Proportion Of Leukocytes Research Articles

Association of innate versus specific immunity with heart failure incidence: a prospective study.

Immune disorders are key heart failure (HF) triggers, but little is known about whether the status of immunity affects the incidence of HF. To explore this, we used blood cell counts and derived ratios to investigate the association between immunity status markers and HF incidence. The number and proportion of peripheral blood leucocytes in a physiological state are related to the body's immune status. Neutrophils, monocytes, SII (systemic immune-inflammatory index), NLR (neutrophil-to-lymphocyte ratio), and PLR (platelet-to-lymphocyte ratio) serve as innate immunity status markers, while lymphocytes and LMR (lymphocyte-to-monocyte ratio) serve as specific immunity status markers. 330 362 UK Biobank (UKB) participants were finally examined. Cox proportional hazard models were used to explore the relationship between immunity status markers and HF incidence. Flexible parametric survival models were used to capture time-varying relationships between blood cell ratios and HRs for HF. Subgroup analyses were conducted by age, sex, and body mass index. Finally, sensitivity analyses were performed to validate the results. During a median follow-up of 14.1 years, 9611 (2.9%) participants developed HF. Neutrophils, monocytes, SII, and NLR were positively associated with HF incidence, with fully adjusted per SD increment HR (95% CI) of 1.20 (1.17 to 1.22), 1.09 (1.07 to 1.12), 1.12 (1.10 to 1.14), and 1.16 (1.14 to 1.18), respectively. Platelets, lymphocytes, and LMR were inversely correlated with HF incidence, with fully adjusted per SD increment HR (95% CI) of 0.97 (0.95 to 1.00), 0.97 (0.95 to 0.99), and 0.90 (0.88 to 0.92), respectively. The innate immunity status markers were positively associated with HF incidence, while specific immunity status markers exhibited an inverse association, offering novel insights for HF prediction and intervention.

The microbiome biomarkers of pregnant women’s vaginal area predict preterm prelabor rupture in Western China

IntroductionIntraamniotic infection is crucial in preterm prelabor rupture of membranes(PPROM), a clinical condition resulting from the invasion of vaginal opportunistic microbes into the amniotic cavity. Although previous studies have suggested potential associations between infection and PPROM, the role of vaginalopportunistic bacteria in PPROM has received limited attention.MethodsThis study aimed to confirm the vaginal bacterial etiology of PPROM. We investigated vaginal microbiotas using automatic analysis of vaginal discharge, microbiological tests, and 16s rRNA genehigh-throughput sequencing.ResultsThe research findings revealed that the proportion of parabasal epitheliocytes, leukocytes, toxic leukocytes, and bacteria with diameters smaller than 1.5 um was significantly higher in the PPROM group than that in the normal full-term labor (TL) group. The top three vaginal opportunistic bacterial isolates in all participants were 9.47% Escherichia coli, 5.99% Streptococcus agalactiae, and 3.57% Enterococcus faecalis. The bacterial resistance differed, but all the isolates were sensitive to nitrofurantoin. Compared with the vaginal microbiota dysbiosis (VMD) TL (C) group, the VMD PPROM (P) group demonstrated more operational taxonomic units, a high richness of bacterial taxa, and a different beta-diversity index. Indicator species analysis revealed that Lactobacillus jensenii, Lactobacillus crispatus, and Veillonellaceae bacterium DNF00626 were strongly associated with the C group. Unlike the C group, the indicator bacteria in the P group were Enterococcus faecalis, Escherichia coli, and Streptococcus agalactiae.DiscussionThese findings provide solidevidence that an abnormal vaginal microbiome is a very crucial risk factorclosely related to PPROM. There were no unique bacteria in the vaginalmicrobiota of the PPROM group; however, the relative abundance of bacteria inthe abnormal vaginal flora of PPROM pregnancies differed. Antibiotics should bereasonably selected based on drug sensitivity testing. The findings presented in this paper enhance our understanding of Streptococcus agalactiae, Enterococcus faecalis, and Escherichia coli vaginal bacterial etiology of PPROM in Western China.

