Iron-related markers of inflammation in horses with colic.

The aim of the study was to compare and correlate levels of ferritin, transferrin, iron and APPs in healthy horses and those surgically treated for strangulating colic. On admission, measurements of inflammatory markers related to iron and total protein, fibrinogen, albumin, haptoglobin and ceruloplasmin were made. The study comprised 22 horses, divided into a control group (CG) of healthy horses (n = 10) and horses with surgically treated acute abdomen (n = 12), obstruction group (OG). The OG was subdivided according to the affected intestinal segment (small vs. large) and according to outcome (survivors vs. non survivors). The OG had higher haptoglobin (34.8±14.2 mg/dL vs 20.8±7.21 mg/dL) and transferrin (487±161 mg/dL vs 369±71.4 mg/dL) values and lower iron (96.9±65 µg/dL vs 218±105 µg/dL) values than the CG. The OG horses with large intestine obstruction had lower values of transferrin (374.6±130 mg/dL) than horses with small intestinal obstruction (598.6±98.9 mg/dL). There was no difference in outcome between horses with large and small intestinal obstruction. Ferritin levels were moderately correlated with total protein (r = 0.594; P = 0.042) and albumin (r = 0.584; P = 0.046) in OG. In the multivariate exploratory analysis, fibrinogen levels were higher in animals that did not survive. In conclusion, haptoglobin, transferrin and iron were useful inflammatory markers for colic in horses. The correlation of ferritin with other APPs shows a possible role of ferritin as an APP in horses. Fibrinogen levels are higher in horses with greater risk of death from strangulating obstructions.

Effects of offering free-choice hay for the first 5 days postpartum on productivity, serum inflammatory markers, gut permeability, and colon gene expression in fresh dairy cows.

The objective of the present study was to evaluate the effects of offering free-choice hay to cows during the first 5 d immediately after calving on feed intake, milk yield, plasma metabolites, serum inflammatory markers, rumination, gut permeability, and colon gene expression. It was hypothesized that cows offered free-choice hay would have lower gut permeability, lower inflammation, and higher milk production, compared with cows not offered hay. Thirty-two multiparous cows were fed a closeup total mixed ration (TMR; 21.5% starch, 32.1% forage neutral detergent fiber [NDF] on a dry matter basis) until calving. In the postpartum period, all cows were fed a fresh cow TMR (26.8% starch and 23.4% forage NDF) from calving until 21 DIM, and were assigned randomly to receive 1 of 2 treatments as follows: (1) free-choice timothy hay (61.6% NDF; 9.6% crude protein), offered outside of the TMR in a separate manger, for the first 5 d postpartum (FCH; n = 20), or 2) no free-choice hay (NH; n = 12). The FCH cows tended to have lower serum haptoglobin concentration on d 3, compared with NH (0.95 vs. 1.52 mg/mL). Within the FCH group, cows with greater hay intake had a smaller increase in serum amyloid A from d 1 to 3 after calving (r = 0.37), and tended to have a smaller increase in serum haptoglobin concentration (r = 0.36). Cows in the FCH group had a lower ratio of starch intake (kg) to forage NDF intake (kg) on d 1 and 2, compared with NH (0.91 vs. 1.14 ± 0.03), and cows that had a lower starch:forage NDF ratio tended to have a smaller increase in serum haptoglobin concentration from d 1 to 3 after calving (r = 0.32). Cows in the FCH group had lower TMR dry matter intake (DMI; 15.0 vs. 17.1 ± 0.93 kg/d) and lower total DMI (TMR + hay DMI; 15.9 vs. 17.1 ± 0.87 kg/d), from d 1 to 5 when free-choice hay was offered, compared with NH. However, the hay treatment did not affect plasma energy metabolite concentration, gut permeability, colon gene expression, milk yield, rumination time, or change in body weight or body condition score. Overall, these findings suggest that offering free-choice hay for the first 5 d after calving may reduce serum inflammatory marker concentration, but milk yield may not increase, due to lower intake.

Associations between serum health biomarker concentrations and reproductive performance, accounting for milk yield, in pasture-based Holstein cows in southeastern Australia.

