condition evaluation and disease diagnosis of tiger frogs.

A physical wiring diagram for the human immune system.

The human immune system is composed of a distributed network of cells circulating throughout the body, which must dynamically form physical associations and communicate using interactions between their cell-surface proteomes1. Despite their therapeutic potential2, our map of these surface interactions remains incomplete3,4. Here, using a high-throughput surface receptor screening method, we systematically mapped the direct protein interactions across a recombinant library that encompasses most of the surface proteins that are detectable on human leukocytes. We independently validated and determined the biophysical parameters of each novel interaction, resulting in a high-confidence and quantitative view of the receptor wiring that connects human immune cells. By integrating our interactome with expression data, we identified trends in the dynamics of immune interactions and constructed a reductionist mathematical model that predicts cellular connectivity from basic principles. We also developed an interactive multi-tissue single-cell atlas that infers immune interactions throughout the body, revealing potential functional contexts for new interactions and hubs in multicellular networks. Finally, we combined targeted protein stimulation of human leukocytes with multiplex high-content microscopy to link our receptor interactions to functional roles, in terms of both modulating immune responses and maintaining normal patterns of intercellular associations. Together, our work provides a systematic perspective on the intercellular wiring of the human immune system that extends from systems-level principles of immune cell connectivity down to mechanistic characterization of individual receptors, which could offer opportunities for therapeutic intervention.

Telomere length and brain aging: A systematic review and meta-analysis.

The current evidence on the association of leukocyte telomere length (LTL) with age-related structural and cognitive changes in the brain is mixed. Herein conforming to PRISMA 2020 guidelines, we performed a systematic review and meta-analysis using data from 27 observational studies in non-demented individuals. We used effect size and p-value based meta-analysis methods considering marked heterogeneity among studies. We found that the longer LTL was associated with higher brain volume (ß = 0.43, 95%CI: 0.36-0.50%, p = 0.008, N = 1102) and with higher global cognition (ß = 0.01; 95%CI: 0.00-0.02, p = 0.03, N = 19609) by effect size based meta-analysis and with brain volume, hippocampal volume, global cognition, cognitive domains of attention/speed as well as executive functions by p-value based meta-analysis. No significant association of LTL with brain white matter hyperintensities was detected. Furthermore, the evidence strongly suggests a subgroup-specific canonical effect of telomeres, notably in older individuals and females. In conclusion, we provide meta-analytic evidence on the beneficial effect of telomeres on brain structure as well as cognition and advocate for a beneficial subgroup-specific effect that warrants further attention.

Quantitation of methotrexate polyglutamates in human whole blood, erythrocytes and leukocytes collected via venepuncture and volumetric absorptive micro-sampling: a green LC-MS/MS-based method.

Low-dose methotrexate (MTX) plays a key role in treatment of rheumatoid arthritis. However, not all patients respond satisfactorily, and no therapeutic drug monitoring has been implemented in clinical practice, despite the fact that MTX therapy has now been available for decades. Analysis of individual intracellular MTX metabolites among rheumatoid arthritis (RA) patients is hampered by the low intracellular concentrations of MTX-PGs which require a highly sensitive method to quantify. Here, we present a rapid and highly sensitive LC (HILIC) MS/MS method with LLOQ 0.1 nM, 0.8 nmol/L for each metabolite of MTX-PG1-5 and MTX-PG6-7 respectively. Over a linear range of 0.1-100 nM, 0.8-100 nmol/L for each metabolite of MTX-PG1-5 and MTX-PG6-7, respectively, the inter- and intra- accuracy and precision were within 15% of the nominal value for all MTX metabolites. The presented assay was used to assess and compare MTX metabolite concentrations extracted from four different matrices: red blood cells, plasma, peripheral blood mononuclear cells, and whole blood that have been collected either using traditional venepuncture or volumetric absorptive micro-sampling (VAMS) sampling techniques. The presented method not only improves analyte coverage and sensitivity as compared to other published methods; it also improves the greenness.

Associations of the Gut Microbiota Composition and Fecal Short-Chain Fatty Acids with Leukocyte Telomere Length in Children Aged 6 to 9 Years in Guangzhou, China: A Cross-sectional Study.