Interactions between rumen epithelium-associated microbiota and host immunological and metabolic adaptations in response to different milk replacer feeding intensities in dairy calves

The milk replacer feeding regime in dairy calves has a great impact on metabolic and immunological functioning and affects animal welfare and lifetime performance. The feeding regime influences the rumen microbial composition, and epithelium-associated microbes may interact with the immune system of the host. We examined the correlations between blood leukocyte counts and the rumen epithelium-associated microbiome in dairy calves fed 2 different milk replacer feeding intensities and if these factors related to metabolic traits. Fourteen newborn female dairy calves were allocated to a group receiving either 10% (n = 7) or 20% (n = 7) milk replacer of their body weight (on average 41 kg) and provided ad libitum access to grass hay and concentrate pellets. At 3 weeks of life, all calves were fitted with a rumen cannula. Calves were weaned at 12 weeks of life and received a total mixed ration for ad libitum intake. Pre- (8 to 10 weeks of life) and post-weaning (21 to 23 weeks of life), methane production was measured in respiration chambers, and rumen epithelium and blood were sampled for 16S rRNA sequencing and leukocyte analyses, respectively. Pre-weaning, the reduced milk replacer feeding intensity was accompanied with higher concentrate intake but lower growth performance (P < 0.001), a higher abundance of amylolytic and lower abundance of cellulolytic epimural microbes. The group fed a low milk replacer intensity had also greater portions of monocytes (P = 0.031), CD8+ (P < 0.001), and CD14+ (P = 0.044) leukocytes, suggesting elevated inflammatory conditions. Correlations between CD8+ T cells and rumen methanogens, Ruminococcaceae, and Lachnospiraceae were recorded, but these were not consistent throughout maturation. Post-weaning, differences in feed intake and rumen microbial composition converged among milk replacer groups, while differences in growth performance (P = 0.040) and CD8+ cells (P < 0.001) were still present. In conclusion, a reduced milk replacer feeding intensity in dairy calves compromised growth performance and immunity and this effect persisted in the long-term. Significant correlations between the proportion of leukocytes and distinct epimural microbe taxa indicated an interplay between rumen epimural colonization and immune functioning of the host. However, further research is required addressing this interplay between rumen epimural microbes and immune functioning in dairy calves.

Comparative morphology of blood cells of Ambassis nalua, A. vachelli, and Inegocia japonica

Hematological investigation is an essential tool for monitoring fish health. This research compared morphometric traits and blood cell characteristics between two representative pelagic fish species (Ambassis nalua and A. vachelli) and one benthic species (Inegocia japonica) collected from seagrass meadows off Libong Island in Thailand, where they coexist with human activity. Blood samples were collected and prepared using the smear technique. The erythrocytes of all observed specimens had elliptical or oval shapes. The benthic fish I. japonica significantly outperformed both pelagic fish in terms of the mean size of many erythrocyte features. However, more erythrocyte nuclear abnormalities were found in I. japonica, along with A. nalua. Lymphocytes made up the highest proportion of leukocytes in I. japonica, followed by neutrophils. The morphometric erythrocyte data of I. japonica possibly indicated the greater oxygen requirement of fish living in demersal habitats. The baseline parameters from the hematological data of the sampled fish will be used to monitor environmental quality.

Immune response to cold exposure: Role of γδ T cells and TLR2-mediated inflammation.

The mammalian body possesses remarkable adaptability to cold exposure, involving intricate adjustments in cellular metabolism, ultimately leading to thermogenesis. However, cold-induced stress can impact immune response, primarily through noradrenaline-mediated pathways. In our study, we utilized a rat model subjected to short-term or long-term mild cold exposure to investigate systemic immune response during the cold acclimation. To provide human relevance, we included a group of regular cold swimmers in our study. Our research revealed complex relationship between cold exposure, neural signaling, immune response, and thermogenic regulation. One-day cold exposure triggered stress response, including cytokine production in white adipose tissue, subsequently activating brown adipose tissue, and inducing thermogenesis. We further studied systemic immune response, including the proportion of leukocytes and cytokines production. Interestingly, γδ T cells emerged as possible regulators in the broader systemic response, suggesting their possible contribution in the dynamic process of cold adaptation. We employed RNA-seq to gain further insights into the mechanisms by which γδ T cells participate in the response to cold. Additionally, we challenged rats exposed to cold with the Toll-like receptor 2 agonist, showing significant modulation of immune response. These findings significantly contribute to understanding of the physiological acclimation that occur in response to cold exposure.

Haemosporidian parasites (Apicomplexa, Haemosporida) of breeding common starling Sturnus vulgaris in Latvia

