Mean Cell Hemoglobin Concentration

About 3 ml of UCB specimen were obtained from each study participants after delivery from the clamped umbilical cord. The two trend Midwifery professionals collected the cord blood sample from the clamped cord through excluding of the placenta. The collected sample was immediately poured into tri-potassium ethylene diamine tetra acetic acid (K3-EDTA) test tube and gently mixed to prevent blood clotting. In addition, 3 ml of venous blood was collected from the mother after delivery with a sterile and disposable syringe. Hematological parameters: total white cell count (WBC), differential white cell count (neutrophils, lymphocytes and mixed which contains eosinophils, monocytes. and basophiles), platelet count, red blood cell count (RBC), Hgb, hematocrit (%), mean cell volume (MCV), mean cell hemoglobin (MCH), mean cell hemoglobin concentration (MCHC), and red cell distribution width (RDW) were determined by using the using Sysmex KX-21N (Sysmex Corporation Kobe, Japan) automated hematological whole blood analyzer based on direct current principle. Three experienced laboratory technologist performs the complete blood count (CBC) by strictly adhering standard operating procedures.

Multi-trait colocalization was performed using the Hypothesis Prioritization Colocalization (HyPrColoc) package (https://github.com/jrs95/hyprcoloc). This package performs multi-trait colocalization in a similar way to moloc, the multi-trait extension to coloc^{52 (link)}, but in a computationally efficient way that allows colocalization of large numbers of traits to be performed. We investigated colocalization between six traits: HDL-cholesterol, triglycerides, CHD risk, mean corpuscular haemoglobin concentration, platelet distribution width, and red cell distribution width. These blood cell traits were selected as variants associated with these traits have the greatest mean posterior probability of belonging to the largest group of variants identified by the contamination mixture method (Supplementary Table 4). Associations with the blood cell traits were estimated in 173,480 unrelated European-descent individuals from the UK Biobank and INTERVAL studies^{53 (link)}. For each gene region, we took all available variants from the relevant recombination window around the gene^{54 (link)}. Colocalization was performed using default settings for the priors in the hyprcoloc function (prior probability of initial trait association 0.0001, conditional probability of subsequent trait having shared association 0.02), and with the uniform priors setting as the default setting can be overly conservative.
While the exact pattern of colocalization differed between the gene regions, colocalization between HDL-cholesterol, CHD risk, and at least one blood cell trait was observed for 3 of the gene regions using the conservative priors, and for 7 regions using uniform priors (Supplementary Table 5). The posterior probability of colocalization was at least 0.7 in all cases, except when using conservative priors in the C5orf67 gene region. For this region, there was evidence of colocalization between HDL-cholesterol, triglycerides, CHD risk, and mean corpuscular haemoglobin concentration at posterior probability 0.59, and evidence of colocalization between HDL-cholesterol, triglycerides, and mean corpuscular haemoglobin concentration only (excluding CHD risk) at posterior probability 0.96. For the two gene regions that did not show evidence of colocalization between these traits, one possible explanation is the presence of multiple causal variants in the region; as the ATXN2 gene region reported colocalization between HDL-cholesterol and CHD risk, and separately between the blood cell traits. For COBLL1, there was colocalization between HDL-cholesterol and the blood cell traits, but not CHD risk.
As this investigation only uses publicly available summarised data on genetic associations with traits and diseases, no specific ethical approval is required.

At the end of the study period (12th week), 24 birds [six birds from each group (1 per replicate)] were selected for sample collection after 12 h fasting. About 5 ml of blood were collected from the jugular veins, then kept in a micro-anticoagulant tube in slanting position for 30 min, centrifuged at 1,500 x g for 15 min (Tang et al., 2017 (link)). The harvested plasma was transferred to Eppendorf tubes (1.5 ml) and stored at –20°C until analysis. The whole blood samples were put together in an ice pack and transported to the laboratory for hematology analysis within 1 h of collection. For hematological indices analysis, an automated hematology analyzer (Model: BC-2800 Vet, Mindray, Shenzhen, China) was used. Red blood cell indices, MCH, MCV and mean cell hemoglobin concentration (MCHC), were calculated (Jain, 1993 ).
Before analysis of biochemical indices, the serum was thawed at 4°C and kept at low temperature during the whole process in order to avoid activation of enzymes. Determination of glutathione peroxidase (GST), glutathione peroxidase (GSH-Px), total antioxidant capacity (T-AOC), total superoxide dismutase (T-SOD), catalase (CAT), and malondialdehyde (MDA) were achieved using an assay kits from ML Bio and Jiancheng Bioengineering Institute (Nanjing, China), and measured spectrophotometrically (Shimadzu, model UV-1800, Tokyo, Japan). Serum concentrations of immunoglobulins A (IgA), immunoglobulins G (IgG), immunoglobulins M (IgM) and complement proteins; C3 and C4 were determined with the corresponding chicken ELISA kits. All standards were tested in duplicate and concentrations of IgA, IgG, IgM, C3, and C4 were determined using standard curves constructed from the standards run on the plate. All the ELISA kits adopted in the study are of high specificity and sensitivity for chickens, and all procedures were done according to the manufacturer’s instructions.

A complete blood count (CBC) was performed as a broad screening test in order to support the assessment of each dog’s health status. This screening included: haematocrit, haemoglobin, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentration, platelet count, red cell distribution width, red blood cell count and white blood cell count. The CBCs were performed by the Diagnostic Laboratory, Department of Clinical Studies, School of Veterinary Medicine, Aristotle University of Thessaloniki, Greece.

The candidate features were collected from the electronic medical record, including age, sex, BMI (BMI = weight/height²), systolic blood pressure (SP), diastolic blood pressure (DP) and blood testing.
The blood testing was as following:(1) complete blood count: WBC, RBC, HGB, HCT, MCV, MCH, mean corpuscular hemoglobin concentration (MCHC), PLT, standard deviation in red cell distribution width (RDW-SD), coefficient variation of red cell volume distribution width (RDW-CV), platelet distribution width (PDW), MPV, P-LCR, PCT, NE, LY, MONO, EO, basophil count (BA), neutrophil ratio (NE%), lymphocyte ratio (LY%), MONO%, EO%, basophil ratio (BA%); (2) liver function: total bilirubin (TBIL), direct bilirubin (DBIL), indirect bilirubin (IDBIL), ALT, AST, ratio of AST/ALT, γ-GT, alkaline phosphatase (ALP), TP, albumin (ALB), GLO, ratio of ALB/GLO; (3) lipid panel: TG, TC, HDL, LDL, VLDL.