BACKGROUND: Telomere length (TL) serves as a marker of cellular senescence and appears to plateau between the age of 4 y and young adulthood, after which the gut microbiota are supposed to be established. However, scarce data are available regarding the correlation between gut microbiota composition and TL in the pediatric population. OBJECTIVES: We aimed to investigate whether the gut microbiota and the concentrations of SCFAs in feces are associated with leukocyte TL in children. METHODS: In total, 401 children aged 6-9 y from Guangzhou were enrolled in this cross-sectional study. qPCR was used to determine relative TL in peripheral blood leukocytes. The gut microbiota was characterized by 16S ribosomal RNA amplicon sequencing and the fecal concentrations of total SCFAs and SCFA subtypes were determined using HPLC. The multivariate methods with an unbiased variable selection (MUVR) algorithm and partial least square models were used to select predictable operational taxonomic units (OTUs). Further correlation analyses were performed based on multiple linear regression models with adjustment for covariates and false discovery rate. RESULTS: With the use of MUVR, 35 relevant and minimal optimal OTUs were finally selected. Multiple linear regression analysis showed that the abundance of several OTUs, including OTU334 (belonging to the genus Family XIII AD3011 group), OTU726 (belonging to the species Lachnoclostridium phocaeense), OTU1441 (belonging to the genus Ruminococcus torques group), OTU2553 (belonging to the genus Lachnospiraceae UCG-010), and OTU3375 (belonging to the family Lachnospiraceae), was negatively associated with leukocyte TL (ß: -0.187 to -0.142; false discovery rate (FDR)-corrected P value (PFDR) = 0.009-0.035]. However, neither SCFA subtype nor total SCFA content in feces exhibited significant associations with TL (ß: -0.032 to 0.048; PFDR = 0.915-0.969). CONCLUSIONS: The gut microbiota, but not fecal SCFA concentration, was significantly associated with TL in this pediatric population.

[Comparison of initial periodontal therapy and its correlation with white blood cell level in periodontitis patients with or without diabetes mellitus].

OBJECTIVE: To compare the clinical efficacy of initial periodontal therapy in periodontitis patients with or without type 2 diabetes mellitus and its correlation with white blood cell counts. METHODS: In this study, 32 chronic periodontitis patients without systemic disease (CP group) and 27 chronic periodontitis patients with type 2 diabetes mellitus (CP+DM group) were enrolled. At admission, all the patients went through periodontal examination and fasting blood examination(baseline). Probing depth (PD), attachment loss (AL), bleeding index (BI), plaque index (PLI), white blood cells (WBC) counts and fasting blood glucose (FBG) were recorded respectively, while hemoglobin A1c (HbA1c) was recorded only in CP+DM group. After that, initial periodontal therapy was performed. All the tests were repeated 3 and 6 months after treatment. The changes of periodontal clinical indexes and WBC levels were compared between the two groups before and after treatment, and the correlation between WBC and periodontal clinical indexes and glucose metabolism indexes were analyzed by generalized linear mixed model. RESULTS: At baseline, the periodontal inflammation and destruction were similar in CP and CP+DM group, but the WBC level was significantly higher in CP+DM groups [(6.01±1.26)×109/L vs. (7.14±1.99)×109/L, P=0.01]. After 3 and 6 months of initial periodontal therapy, the mean PD, AL, BI, and PLI in CP+DM and CP groups were significantly lower than the baseline, and the PD in CP+DM group was further decreased by 6 months compared with 3 months [(3.33±0.62) mm vs. (3.61±0.60) mm, P < 0.05]. However, none of these periodontal indexes showed significant difference between the two groups by 3 or 6 months. In CP+DM group, HbA1c at 3 months and 6 months were significantly lower than the baseline [(7.09±0.79)% vs. (7.64±1.16)%, P < 0.05; (7.06±0.78)% vs. (7.64±1.16)%, P < 0.05], and FBG was significantly lower than the baseline by 6 months [(7.35±1.14) mmol/L vs. (8.40±1.43) mmol/L, P < 0.05]. The WBC level in CP group was significantly lower than the baseline level by 3 months [(5.35±1.37)×109/L vs. (6.01±1.26)×109/L, P < 0.05], while that in CP+DM group was significantly lower than the baseline level by 6 months [(6.00±1.37)×109/L vs. (7.14±1.99)×109/L, P < 0.05]. The analysis of genera-lized linear mixed model showed that WBC level was significantly positively correlated with PD and FBG (P < 0.05). CONCLUSION: Initial periodontal therapy can effectively improve the periodontal clinical status of patients with or without type 2 diabetes mellitus, and have benefits on glycemic control in diabetic patients. However, the response of periodontal indexes and WBC level to initial therapy were relatively delayed in diabetic patients. WBC plays an important role in the correlation between diabetes mellitus and periodontitis.

Identification of glucocorticoid-related molecular signature by whole blood methylome analysis.