Migratory behaviour in seasonal environments affects host–pathogen relationships, especially for vector‐transmitted blood parasites of the order Haemosporida. The common starling Sturnus vulgaris is a short‐distance migrant where the north‐eastern European breeding population spend the non‐breeding season in temperate mild western Europe. Despite the high abundance and known susceptibility as a host, blood parasitism in wild starlings has rarely been studied with molecular methods. Here, we monitored haemosporidian parasitism in a Latvian starling population over four breeding seasons. We found a total annual parasite prevalence of 2.7–15.7% caused by four Haemoproteus, three Plasmodium, and one Leucocytozoon cytochrome‐b (cyt‐b) genetic lineages. Herein, seven of these lineages have been recorded for the first time in the common starling as host. Lineage‐specific parasitemia was generally low (Haemoproteus range: 0.008–1.028%, Plasmodium range: 0.002–0.005%, Leucocytozoon range: 0.003–0.004%) indicating chronic infection stages in all parasitised hosts during the breeding season. Additionally, the proportion of leukocytes in peripheral blood was enhanced in infected compared to non‐infected hosts indicating activated immune defence during the chronic infection stage. Finally, 11% (3 out of 27) of individuals had cleared the infection from peripheral blood after one year. Causes for the variability in infection prevalence in common starlings across years, as well as the transmission period during the host annual cycle, are still open for future studies.

VEGFA, VEGFR1, VEGFR2 serum and cerebrospinal fluid concentration in patients with acute leukemia

Background. Vascular endothelial growth factor A (VEGFA) is one of the most important factors for regulation of hematopoietic stem cells differentiation. It is involved in leukemogenesis and central nervous system (CNS) damage in acute leukemia. According to the literature, the VEGFA production by blast cells is increased, but the values of serum concentration and the associations with CNS involvement are contradictory.Aim. evaluate the VEGFA, VEGFR1, VEGFR2 concentration in serum and cerebrospinal fluid of patient with different types of acute leukemia in disease onset and during treatment.Materials and methods. The concentration of VEGFA in serum and cerebrospinal fluid was studied in 74 primary patients with acute leukemia. The comparison group consisted of 67 healthy donors. VEGFR1, VEGFR2 were studied in serum and cerebrospinal fluid in 34 patients at the onset of the disease. The comparison group consisted of 10 healthy donors. For the analysis, an enzyme immunoassay was used on a semi-automatic Personal Lab analyzer (Adaltis) and Affymetrix eBioscience human VEGF-A Platinum ELISA reagents.Results. Serum VEGFA concentration was statistically significantly lower in acute leukemia patients than that of donors (median 149.78 and 432.19 pg/ml respectively; p &lt;0.0001). Factor deficiency was significantly more pronounced in patients with blastemia (p &lt;0.015). During antitumor therapy, there was a tendency to increase the amount of the factor in the blood serum. Serum concentration of soluble VEGFR2 was also lower in patients than that of donors (6949.9 and 8795.9 pg/ml respectively; p = 0.0026). For concentration of VEGFR1 such deviations were not found. The concentrations of VEGFR1 and VEGFR2 in serum were higher than in cerebrospinal fluid (p &lt;0.0001), while VEGFR1 showed a positive correlation between serum and cerebrospinal fluid concentrations. the concentration of VEGFR1 in the cerebrospinal fluid was significantly lower in patients with B-lymphoblastic leukemia/lymphoma compared to other types of leukemia.Conclusion. the concentration of VEGFA in serum decreases in patients with blastemia, this may indicate a lack of secretion and excessive consumption of the factor by blast cells with a decrease in the proportion of leukocytes that normally secrete the factor. In the cerebrospinal fluid, the concentrations of VEGFR1 and VEGFR2 are lower than in serum, with the lowest values being found in patients with B-lymphoblastic leukemia/lymphoma, but no relationship with the development of CNS involvement was found.

Abstract 3424: Assessing a DNA methylation-based telomere length estimator in individuals with a heavy smoking history