In this single cohort study, we investigated associations between the concentrations of a suite of serum biomarkers measured in the first 30 d of lactation and subsequent reproductive performance measured as mating start date to conception intervals, in pasture-based Holstein cows. A secondary objective was to examine associations between biomarker concentrations and 305-d milk yield to assess whether any positive associations between biomarker concentration and reproductive performance were explained by reduced milk production. The data used had been collected as part of an ongoing project from 2017 to 2020 to compile a data set from a large population of lactating dairy cows. Biomarkers measured were those associated with energy balance (ß-hydroxybutyrate [BHB] and nonesterified fatty acids [NEFA]), protein nutritional status (urea and albumin), immune status (globulin, albumin to globulin ratio and haptoglobin), and macromineral status (calcium and magnesium). Associations between biomarker concentrations and mating start date to conception interval were investigated using Cox proportional hazard models, using between 634 and 1,121 lactations (varying by biomarker) from 632 to 1,103 cows and 11 to 17 mating periods from 10 to 13 herds. Based on hazard ratio (HR) estimates and associated 95% confidence intervals (CI), hazard of conception on any particular day of the herds' mating periods was positively associated with the concentrations of albumin (HR = 1.09; 95% CI: 1.05-1.12), albumin to globulin ratio (HR = 2.82; 95% CI: 1.66-4.79), calcium (HR = 2.01; 95% CI: 1.18-3.43), and magnesium (HR = 2.17; 95% CI: 1.01-4.66), and negatively associated with globulin concentration (HR = 0.98; 95% CI: 0.97 to 1.00). There was also some evidence that NEFA concentration was negatively associated (HR = 0.76; 95% CI: 0.57 to 1.01), and urea concentration positively associated (HR = 1.05; 95% CI: 0.99 to 1.11), with reproductive performance, but no evidence that BHB and haptoglobin concentrations were associated with reproductive performance. Except for NEFA, presence and direction of the associations between the biomarker and milk yield were not discordant with that for reproductive performance. Also, except for NEFA, we found no substantial evidence of nonlinear relationships between biomarker concentration and either reproductive performance or milk yield. Correlations between biomarker concentrations were generally weak, indicating that multibiomarker panels may collectively predict reproductive performance better than any single biomarker. We noted substantial variation in the concentrations of all biomarkers within, and for some biomarkers, between herd-year groups. Collectively, these results indicate that there may be scope to improve biomarker concentrations through nutritional, management, and genetic interventions, and by association, reproductive performance and milk yield may also improve.

Observational study on the associations between milk yield, composition, and coagulation properties with blood biomarkers of health in Holstein cows.

The considerable increase in the production capacity of individual cows owing to both selective breeding and innovations in the dairy sector has posed challenges to management practices in terms of maintaining the nutritional and metabolic health status of dairy cows. In this observational study, we investigated the associations between milk yield, composition, and technological traits and a set of 21 blood biomarkers related to energy metabolism, liver function or hepatic damage, oxidative stress, and inflammation or innate immunity in a population of 1,369 high-yielding Holstein-Friesian dairy cows. The milk traits investigated in this study included 4 production traits (milk yield, fat yield, protein yield, daily milk energy output), 5 traits related to milk composition (fat, protein, casein, and lactose percentages and urea), 11 milk technological traits (5 milk coagulation properties and 6 curd-firming traits). All milk traits (i.e., production, composition, and technological traits) were analyzed according to a linear mixed model that included the days in milk, the parity order, and the blood metabolites (tested one at a time) as fixed effects and the herd and date of sampling as random effects. Our findings revealed that milk yield and daily milk energy output were positively and linearly associated with total cholesterol, nonesterified fatty acids, urea, aspartate aminotransferase, γ-glutamyl transferase, total bilirubin, albumin, and ferric-reducing antioxidant power, whereas they were negatively associated with glucose, creatinine, alkaline phosphatase, total reactive oxygen metabolites, and proinflammatory proteins (ceruloplasmin, haptoglobin, and myeloperoxidase). Regarding composition traits, the protein percentage was negatively associated with nonesterified fatty acids and ß-hydroxybutyrate (BHB), while the fat percentage was positively associated with BHB, and negatively associated with paraoxonase. Moreover, we found that the lactose percentage increased with increasing cholesterol and albumin and decreased with increasing ceruloplasmin, haptoglobin, and myeloperoxidase. Milk urea increased with an increase in cholesterol, blood urea, nonesterified fatty acids, and BHB, and decreased with an increase in proinflammatory proteins. Finally, no association was found between the blood metabolites and milk coagulation properties and curd-firming traits. In conclusion, this study showed that variations in blood metabolites had strong associations with milk productivity traits, the lactose percentage, and milk urea, but no relationships with technological traits of milk. Specifically, increasing levels of proinflammatory and oxidative stress metabolites, such as ceruloplasmin, haptoglobin, myeloperoxidase, and total reactive oxygen metabolites, were shown to be associated with reductions in milk yield, daily milk energy output, lactose percentage, and milk urea. These results highlight the close connection between the metabolic and innate immunity status and production performance. This connection is not limited to specific clinical diseases or to the transition phase but manifests throughout the entire lactation. These outcomes emphasize the importance of identifying cows with subacute inflammatory and oxidative stress as a means of reducing metabolic impairments and avoiding milk fluctuations.

Relationships between haptoglobin and inter-alpha trypsin inhibitor heavy chain 4 levels in local and peripheral blood and systemic inflammatory response syndrome in bitches with pyometra.