OBJECTIVE: Cushing's syndrome represents a state of excessive glucocorticoids related to glucocorticoid treatments or to endogenous hypercortisolism. Cushing's syndrome is associated with high morbidity, with significant inter-individual variability. Likewise, adrenal insufficiency is a life-threatening condition of cortisol deprivation. Currently, hormone assays contribute to identify Cushing's syndrome or adrenal insufficiency. However, no biomarker directly quantifies the biological glucocorticoid action. The aim of this study was to identify such markers. DESIGN: We evaluated whole blood DNA methylome in 94 samples obtained from patients with different glucocorticoid states (Cushing's syndrome, eucortisolism, adrenal insufficiency). We used an independent cohort of 91 samples for validation. METHODS: Leukocyte DNA was obtained from whole blood samples. Methylome was determined using the Illumina methylation chip array (~850 000 CpG sites). Both unsupervised (principal component analysis) and supervised (Limma) methods were used to explore methylome profiles. A Lasso-penalized regression was used to select optimal discriminating features. RESULTS: Whole blood methylation profile was able to discriminate samples by their glucocorticoid status: glucocorticoid excess was associated with DNA hypomethylation, recovering within months after Cushing's syndrome correction. In Cushing's syndrome, an enrichment in hypomethylated CpG sites was observed in the region of FKBP5 gene locus. A methylation predictor of glucocorticoid excess was built on a training cohort and validated on two independent cohorts. Potential CpG sites associated with the risk for specific complications, such as glucocorticoid-related hypertension or osteoporosis, were identified, needing now to be confirmed on independent cohorts. CONCLUSIONS: Whole blood DNA methylome is dynamically impacted by glucocorticoids. This biomarker could contribute to better assessment of glucocorticoid action beyond hormone assays.

Comparison of initial periodontal therapy and its correlation with white blood cell level in periodontitis patients with or without diabetes mellitus / 北京大学学报(医学版)

OBJECTIVE@#To compare the clinical efficacy of initial periodontal therapy in periodontitis patients with or without type 2 diabetes mellitus and its correlation with white blood cell counts.@*METHODS@#In this study, 32 chronic periodontitis patients without systemic disease (CP group) and 27 chronic periodontitis patients with type 2 diabetes mellitus (CP+DM group) were enrolled. At admission, all the patients went through periodontal examination and fasting blood examination(baseline). Probing depth (PD), attachment loss (AL), bleeding index (BI), plaque index (PLI), white blood cells (WBC) counts and fasting blood glucose (FBG) were recorded respectively, while hemoglobin A1c (HbA1c) was recorded only in CP+DM group. After that, initial periodontal therapy was performed. All the tests were repeated 3 and 6 months after treatment. The changes of periodontal clinical indexes and WBC levels were compared between the two groups before and after treatment, and the correlation between WBC and periodontal clinical indexes and glucose metabolism indexes were analyzed by generalized linear mixed model.@*RESULTS@#At baseline, the periodontal inflammation and destruction were similar in CP and CP+DM group, but the WBC level was significantly higher in CP+DM groups [(6.01±1.26)×109/L vs. (7.14±1.99)×109/L, P=0.01]. After 3 and 6 months of initial periodontal therapy, the mean PD, AL, BI, and PLI in CP+DM and CP groups were significantly lower than the baseline, and the PD in CP+DM group was further decreased by 6 months compared with 3 months [(3.33±0.62) mm vs. (3.61±0.60) mm, P < 0.05]. However, none of these periodontal indexes showed significant difference between the two groups by 3 or 6 months. In CP+DM group, HbA1c at 3 months and 6 months were significantly lower than the baseline [(7.09±0.79)% vs. (7.64±1.16)%, P < 0.05; (7.06±0.78)% vs. (7.64±1.16)%, P < 0.05], and FBG was significantly lower than the baseline by 6 months [(7.35±1.14) mmol/L vs. (8.40±1.43) mmol/L, P < 0.05]. The WBC level in CP group was significantly lower than the baseline level by 3 months [(5.35±1.37)×109/L vs. (6.01±1.26)×109/L, P < 0.05], while that in CP+DM group was significantly lower than the baseline level by 6 months [(6.00±1.37)×109/L vs. (7.14±1.99)×109/L, P < 0.05]. The analysis of genera-lized linear mixed model showed that WBC level was significantly positively correlated with PD and FBG (P < 0.05).@*CONCLUSION@#Initial periodontal therapy can effectively improve the periodontal clinical status of patients with or without type 2 diabetes mellitus, and have benefits on glycemic control in diabetic patients. However, the response of periodontal indexes and WBC level to initial therapy were relatively delayed in diabetic patients. WBC plays an important role in the correlation between diabetes mellitus and periodontitis.