Abstract Telomere length (TL) and DNA methylation are associated with age and smoking exposure, as well as aging-related outcomes including lung cancer and mortality. A DNA methylation-based TL score (DNAmTL) was developed to estimate TL using DNA methylation array data, both in blood. DNAmTL effectively estimates telomere-related aging using 140 CpGs that are moderately enriched in sub-telomeric regions and not included in existing epigenetic clocks. We examined DNAmTL in individuals with a heavy smoking history (mean 56 pack years, range 11-185) in a nested lung cancer case-control study of 350 cases and 350 controls from the β-Carotene and Retinol Efficacy Trial identified through 2005. Twenty-two controls became cases with additional follow-up through 2013, so 328 controls and 372 cases were considered for these analyses. Leukocyte TL was measured using quantitative polymerase chain reaction (T/S ratio, log2-transformed), and DNA methylation was assayed on the Illumina EPIC array, in blood samples obtained on average 4.9 years prior to diagnosis in cases. We observed weak positive Pearson correlations between DNAmTL and TL in cases (r=0.19, p=0.0003) and controls (r=0.14, p=0.009) that persisted after age-adjustment: r=0.16, p=0.002 and r=0.14, p=0.014, respectively. Age-adjusted DNAmTL, but not TL, was longer for female compared to male cases (T=3.6, p=0.0004) and controls (T=2.2, p=0.03), and for those in the placebo compared to the intervention arm in cases (T=2.1, p=0.03) and controls (T=2.3, p=0.02). Linear regression models of TL and DNAmTL adjusted for age, sex, and race and ethnicity were not statistically significant for smoking variables at blood draw (pack years, current vs former smoking status, cigarettes smoked per day during smoking history, years smoked, years since quit), nor asbestos exposure in either cases or controls. Longer age-adjusted DNAmTL was associated with a lower hazard ratio (HR) for time to lung cancer death (0.60, 95% Confidence Interval (CI): 0.37-0.97) among cases (n=313 lung cancer-specific mortality events) in Cox proportional hazards models adjusted for age, sex, race and ethnicity, and time from blood draw to diagnosis with stage as a stratification variable (early I/II, advanced III/IV, unknown). Analogous age-adjusted TL model showed a similar direction but did not reach statistical significance (HR 0.86, CI: 0.71-1.05). Further adjustment of these models by DNA methylation-estimated leukocyte proportions of granulocytes, monocytes, natural killer, CD8+T, and CD4+T (excluding B cells) yielded similar results for age-adjusted DNAmTL (HR 0.54, CI: 0.31-0.92) and age-adjusted TL (HR 0.87, CI: 0.71-1.05). Our results illustrate that while DNAmTL may not be closely associated with smoking exposure among heavily exposed individuals, shorter DNAmTL may be more strongly associated with lung cancer mortality than TL in individuals with a heavy smoking history. Citation Format: Laurie Grieshober, Stefan Graw, Matt J. Barnett, Gary E. Goodman, Chu Chen, Devin C. Koestler, Carmen J. Marsit, Jennifer A. Doherty. Assessing a DNA methylation-based telomere length estimator in individuals with a heavy smoking history [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3424.

Testing androgen-induced immunosuppression: Environmental androgens as a model system for steroid-immune interaction.

It is well known that hormones influence and direct most facets of physiology; however, there is still contention regarding the directions of certain relationships, for example, between gonadal hormones and immunity. Among the many proposed relationships relating to gonadal-immune interactions, support for immunosuppressive effects of androgens remains prominent within physiological literature. Although ample study has been directed towardthe immunosuppressive effects of androgens, considerable disagreement remains regarding their influence on immune function. In this study, we test the hypothesis that androgens inhibit immunocompetence in the American alligator (Alligator mississippiensis). Developing alligators were incubated at female-producing temperatures with a subset of individuals being exposed to 17-α-methyltestosterone (MT) before sexual determination. 17-α-methyltestosterone is a potent androgen, not aromatizable by crocodilians, that has been found to exert masculinizing effects in exposed crocodilian populations in vivo and in vitro. Additionally, a subset of animals was exposed to a novel antigen to quantify innate and acquired immune function. We recovered no significant differences in leukocyte ratios or proportions between groups and found no significant differences in innate immune function as measured by hemolysis-hemagglutination. However, we did find significant differences in acquired immune function, where masculinized individuals expressed greater antibody titers. Our findings reject the hypothesis that androgens suppress immune function; rather, androgens may be immunoenhancing to acquired humoral responses and neutral to innate humoral immunity in crocodilians.

Genetic parameters for novel mastitis traits defined by combining test-day somatic cell score and differential somatic cell count in the first lactation of Japanese Holsteins