We aimed to assess the usefulness of monitoring inter-alpha trypsin inhibitor heavy chain 4 (ITIH4) and haptoglobin (Hp) in peripheral and local blood in canine pyometra, and evaluation the relationships among acute phase proteins (APPs), systemic inflammatory response syndrome (SIRS) and the presence of bacteria. The material was collected from bitches with pyometra and from healthy ones. Blood was taken from the cephalic and uterine veins. APPs levels were quantified by ELISA. In the peripheral circulation, the Hp was higher in animals with open-cervix pyometra (OCP) than in the closed-cervix pyometra (CCP) and the control group. The Hp concentration was not correlated with age, with the presence of SIRS or with the type of bacteria (Gram-negative, Gram-positive or mixed flora). The ITIH4 concentrations in the peripheral blood did not differ significantly in the cases of pyometra. The Hp concentration in the local circulation increased in the OCP but not in the CCP groups, although the histopathological changes in the endometrium were similar. Peripheral Hp concentrations may be a useful tool in differentiating between the types of pyometra.

Haptoglobin 2-2 Genotype is Related to the Severity of Liver Damage in Hepatitis B Patients with Liver Steatosis.

BACKGROUND: The Haptoglobin (Hp) genotypes have been linked to immune diseases and play a significant role in metabolic diseases. This study aimed to analyze the correlation between Hp gene polymorphism and the severity of hepatitis B accompanied by liver steatosis. METHODS: A total of 182 with Hepatitis B and concurrent hepatic steatosis were included in the study. Clinical biochemical indices for each participant were recorded. DNA was extracted from peripheral blood leukocytes for globin genotyping. Of these participants, 128 underwent biopsy from which histological data were collected. RESULTS: Subjects with hepatitis B and hepatic steatosis carrying the Hp 2-2 genotype exhibited elevated alanine transaminase (ALT), c-glutamyl transferase (GGT), and aspartate amino transferase (AST) levels. In contrast, high-density lipoprotein (HDL) levels and the copy number of Hepatitis B Virus (HBV)-DNA were significantly reduced in those with the Hp 2-2 genotype (p < 0.05). Furthermore, individuals processing the Hp 2-2 genotype demonstrated a heightened hepatitis score and advanced fibrosis stage (p < 0.05). Notably, the Hp 2-2 genotype was independently associated with increased inflammation (odds ratio (OR) = 7.059, p < 0.001) and progressive fibrosis (OR = 3.05, p < 0.022). CONCLUSIONS: The Hp 2-2 genotype is significantly associated with increased severity in cases of hepatitis B with coexisting hepatic steatosis.

The Relationships between Intestinal Permeability and Target Antibodies for a Spectrum of Autoimmune Diseases.

The worldwide prevalence of autoimmune diseases that have limited treatment options and preventive strategies is rapidly rising. There is growing evidence that the microbiota and the integrity of the intestinal barrier play a role in autoimmune diseases. The potential to evaluate intestinal barrier integrity for susceptible individuals and to determine whether restoring intestinal junction integrity impacts autoimmune diseases is an important area of research that requires further attention. In the intestinal permeability model of autoimmune diseases, the breakdown of the intestinal tight junction proteins (zonulin/occludin) allows bacteria, toxins, undigested dietary proteins, and other antigens to pass into the lumen, thereby increasing the number of inflammatory reactions and the activation of immune cells throughout the body. In this study, we investigate the relationship between zonulin/occludin antibodies, which are used to determine intestinal permeability, with autoantibodies used to diagnose autoimmunity. Our investigation may identify significant levels of circulating autoantibodies in human subjects with intestinal permeability compared to those without intestinal permeability. Furthermore, we identified that significant positive linear correlations between serum occludin/zonulin antibodies and circulating autoantibodies could be used to determine autoimmune diseases.

Acute phase protein concentration in pharyngeal swabs from clinically healthy commercial dairy calves.

BACKGROUND: Early diagnosis of disease in calves is crucial for fast recovery and prudent use of antibiotics. The serum concentration of acute phase proteins (APPs) is up- or downregulated in response to tissue injury and has been studied widely in human medicine. There is growing interest in using APPs as biomarkers for different diseases and as a tool to initiate and monitor treatment in veterinary medicine as well. The concentration of APPs in saliva in healthy calves has not been established and the use of pharyngeal swabs offers a non-invasive alternative to blood sampling. Pharyngeal swabs, tracheal aspirate (TA) and blood samples were collected from 84 clinically healthy commercial dairy calves and analyzed for the APPs serum amyloid A (SAA), haptoglobin (Hp) and lipopolysaccharide binding protein (LBP). RESULTS: We found detectable concentrations of SAA, Hp and LBP in pharyngeal swabs from calves, as well as in TA and serum. There were no biologically interesting correlations between the SAA concentrations in serum and TA or pharyngeal swabs. This also applied to Hp and LBP concentrations in serum and TA or pharyngeal swabs. CONCLUSIONS: SAA, Hp and LBP can be measured in saliva and TA from calves, but there was no correlation between the specific APP concentration in serum and pharyngeal swab or TA. There was a considerable technical variation in the sampling method for both pharyngeal swab and TA, and further validation of the methods is needed.

A review on acute phase response in parasitic blood diseases of ruminants.