Lyophilised Platelet-Rich Fibrin: Physical and Biological Characterisation.

BACKGROUND: Platelet-rich fibrin (PRF) has gained popularity in craniofacial surgery, as it provides an excellent reservoir of autologous growth factors (GFs) that are essential for bone regeneration. However, the low elastic modulus, short-term clinical application, poor storage potential and limitations in emergency therapy use restrict its more widespread clinical application. This study fabricates lyophilised PRF (Ly-PRF), evaluates its physical and biological properties, and explores its application for craniofacial tissue engineering purposes. MATERIAL AND METHODS: A lyophilisation method was applied, and the outcome was evaluated and compared with traditionally prepared PRF. We investigated how lyophilisation affected PRF's physical characteristics and biological properties by determining: (1) the physical and morphological architecture of Ly-PRF using SEM, and (2) the kinetic release of PDGF-AB using ELISA. RESULTS: Ly-PRF exhibited a dense and homogeneous interconnected 3D fibrin network. Moreover, clusters of morphologically consistent cells of platelets and leukocytes were apparent within Ly-PRF, along with evidence of PDGF-AB release in accordance with previously reports. CONCLUSIONS: The protocol established in this study for Ly-PRF preparation demonstrated versatility, and provides a biomaterial with growth factor release for potential use as a craniofacial bioscaffold.

Platelet activation and antimicrobial activity of L-PRF: a preliminary study.

Leukocyte and platelet rich fibrin (L-PRF) is one of the platelet concentrates used to support regeneration and healing process. Many studies showed possible immunological and antibacterial properties of L-PRF. We perform an in vitro study to analyze the effect of L-PRF on platelet activation, platelet-leukocytes interactions and antimicrobial activity, important components in the healing process. Molecular biomarkers related with platelet activation and platelet-leukocyte interactions were analyzed by means of flow cytometry when L-PRF exudate was added to whole blood platelets. L-PRF membrane was used to evaluate antimicrobial activity using Enterococcus faecalis (ATCC 29212), Pseudomonas aeruginosa (ATCC 27853) and Candida albicans (ATCC 90028). Our experimental design allows to evaluate platelet activation and analyze molecular biomarkers of other immune cells and platelet-leukocyte interactions. From the results obtained we can conclude that L-PRF can be a valuable tool in healing process, efficient in activating platelets of whole blood and inhibiting microbial growth. In our opinion, the use of L-PRF exudate, in addition to L-PRF membrane, presents some advantages that have to be considered in clinical trials. Additional research on the characterization and quantification of cells and its products present in the L-PRF exudate, as well as on the temporal factor released. Also, further studies using strains isolated from clinical cases are needed.

Impact of g force and timing on the characteristics of platelet-rich fibrin matrices.

Recently, new centrifugation protocols for the preparation of platelet-rich fibrin (PRF) have been introduced in an attempt to further improve the beneficial impact of these 2nd generation platelet concentrate membranes. This in-vitro study aimed to compare the biological and physical characteristics of three types of PRF membranes using two different centrifuges with adapted relative centrifugal forces (RCF): leucocyte- and platelet-rich fibrin, advanced platelet-rich fibrin, and advanced platelet-rich fibrin+. Release of growth factors, macroscopic dimensions, cellular content and mechanical properties of the respective membranes, prepared from blood of the same individual were explored. Furthermore, the impact of timing (blood draw-centrifugation and centrifugation-membrane preparation) was assessed morphologically as well as by electron microscopy scanning. No statistically significant differences amongst the three PRF modifications could be observed, neither in their release of growth factors or the cellular content, nor in clot/membrane dimensions. The difference between both centrifuges were negligible when the same g-force was used. A lower g-force, however, reduced membrane tensile strength. Timing in the preparation process had a significant impact. Adaptation of RCF only had a minimal impact on the final characteristics of PRF membranes.

DNA translated: Friedrich Miescher's discovery of nuclein in its original context.

In 1871, the Swiss physiological chemist Friedrich Miescher published the results of a detailed chemical analysis of pus cells, in which he showed that the nuclei of these cells contained a hitherto unknown phosphorus-rich chemical which he named 'nuclein' for its specific localisation. Published in German, 'Ueber Die Chemische Zusammensetzung Der Eiterzellen', [On the Chemical Composition of Pus Cells] Medicinisch-Chemische Untersuchungen (1871) 4: 441-60, was the first publication to describe DNA, and yet remains relatively obscure. We therefore undertook a translation of the paper into English, which, together with the original article, can be accessed via the following link https://doi.org/10.1017/S000708742000062X. In this paper, we offer some intellectual context for its publication and immediate reception.