In this study, we aimed to improve current udder health genetic evaluations by addressing the limitations of monthly sampled somatic cell score (SCS) for distinguishing cows with robust innate immunity from those susceptible to chronic infections. The objectives were to (1) establish novel somatic cell traits by integrating SCS and the differential somatic cell count (DSCC), which represents the combined proportion of polymorphonuclear leukocytes and lymphocytes in somatic cells and (2) estimate genetic parameters for the new traits, including their daily heritability and genetic correlations with milk production traits and SCS, using a random regression test-day model (RRTDM). We derived 3 traits, namely ML_SCS_DSCC, SCS_4_DSCC_65_binary, and ML_SCS_DSCC_binary, by using milk loss estimates at corresponding SCS and DSCC levels, thresholds established in previous studies, and a threshold established from milk loss estimates, respectively. Data consisted of test-day records collected during January 2021 through March 2022 from 265 herds in Hokkaido, Japan. From these records, we extracted records between 7 to 305 d in milk (DIM) in the first lactation to fit the RRTDM. The model included the random effect of herd-test-day, the fixed effect of year-month, fixed lactation curves nested with calving age groups, and random regressions with Legendre polynomials of order 3 for additive genetic and permanent environmental effects. The analysis was performed using Gibbs sampling with Gibbsf90+ software. The averages (ranges) of daily heritability estimates over lactation were 0.086 (0.075 to 0.095) for SCS, 0.104 (0.073 to 0.127) for ML_SCS_DSCC, 0.137 (0.014 to 0.297) for SCS_4_DSCC_65_binary, and 0.138 (0.115 to 0.185) for ML_SCS_DSCC_binary; the heritability curve for SCS_4_DSCC_65_binary was erratic. Genetic correlations within the trait decreased as the DIM interval widened, especially for those integrating DSCC, indicating that these traits should be analyzed using RRTDM rather than repeatability models. The averages (ranges) of genetic correlations with milk yield over lactation were 0.01 (-0.22 to 0.28) for SCS, -0.05 (-0.40 to 0.13) for ML_SCS_DSCC, -0.08 (-0.17 to 0.09) for SCS_4_DSCC_65_binary, and -0.08 (-0.22 to 0.27) for ML_SCS_DSCC_binary. Compared with SCS, the newly defined traits exhibited slightly stronger negative genetic correlations with milk yield. Especially in late lactation stages, the genetic correlation between ML_SCS_DSCC and milk yield was significantly below zero, with a posterior median of -0.40. Furthermore, the new traits showed positive correlations with SCS, having estimates varying from 0.68 to 0.85 for ML_SCS_DSCC, 0.14 to 0.47 for SCS_4_DSCC_65_binary, and 0.61 to 0.66 for ML_SCS_DSCC_binary, depending on DIM. Considering that ML_SCS_DSCC and ML_SCS_DSCC_binary have relatively high heritability (compared with SCS) and favorable genetic correlations with milk production traits and SCS, their incorporation into breeding programs appears promising. Nevertheless, their genetic relationships with (sub)clinical mastitis require further investigation.

Different leukocyte subsets are targeted by systemic and locoregional administration despite conserved nanomaterial characteristics optimal for lymph node delivery.

Lymph nodes (LNs) house a large proportion of the body's leukocytes. Accordingly, engineered nanomaterials are increasingly developed to direct therapeutics to LNs to enhance their efficacy. Yet while lymphatic delivery of nanomaterials to LNs upon locoregional injection has been extensively evaluated, nanomaterial delivery to LN-localized leukocytes after intravenous administration has not been systematically explored nor benchmarked. In this work, a panel of inert, fluorescent nanoscale tracers and drug delivery vehicles were utilized to interrogate intravenous versus locoregionally administered nanomaterial access to LNs and leukocyte subsets therein. Hydrodynamic size and material effects on LN accumulation extents were similar between intravenous versus intradermal injection routes. Nanomaterial distribution to various LN leukocyte subsets differed substantially with injection route, however, in a manner not proportional to total LN accumulation. While intravenously administered nanomaterials accumulated in LNs lowly compared to systemic tissues, in sharp contrast to locoregional delivery, they exhibited size-dependent but material-independent access to immune cells within the LN parenchyma, which are not easily accessed with locoregional delivery.

Baseline haematological parameters in three common Australian frog species.

Amphibians are experiencing declines globally, with emerging infectious diseases as one of the main causes. Haematological parameters present a useful method for determining the health status of animals and the effects of particular diseases, but the interpretation of differential cell counts relies on knowing the normal ranges for the species and factors that can affect these counts. However, there is very little data on either normal haematological parameters or guides for blood cell types for free-ranging frog species across the world. This study aims to 1) create a visual guide for three different Australian frog species: Litoria paraewingi, Limnodynastes dumerilii, and Crinia signifera, 2) determine the proportions of erythrocytes to leukocytes and 3) differential leukocytes within blood smears from these three species and 4) assess the association between parasites and differential counts. We collected blood samples from free-ranging frogs and analysed blood smears. We also looked for ectoparasites and tested for the fungal disease chytridiomycosis. Overall, we found that the differentials of erythrocytes to leukocytes were not affected by species, but the proportions of different leukocytes did vary across species. For example, while lymphocytes were the most common type of leukocyte across the three species, eosinophils were relatively common in Limnodynastes dumerilii but rarely present in the other two species. We noted chytridiomycosis infection as well as ectoparasites present in some individuals but found no effect of parasites on blood parameters. Our results add baseline haematological parameters for three Australian frog species and provide an example of how different frog species can vary in their differential blood cell counts. More information is needed on frog haematological data before these parameters can be used to determine the health status of wild or captive frogs.