Parasitic blood diseases (theileriosis, babesiosis, anaplasmosis, and trypanosomiasis) are common in regions where the distributions of the hosts, parasites, and vectors are convergent. They endanger animal production, and a few are also harmful to public health. The acute phase reaction (APR) is a complex, non-specific reaction that occurs in various events, including surgical trauma, infection, stress, inflammation, and neoplasia. To understand pathogenesis, we must study APR effects and acute phase proteins (APPs) alterations in naturally occurring and experimental infections. The elevation of haptoglobin (Hp), Serum amyloid A (SAA), and fibrinogen concentrations was markedly significant in bovine and ovine theileriosis. Hp, SAA, ceruloplasmin, and fibrinogen concentrations in anaplasmosis were dramatically elevated. A significant increase in SAA was observed in bovine babesiosis, while ovine babesiosis showed a significant rise in sialic acid levels. In cases of trypanosomiasis caused by T. vivax, there have been reports of elevated levels of Hp, complement C3, and antitrypsin. Improving our understanding of APR could result in more effective methods for diagnosis, treatment, control, and eradication of diseases. The article provides an overview of APPs alterations and other inflammation-related parameters (some cytokines, adenosine deaminase, and sialic acids) in parasitic blood diseases of ruminants.

Prognostic significance of serum fucosylated pro-haptoglobin in advanced renal cell carcinoma patients treated with immune checkpoint inhibitors.

With the widespread use of immune checkpoint inhibitors (ICIs), identifying predictive biomarkers is critical. Recently, serum fucosylated haptoglobin (Fuc-Hp) was thought to play an important role in tumour immunity in several types of cancer. Therefore, evaluating serum Fuc-Hp in the peripheral blood can potentially identify non-invasive predictive biomarkers for the clinical efficacy of ICIs. In this study, 31 patients with advanced renal cell carcinoma (RCC) treated with nivolumab were enrolled and defined as responders or non-responders according to RECIST criteria. Serum samples were collected before and 1 month after treatment initiation, and an ELISA assay was performed using Aleuria Aurantia Lectin (AAL) and 10-7G monoclonal antibodies that recognise Fuc-mature Hp (Fuc-mHp) and Fuc-pro Hp (Fuc-pHp), respectively. We first measured AAL-haptoglobin (Fuc-mHp) and total haptoglobin levels before nivolumab and found that neither value could predict the clinical response. Notably, serum 10-7G levels were significantly lower in the responder group (p = 0.035). We also confirmed the use of serum 10-7G levels for predicting progressive disease after nivolumab (area under the curve, 0.816). Accordingly, low 10-7G levels were significantly correlated with better progression-free survival (p = 0.041). In conclusion, serum Fuc-pHp analysis may identify patients with advanced RCC who benefit from ICIs.

Myeloid cell iron uptake pathways and paramagnetic rim formation in multiple sclerosis.

In multiple sclerosis (MS), sustained inflammatory activity can be visualized by iron-sensitive magnetic resonance imaging (MRI) at the edges of chronic lesions. These paramagnetic rim lesions (PRLs) are associated with clinical worsening, although the cell type-specific and molecular pathways of iron uptake and metabolism are not well known. We studied two postmortem cohorts: an exploratory formalin-fixed paraffin-embedded (FFPE) tissue cohort of 18 controls and 24 MS cases and a confirmatory snap-frozen cohort of 6 controls and 14 MS cases. Besides myelin and non-heme iron imaging, the haptoglobin-hemoglobin scavenger receptor CD163, the iron-metabolizing markers HMOX1 and HAMP as well as immune-related markers P2RY12, CD68, C1QA and IL10 were visualized in myeloid cell (MC) subtypes at RNA and protein levels across different MS lesion areas. In addition, we studied PRLs in vivo in a cohort of 98 people with MS (pwMS) via iron-sensitive 3 T MRI and haptoglobin genotyping by PCR. CSF samples were available from 38 pwMS for soluble CD163 (sCD163) protein level measurements by ELISA. In postmortem tissues, we observed that iron uptake was linked to rim-associated C1QA-expressing MC subtypes, characterized by upregulation of CD163, HMOX1, HAMP and, conversely, downregulation of P2RY12. We found that pwMS with [Formula: see text] 4 PRLs had higher sCD163 levels in the CSF than pwMS with [Formula: see text] 3 PRLs with sCD163 correlating with the number of PRLs. The number of PRLs was associated with clinical worsening but not with age, sex or haptoglobin genotype of pwMS. However, pwMS with Hp2-1/Hp2-2 haplotypes had higher clinical disability scores than pwMS with Hp1-1. In summary, we observed upregulation of the CD163-HMOX1-HAMP axis in MC subtypes at chronic active lesion rims, suggesting haptoglobin-bound hemoglobin but not transferrin-bound iron as a critical source for MC-associated iron uptake in MS. The correlation of CSF-associated sCD163 with PRL counts in MS highlights the relevance of CD163-mediated iron uptake via haptoglobin-bound hemoglobin. Also, while Hp haplotypes had no noticeable influence on PRL counts, pwMS carriers of a Hp2 allele might have a higher risk to experience clinical worsening.