Diet Quality and Epigenetic Aging in the Women’s Health Initiative

BackgroundHigher diet quality scores are associated with a lower risk for many chronic diseases and all-cause mortality; however, it is unclear if diet quality is associated with aging biology. ObjectiveThis study aimed to examine the association between diet quality and a measure of biological aging known as epigenetic aging. DesignA cross-sectional data analysis was used to examine the association between three diet quality scores based on self-reported food frequency questionnaire data and five measures of epigenetic aging based on DNA methylation (DNAm) data from peripheral blood. Participants/settingThis study included 4,500 postmenopausal women recruited from multiple sites across the United States (1993-98), aged 50 to 79 years, with food frequency questionnaire and DNAm data available from the Women’s Health Initiative baseline visit. Main outcome measuresFive established epigenetic aging measures were generated from HumanMethylation450 Beadchip DNAm data, including AgeAccelHannum, AgeAccelHorvath, AgeAccelPheno, AgeAccelGrim, and DunedinPACE. Statistical analyses performedLinear mixed models were used to test for associations between three diet quality scores (Healthy Eating Index, Dietary Approaches to Stop Hypertension, and alternate Mediterranean diet scores) and epigenetic aging measures, adjusted for age, race and ethnicity, education, tobacco smoking, physical activity, Women’s Health Initiative substudy from which DNAm data were obtained, and DNAm-based estimates of leukocyte proportions. ResultsHealthy Eating Index, Dietary Approaches to Stop Hypertension, and alternate Mediterranean diet scores were all inversely associated with AgeAccelPheno, AgeAccelGrim, and DunedinPACE (P < 0.05), with the largest effects with DunedinPACE. A one standard deviation increment in diet quality scores was associated with a decrement (β ± SE) in DunedinPACE z score of –0.097 ± 0.014 (P = 9.70 x 10-13) for Healthy Eating Index, –0.107 ± 0.014 (P = 1.53 x 10-14) for Dietary Approaches to Stop Hypertension, and –0.068 ± 0.013 (P = 2.31 x 10-07) for the alternate Mediterranean diet. ConclusionsIn postmenopausal women, diet quality scores were inversely associated with DNAm-based measures of biological aging, particularly DunedinPACE.

Cytology of Turkey Blood-Reactive Hemograms and Measures of Stress

The current study was intended to offer a cytological counterbalance to published descriptions of how processing or other procedures affect turkey welfare. Cytology represents a detailed description of morphological atypia or unusual intracellular or intercellular behavior. The study aimed to describe the variation of blood cells of commercial turkeys. Blood films were collected from 4 turkeys at four different ages of 6, 12, 17, and 19 weeks at commercial farms by a qualified veterinarian. The slides, stained by Wright-Giemsa, were photographed and interpreted off-site. Normal cells of the lymphocyte (L) and heterophil (H) series were described first, followed by examples of atypical cells of other series. These were shown with descriptions of cellularity defined as the proportion of leukocytes in each microscopic field. The results indicated examples of cells whose presence in a standard differential count (SDC) was important enough to disqualify the simple H/L ratio as a stress measure. These cells were atypical members of the lymphoid series, plasmacytes, and other cell types. Atypical granulocytes were heterophils with irregular shapes and faint nuclear staining (hypochromia). An example of a representative total white count revealed how the H/L value could depend on where the cells were counted on the slide. In conclusion, the cytology clearly shows that the presence of atypical cells in a hemogram highlights the inadequacy of relying solely on the simple H/L ratio to estimate stress status.

Classification of Peripheral Blood Leukocyte Phenotypes and Serum Cytokines in Vogt-Koyanagi-Harada Disease before and after Glucocorticoid Therapy.

Vogt-Koyanagi-Harada disease (VKH) is an autoimmune disease, and glucocorticoid therapy (GC) is widely used for VKH. We provided a profile of leukocyte populations and serum cytokines in VKH patients under GC. A prospective observational study was conducted on three treatment-naïve VKH patients. Peripheral blood samples were collected from the patients before GC (VKH-acute) and after 6 months (VKH-remission), and healthy individuals were used as controls. Proportions of 37-type leukocytes and levels of 27-kind cytokines were measured by mass cytometry and multiplex bead analysis. Property similarity was analyzed using hierarchical cluster analysis. The leukocytes and cytokines were broadly classified into four and three clusters: (1) a cluster with high intensity in VKH-acute consisting of B cells, Th2-like, Th17-like, basophils, and IL-7 and IP-10; (2) a cluster with high intensity in VKH-remission composed of monocytes, neutrophils, IL-4, and TNFα; in leukocytes, (3) a cluster with low intensity in VKH-acute and -remission consisting of CD8+ T cells, Th1-like, and NKT cells; (4) a cluster with low intensity in VKH-remission composed of NK cells, Tregs, and DCs; and in cytokines, (5) a cluster with high intensities in VKH-acute and -remission comprising G-CSF, MCP-1, eotaxin, and IL-17A. These findings suggest that inflammatory composition in blood during the acute phase of VKH represents complex hyperimmune responses dominantly driven by Th and B cells.