Effect of pre- and postpartum supplementation of a pure glycerol product to dairy cows on feed intake, metabolic markers, and milk yield and components.

The objective of this study was to quantify the effects of supplementing transition dairy cows with a low inclusion dry glycerol product in the pre- and postpartum periods on feed intake, metabolic markers, and milk yield and components. Multiparous Holstein dairy cows (n = 60) were enrolled in a 2-by-2 factorial design study. Starting 21 d before expected parturition, cows individually received a dry cow diet with (1) 250 g/d glycerol product supplementation [66% pure glycerol (United States Pharmacopeia grade); GLY], or (2) no supplementation (CON) mixed to their total mixed ration. After parturition, cows, again, were individually assigned to either GLY, or (2) no supplementation (CON) to their partial mixed ration for the first 21 d in milk (DIM). Cows were milked by an automated milking system and offered a target of 5.4 kg DM/d pellet (23% of target total dry matter intake, DMI) in the automated milking system and followed for 42 d into lactation. Blood samples were collected 6.3 ± 3.47 d before calving for all blood measures and 3, 7, 10, and 14 DIM for analysis of glucose and ß-hydroxybutyrate, as well as 3 and 7 DIM for nonesterified fatty acids (NEFA) and haptoglobin. Initial dry cow body weight (BW), calf birth weight, previous 305-d milk, and month of parturition were used as covariates in the statistical model. Cows supplemented with GLY prepartum lost less BW and consumed more DMI pre- and postpartum, as well as had lower postpartum blood ß-hydroxybutyrate and NEFA concentrations compared with those fed the CON treatment prepartum. Cows supplemented with GLY postpartum had lesser DMI in the first 42 DIM than cows fed CON postpartum, but also had reduced blood NEFA concentrations, odds of a high haptoglobin test, odds of a low blood glucose test, and lesser preformed fatty acid concentrations and yields in their milk. Cows supplemented glycerol both pre- and postpartum lost the least total BW from -21 to 21 DIM. No treatment effects were detected for milk yield; however, cows receiving GLY postpartum had lower milk fat. Overall, glycerol supplementation during the transition period, particularly during the 21 d before calving, was associated with markers of improved metabolic status.

Fecal acute phase proteins in cats with chronic enteropathies.

BACKGROUND: Chronic enteropathies (CE) are common in cats and reliable biomarkers that can distinguish different causes and predict or monitor response to treatment are currently lacking. HYPOTHESIS: To evaluate certain acute phase proteins in feces that could potentially be used as biomarkers in cats with CE. ANIMALS: Twenty-eight cats with either inflammatory bowel disease (IBD; n = 13), food-responsive enteropathy (FRE; n = 3) or small cell gastrointestinal lymphoma (SCGL; n = 12) and 29 healthy control cats were prospectively enrolled. METHODS: Fecal concentrations of haptoglobin, alpha-1-acid-glycoprotein (AGP), pancreatitis-associated protein-1 (PAP-1), ceruloplasmin, and C-reactive protein (CRP) were measured using Spatial Proximity Analyte Reagent Capture Luminescence (SPARCL) immunoassays before and after initiation of treatment. Cats were treated with diet and/or prednisolone (IBD cats), plus chlorambucil (SCGL cats). RESULTS: Compared with controls, median fecal AGP concentrations were significantly lower (25.1 vs 1.8 µg/g; P = .003) and median fecal haptoglobin (0.17 vs 0.5 µg/g), PAP-1 (0.04 vs 0.4 µg/g) and ceruloplasmin (0.15 vs 4.2 µg/g) concentrations were significantly higher (P < .001) in cats with CE. Median fecal AGP concentrations were significantly lower (P = .01) in cats with IBD and FRE (0.6 µg/g) compared with cats with SCGL (10.75 µg/g). A significant reduction was found in CE cats after treatment for median fecal ceruloplasmin concentrations (6.36 vs 1.16 µg/g; P = .04). CONCLUSIONS: Fecal AGP concentration shows promise to differentiate cats with SCGL from cats with IBD and FRE. Fecal ceruloplasmin concentrations may be useful to objectively monitor response to treatment in cats with CE.

Association of time to metritis diagnosis with circulating concentration of metabolites, minerals, and haptoglobin in Jersey cows.

Our objective was to investigate the association of early metritis [EMET, diagnosed at <5 d in milk (DIM)] and late metritis (LMET, diagnosed at ≥5 DIM) with circulating concentrations of energy metabolites, minerals, and haptoglobin (Hp) throughout the first 14 d postpartum. A total of 379 purebred Jersey cows were enrolled in a prospective cohort study from a single herd in west Texas. Cows were examined for metritis using the Metricheck device (Simcro Ltd.) at 4, 7, and 10 DIM. Cows identified by farm employees as possible metritis cases were also evaluated for metritis. Blood samples were collected for analysis of concentrations of Ca, Mg, and glucose at DIM 1 through 5, 7, 10, and 14. Albumin, urea, fructosamine, free fatty acids (FFA), creatinine, and ß-hydroxybutyrate (BHB) were analyzed at DIM 3, 5, 7, 10, and 14, and Hp at DIM 1 through 5 and 7. Data were analyzed using the MIXED and PHREG procedures of SAS (SAS Institute Inc.). A series of mixed general linear models accounting for repeated measures were fitted to the data. The independent variables metritis [no metritis (NMET), EMET, and LMET], DIM of analyte assessment, and parity were forced in all models. Multivariable Cox proportional hazard models were built to assess the risk of pregnancy and culling within 150 DIM. The overall metritis incidence was 26.9% (EMET = 49; LMET = 53; NMET = 277). Average concentrations of glucose, Mg, and urea were not associated with metritis. The associations of Ca, creatinine, BHB, and fructosamine with metritis were dependent on the DIM of analyte assessment. Cows categorized as EMET and LMET had, on average, lower albumin and fructosamine compared with NMET cows. Both EMET and LMET cows had, on average, greater BHB than NMET cows. A greater FFA concentration was only observed in cows diagnosed with EMET compared with NMET cows (EMET = 0.58, LMET = 0.52, NMET = 0.48 mmol/L). Additionally, circulating Hp concentration was greater for LMET and EMET compared with NMET cows, and EMET cows had greater Hp compared with LMET cows (EMET = 1.15; LMET = 1.00; NMET = 0.84). In conclusion, several blood biomarkers were temporally associated with early versus late metritis diagnosis in postpartum Jersey cows. No meaningful differences were observed in production, reproduction, or culling between EMET and LMET cows. These results suggest that cows with EMET undergo a more severe degree of inflammation and negative energy balance compared with NMET cows.

Disease-Specific haptoglobin N-Glycosylation in inflammatory disorders between cancers and benign diseases of 3 types of female internal genital organs.

BACKGROUND: N-glycosylation of the haptoglobin is closely related to pathological states. This study aims to evaluate the association of glycosylation of disease-specific Hp (DSHp) ß chain with different pathological states of the cervix, uterus, and ovary to explore differences in their inflammatory responses and to screen potential biomarkers to distinguish cancer from benign diseases. METHODS: DSHp-ß chains of 1956 patients with cancers and benign diseases located in the cervix, uterus, and ovary organs were separated from serum immunoinflammatory-related protein complexes (IIRPCs). The N-glycopeptides from DSHp-ß chains were detected using mass spectrometry, followed by an analysis of machine learning algorithms. RESULTS: 55 N-glycopeptides at N207/N211, 19 at N241, and 21 at N184 glycosylation sites of DSHp for each sample were identified. Fucosylation and sialylation of DSHp in cervix, uterus, and ovary cancer were significantly increased compared to their corresponding benign diseases (p < 0.001). The cervix diagnostic model, a combination of G2N3F, G4NFS, G7N2F2S5, GS-N&GS-N, G2N2&G4N3FS, G7N2F2S5, G2S2&G-N, and GN2F&G2F at N207/N211 sites, G3NFS2 and G3NFS at N241site, G9N2S, G6N3F6, G4N3F5S, G4N3F4S2, and G6N3F4S at N184 site), has shown a good diagnostic capability to distinguish cancer from benign diseases, with the area under curve (AUC) of 0.912. The uterus diagnostic model including G4NFS, G2S2&G2S2, G3N2S2, GG5N2F5, G2&G3NFS, and G5N2F3S3 at N207/N211 sites, and G2NF3S2 at N184 site, with an AUC of 0.731. The ovary diagnostic model including G2N3F, GF2S-N &G2F3S2, G2S&G2, and G2S&G3NS at N207/N211 sites; G2S and G3NFS at N241 site, G6N3F4S at N184 site, with an AUC of 0.747. CONCLUSIONS: These findings provide insights into differences in organ-specific inflammatory responses of DSHp for different pathological states among the organs of the cervix, uterus, and ovary.

Prohaptoglobin is a possible prognostic biomarker for colorectal cancer.

BACKGROUND AND AIMS: Fucosylated haptoglobin is a novel glycan biomarker for colorectal and other cancers, while the significance of its precursor, prohaptoglobin (proHp), remains to be elucidated. In this study, we investigated whether proHp can be a colorectal cancer (CRC) biomarker and the biological functions of proHp in CRC using 10-7G, a monoclonal antibody recently developed in our laboratory. MATERIALS AND METHODS: Serum proHp level in 74 patients with CRC was semi-quantified by western blotting, and 5-year recurrence-free survival and overall survival were analyzed for groups stratified by proHp status (high vs. low). We also performed immunohistochemical analyses of 17 CRC tissue sections using 10-7G mAb. The biological functions of proHp were evaluated by overexpressing proHp in CRC cell lines. RESULTS: Serum proHp correlated with the clinical stage and poorer prognosis of CRC. In the primary CRC sections, immune cells were stained positive for 10-7G in ∼50% of the cases. Overexpression of proHp in HCT116 human CRC cells induced epithelial-mesenchymal transition-like changes and promoted cell migration in CRC cells. CONCLUSION: We provide evidence for the first time that proHp has potential as a prognostic biomarker for CRC and demonstrated specific biological activities of proHp.