Calprotectin blockade inhibits long-term vascular pathology following peritoneal dialysis-associated bacterial infection.

Bacterial infections and the concurrent inflammation have been associated with increased long-term cardiovascular (CV) risk. In patients receiving peritoneal dialysis (PD), bacterial peritonitis is a common occurrence, and each episode further increases late CV mortality risk. However, the underlying mechanism(s) remains to be elucidated before safe and efficient anti-inflammatory interventions can be developed. Damage-Associated Molecular Patterns (DAMPs) have been shown to contribute to the acute inflammatory response to infections, but a potential role for DAMPs in mediating long-term vascular inflammation and CV risk following infection resolution in PD, has not been investigated. We found that bacterial peritonitis in mice that resolved within 24h led to CV disease-promoting systemic and vascular immune-mediated inflammatory responses that were maintained up to 28 days. These included higher blood proportions of inflammatory leukocytes displaying increased adhesion molecule expression, higher plasma cytokines levels, and increased aortic inflammatory and atherosclerosis-associated gene expression. These effects were also observed in infected nephropathic mice and amplified in mice routinely exposed to PD fluids. A peritonitis episode resulted in elevated plasma levels of the DAMP Calprotectin, both in PD patients and mice, here the increase was maintained up to 28 days. In vitro, the ability of culture supernatants from infected cells to promote key inflammatory and atherosclerosis-associated cellular responses, such as monocyte chemotaxis, and foam cell formation, was Calprotectin-dependent. In vivo, Calprotectin blockade robustly inhibited the short and long-term peripheral and vascular consequences of peritonitis, thereby demonstrating that targeting of the DAMP Calprotectin is a promising therapeutic strategy to reduce the long-lasting vascular inflammatory aftermath of an infection, notably PD-associated peritonitis, ultimately lowering CV risk.

RNA sequencing of peripheral blood in amyotrophic lateral sclerosis reveals distinct molecular subtypes: Considerations for biomarker discovery.

Amyotrophic lateral sclerosis (ALS) is a heterogeneous neurodegenerative disease with limited therapeutic options. A key factor limiting the development of effective therapeutics is the lack of disease biomarkers. We sought to assess whether biomarkers for diagnosis, prognosis or cohort stratification could be identified by RNA sequencing (RNA-seq) of ALS patient peripheral blood. Whole blood RNA-seq data were generated for 96 Australian sporadic ALS (sALS) cases and 48 healthy controls (NCBI GEO accession GSE234297). Differences in sALS-control gene expression, transcript usage and predicted leukocyte proportions were assessed, with pathway analysis used to predict the activity state of biological processes. Weighted Gene Co-expression Network Analysis (WGCNA) and machine learning algorithms were applied to search for diagnostic and prognostic gene expression patterns. Unsupervised clustering analysis was employed to determine whether sALS patient subgroups could be detected. Two hundred and forty-five differentially expressed genes were identified in sALS patients relative to controls, with enrichment of immune, metabolic and stress-related pathways. sALS patients also demonstrated switches in transcript usage across a small set of genes. We established a classification model that distinguished sALS patients from controls with an accuracy of 78% (sensitivity: 79%, specificity: 75%) using the expression of 20 genes. Clustering analysis identified four patient subgroups with gene expression signatures and immune cell proportions reflective of distinct peripheral effects. Our findings suggest that peripheral blood RNA-seq can identify diagnostic biomarkers and distinguish molecular subtypes of sALS patients however, its prognostic value requires further investigation.

Cold Mechanical Isolation of Placental Macrophages as a Method to Limit Procedure-Induced Activation of Macrophages.

Isolation of placental macrophages using enzymatic digestion at warm temperatures is widely used for invitro studies. However, studies in brain and kidney tissue show that this method activates immune cells, immediate early genes, and heat shock proteins. Isolating placental macrophages while preserving their tissue-specific characteristics as much as possible is pivotal to reliably studying their functions. We therefore developed a mechanical dissociation protocol at low temperatures and compared this to enzymatic digestion at high temperatures. Decidual and villous macrophages were isolated from term human placentas. A cell suspension was generated by mechanical dissociation using a gentleMACS. For warm enzymatic digestion, Accutase was added, followed by incubation at 37°C. Macrophages were isolated after Ficoll density gradient centrifugation. Cell types were analyzed with flow cytometry (CD45, CD14, CD80, CD86, CD163, and CD206) and their activation status with real-time PCR (FOS, JUN, HSP27, HSP70, IL1β, TNFα, IL10, and TGFβ) after cell sorting. A higher proportion of leukocytes and macrophages was obtained from the villi with cold mechanical dissociation (p < 0.05). Compared to warm enzymatic digestion, cold mechanical dissociation resulted in a higher expression of CD163 in villous and decidual macrophages (p < 0.05). Warm enzymatic digestion showed higher levels of TNFα, IL1β, and IL10 in decidual and villous macrophages, and HSP70 in villous macrophages. Our data show that mechanical dissociation of placental tissue at low temperatures is associated with less activation of placental macrophages. This suggests that cold mechanical dissociation is a preferred method, resulting in macrophages that more closely resemble their in-tissue state.