Engineering Therapeutics to Detoxify Hemoglobin, Heme, and Iron.

Hemolysis (i.e., red blood cell lysis) can increase circulatory levels of cell-free hemoglobin (Hb) and its degradation by-products, namely heme (h) and iron (Fe). Under homeostasis, minor increases in these three hemolytic by-products (Hb/h/Fe) are rapidly scavenged and cleared by natural plasma proteins. Under certain pathophysiological conditions, scavenging systems become overwhelmed, leading to the accumulation of Hb/h/Fe in the circulation. Unfortunately, these species cause various side effects such as vasoconstriction, hypertension, and oxidative organ damage. Therefore, various therapeutics strategies are in development, ranging from supplementation with depleted plasma scavenger proteins to engineered biomimetic protein constructs capable of scavenging multiple hemolytic species. In this review, we briefly describe hemolysis and the characteristics of the major plasma-derived protein scavengers of Hb/h/Fe. Finally, we present novel engineering approaches designed to address the toxicity of these hemolytic by-products.

Effect of source and amount of rumen-protected choline on hepatic metabolism during induction of fatty liver in dairy cows.

Objectives were to determine the effect of supplementing increased amounts of rumen-protected choline (RPC) from sources with low (L, 28.8%) or high (H, 60.0%) concentration of choline chloride on hepatic metabolism when cows were subjected to feed restriction to develop fatty liver. It was hypothesized that increased supplementation of RPC reduces hepatic triacylglycerol and enhances glycogen concentrations. Pregnant, nonlactating multiparous Holstein cows (n = 110) at mean (± standard deviation) 232 ± 3.9 d of gestation were blocked by body condition (4.01 ± 0.52) and assigned to receive 0 (CON), 12.9 (L12.9 or H12.9), or 25.8 (L25.8 or H25.8) g/d of choline ion. Cows were fed for ad libitum intake on d 1 to 5 and restricted to 50% of the NEL required for maintenance and pregnancy from d 6 to 13. Intake of metabolizable methionine was maintained at 19 g/d during the feed restriction period by supplying rumen-protected methionine. Hepatic tissue was sampled on d 6 and 13 and analyzed for triacylglycerol, glycogen, and mRNA expression of genes involved in choline, glucose, and fatty acids metabolism, cell signaling, inflammation, autophagy, lipid droplet dynamics, lipophagy, and endoplasmic reticulum stress response. Blood was sampled and analyzed for concentrations of fatty acids, ß-hydroxybutyrate (BHB), glucose, triacylglycerol, total cholesterol, and haptoglobin. Orthogonal contrasts evaluated the effect of supplementing RPC [CON vs. (1/4·L12.9 + 1/4·L25.8 + 1/4·H12.9 + 1/4·H25.8)], source of RPC [(1/2·L12.9 + 1/2·L25.8) vs. (1/2·H12.9 + 1/2·H25.8)], amount of RPC [(1/2·L12.9 + 1/2·H12.9) vs. (1/2·L25.8 + 1/2·H25.8)], and interaction between source and amount [(1/2·L12.9 + 1/2·H25.8) vs. (1/2·H12.9 + 1/2·L25.8)]. Least squares means and standard error of the means are presented in sequence as CON, L12.9, L25.8, H12.9, H25.8. Supplementation of RPC reduced hepatic triacylglycerol (9.3 vs. 6.6 vs. 5.1 vs. 6.6 vs. 6.0 ± 0.6% as-is) and increased glycogen contents (1.8 vs. 2.6 vs. 3.6 vs. 3.1 vs. 4.1 ± 0.2% as-is) on d 13 of the experiment. Feeding RPC reduced serum haptoglobin (136.6 vs. 85.6 vs. 80.6 vs. 82.8 vs. 81.2 ± 4.6 µg/mL) during the feed restriction period; however, blood concentrations of fatty acids, BHB, glucose, triacylglycerol, and total cholesterol did not differ among treatments. During feed restriction, supplementation of RPC enhanced the mRNA expression of genes related to choline metabolism (BHMT), uptake of fatty acids (CD36), and autophagy (ATG3), and reduced the expression of a transcript associated with endoplasmic reticulum stress response (ERN1). An increase in the amount of choline ion from 12.9 to 25.8 g/d enhanced the mRNA expression of genes associated with synthesis and assembly of lipoproteins (APOB100), and inflammation (TNFA), whereas it reduced the expression of genes linked to gluconeogenesis (PC), oxidation of fatty acids (ACADM, MMUT), ketogenesis (ACAT1), and synthesis of antioxidants (SOD1) on d 13 of the experiment. Feeding RPC, independent of the product used, promoted lipotropic effects that reduced hepatic lipidosis in dairy cows.