Single-cell transcriptomics reveals subtype-specific molecular profiles in Nrf2-deficient macrophages from murine atherosclerotic aortas.

Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcriptional regulator of antioxidant and anti-inflammatory response in all cell types. It also activates the transcription of genes important for macrophage function. Nrf2 activity declines with age and has been closely linked to atherosclerosis, but its specific role in this vascular pathology is not clear. Atherosclerotic plaques contain several macrophage subsets with distinct, yet not completely understood, functions in the lesion development. The aim of this study was to analyze the transcriptome of diverse Nrf2-deficient macrophage subpopulations from murine atherosclerotic aortas. Mice with transcriptionally inactive Nrf2 in Cdh5-expressing cells (Nrf2 Cdh5tKO) were used in the experiments. These mice lack transcriptional Nrf2 activity in endothelial cells, but also in a proportion of leukocytes. We confirmed that the bone marrow-derived and tissue-resident macrophages isolated from Nrf2 Cdh5tKO mice exhibit a significant decline in Nrf2 activity. Atherosclerosis was induced in Nrf2 Cdh5tKO and appropriate control mice via adeno-associated viral vector (AAV)-mediated overexpression of murine proprotein convertase subtilisin/kexin type 9 (Pcsk9) in the liver and high-fat diet feeding. After 21 weeks, live aortic cells were sorted on FACS and single-cell RNA sequencing (scRNA-seq) was performed. Unsupervised clustering singled out 13 distinct aortic cell types. Among macrophages, 9 subclusters were identified. Differential gene expression analysis revealed cell subtype-specific expression patterns. A subset of inflammatory macrophages from atherosclerotic Nrf2 Cdh5tKO mice demonstrated downregulation of DNA replication genes (e.g. Mcm7, Lig1, Pola1) concomitant with upregulation of DNA damage sensor Atr gene. Atherosclerotic Nrf2 Cdh5tKO Lyve1+ resident macrophages showed strong upregulation of IFN-stimulated genes, as well as changes in the expression of death pathways-associated genes (Slc40a1, Bcl2a1). Furthermore, we observed subtype-specific expression of core ferroptosis genes (e.g. Cp, Hells, Slc40a1) in inflammatory versus tissue resident macrophages. This observation suggested a link between ferroptosis and inflammatory microenvironment appearing at a very early stage of atherogenesis. Our findings indicate that Nrf2 deficiency in aortic macrophages leads to subtype-specific transcriptomic changes associated with inflammation, iron homeostasis, cell injury or death pathways. This may help understanding the role of aging-associated decline of Nrf2 activity and the function of specific macrophage subtypes in atherosclerotic lesion development.

Hematological Evaluation of Three Common Teleosts in Relation to The Environmental Changes from Trang Province, Thailand.

Hematological evaluation of fish is essential to the assessment of their physiological status. This study describes the morphometric analysis and comparison of blood cell characteristics in Zanarchopterus sp., Gerres filamentosus Cuvier, 1829 and Leiognathus decorus (De Vis, 1884). The species were collected at two locations off the coast of Trang Province, Thailand. A comparative hematological evaluation was made to assess the effects of environmental conditions on the blood of the fish. Ten individuals of each species were collected from a seagrass bed at Libong Island, where human activities are increasing, and from a secluded sandy beach. Their blood samples were analysed using the blood smear technique. Erythrocytes of all the studied fishes were either elliptical or oval. The morphometric data from both locations showed that erythrocytes were of similar size, except for those of Zanarchopterus sp. Fish from both stations showed several types of leukocytes, including neutrophils and lymphocytes.The highest proportion of leukocytes was made up of lymphocytes, followed by neutrophils. However, monocytes were only observed in fish from Libong Island and the erythrocytic nuclei of fish collected from Libong Island were both reniform and lobate. Our results show the potential of hematological evaluation as an early warning signal of environmental impacts on aquatic animals. The determination of baseline parameters could provide a tool for the monitoring of environmental quality.