Proteomic analysis of plasma to identify novel biomarkers for intra-amniotic infection and/or inflammation in preterm premature rupture of membranes.

To identify potential plasma biomarkers associated with microbial invasion of the amniotic cavity (MIAC) and/or intraamniotic inflammation (IAI) in women with preterm premature rupture of membranes (PPROM). This retrospective cohort study included 182 singleton pregnant women with PPROM (23-33 weeks) who underwent amniocentesis. Plasma samples; all subjects were chosen from these participants and were analyzed using label-free liquid chromatography-tandem mass spectrometry for proteome profiling using a nested case-control study design (cases with MIAC/IAI vs. non-MIAC/IAI controls [n = 9 each]). Three identified target molecules for MIAC/IAI were further verified by ELISA in the study cohort (n = 182). Shotgun proteomic analysis revealed 17 differentially expressed proteins (P < 0.05) in the plasma of MIAC/IAI cases. In particular, the levels of FCGR3A and haptoglobin, but not LRP1, were found to be increased in the plasma of patients with MIAC, IAI, and both MIAC/IAI compared with those without these conditions. Moreover, these differences remained significant after adjusting for gestational age at sampling. The area under the curves of plasma FCGR3A and haptoglobin ranged within 0.59-0.65 with respect to each of the three outcome measures. Plasma FCGR3A and haptoglobin were identified as potential independent biomarkers for less-invasively detecting MIAC/IAI in women with PPROM.

Induced endometritis in early lactation compromises production and reproduction in dairy cows.

Objectives of this experiment were to study the effect of infusing utero-pathogenic bacteria to induce endometrial inflammation on productive performance in early lactation and subsequent reproduction. Although endometritis is associated with perturbed reproduction, numerous factors may contribute to the observed association. It was hypothesized that induced endometrial inflammation, resulting in localized and systemic inflammatory responses, compromises production and reproduction. Holstein cows without clinical disease and with less than 18% polymorphonuclear leukocytes (PMN) in endometrial cytology on d 31 ± 3 postpartum had their estrous cycle synchronized. Cows were blocked by parity and genomic breeding value for cow conception rate and, within block, assigned randomly to remain as untreated controls (CON; n = 37) or to receive an intrauterine infusion of 5.19 × 108 cfu Escherichia coli and 4.34 × 108 cfu Trueperella pyogenes during the luteal phase to induce endometrial inflammation (INF; n = 48). Endometrial cytology was taken on d 2 and 7 after treatment to evaluate the proportion of PMN. Rectal temperature, dry matter intake, and yields of milk and components were measured in the first 7 d after treatment. Blood serum was analyzed for concentration of haptoglobin. Leukocytes were isolated from blood on d 2 and 7 after treatment and on d 19 after artificial insemination (AI) and mRNA was quantified for a select group of genes. Cows received AI and reproduction was followed for 300 d postpartum. Bacterial infusion induced endometrial inflammation with increased proportions of PMN in the endometrial cytology on d 2 (4.4 ± 0.7 vs. 26.3 ± 2.8%) and 7 (10.9 ± 1.7 vs. 17.4 ± 2.1%) after treatment, resulting in increased mean prevalence of subclinical endometritis (>10% PMN; 23.3 ± 6.3 vs. 80.9 ± 5.1%). Rectal temperature did not differ between CON and INF, but the concentration of haptoglobin in serum tended to increase in INF compared with CON (113 ± 14 vs. 150 ± 16 µg/mL). Induced endometrial inflammation reduced yields of milk (44.9 ± 0.8 vs. 41.6 ± 0.8 kg/d), protein (1.19 ± 0.03 vs. 1.12 ± 0.03 kg/d), and lactose (2.17 ± 0.04 vs. 2.03 ± 0.04 kg/d) and tended to reduce dry matter intake (20.7 ± 0.5 vs. 19.4 ± 0.6 kg/d) in the first 7 d after treatment. Indeed, the reduction in milk yield lasted 4 wk. However, treatment did not affect yields of energy-corrected milk or fat because treatment with INF increased the concentration of fat in milk (3.54 ± 0.10 vs. 3.84 ± 0.10%). Induced endometrial inflammation reduced pregnancy per AI at all inseminations (33.4 ± 5.1 vs. 21.6 ± 3.7%) and the hazard of pregnancy (0.61; 95% CI = 0.36-1.04), which extended the median days open by 24 d. Blood leukocytes from INF cows had increased mRNA expression of the pro-inflammatory gene IL1B on d 2 and 7 after treatment, but reduced expression of the IFN-stimulated genes ISG15 and MX2 on d 19 after AI. Induced endometrial inflammation depressed production and caused long-term negative effects on reproduction in lactating dairy cows.