Dynamics of lipid droplets in the endometrium and fatty acids and oxylipins in the uterine lumen, blood, and milk of lactating cows during diestrus.

Our objective was to investigate the lipid content of uterus, blood plasma, and milk at early, mid, and late diestrus. Lactating cows (n = 30) had the estrous cycle and ovulation synchronized by administration of exogenous hormones. Cows were blocked by parity and assigned randomly to receive transcervical uterine flushing and biopsy on d 5 (early diestrus), 10 (mid diestrus) or 15 (late diestrus) of the estrous cycle. Flushing and endometrial biopsy were performed in the uterine horn ipsilateral to the corpus luteum. The recovered flushing was used for analyses of lipid composition by liquid chromatography-tandem mass spectrometry and the biopsy was used for investigation of lipid droplet abundance in endometrial cryosections using a neutral lipid fluorescent dye. In addition, blood and milk samples were collected from all cows on d 5, 10, and 15. All blood samples were used to measure the concentration of progesterone in plasma, and all milk samples were used to determine milk composition. Subsamples of blood plasma and milk were also used to evaluate the composition of fatty acids and oxylipins using the same methodology used for uterine flushing samples. The abundance of lipid droplets in the endometrium increased 1.9-fold from d 5 to 10, and 2-fold from d 10 to 15. Concentration of long-chain fatty acids and oxylipins in uterine flushing were, on average, 2.2 and 2.5 times greater in samples collected on d 15 compared with those collected on d 5 and 10. These differences were not observed in blood and milk, suggesting that accumulation of fatty acids and oxylipins in the uterus is regulated locally. In addition to concentration, the profile of individual fatty acids and oxylipins in uterine lumen changed substantially during diestrus. The main categories with increased abundance at late diestrus were mono- and polyunsaturated fatty acids, and oxylipins derived from arachidonic acid, dihomo-γ-linolenic acid, and docosahexaenoic acid. In conclusion, fatty acids and oxylipins accumulate in the uterine lumen during diestrus and might work as a mechanism to supply these lipids to the developing conceptus at late diestrus, when the onset of elongation occurs and substantial synthesis of biomass and cell signaling by lipid mediators are required.

Associations between maternal characteristics and health, survival, and performance of dairy heifers from birth through first lactation.

The objective of this study was to investigate whether health, survival, and performance of dairy heifers from birth through first lactation are associated with parity and health status of their dams. Holstein heifers (n = 1,811) derived from artificial insemination were categorized as (1) daughters of primiparous cows that, consequently, were nonlactating heifers during gestation (Prim-NoL; n = 787); (2) daughters of multiparous cows that did not have any clinical diseases in the previous lactation (Mult-NoCD; n = 638); and (3) daughters of multiparous cows that had at least one clinical disease in the previous lactation (Mult-CD; n = 386). Clinical diseases of the multiparous dams included retained placenta, metritis, mastitis, lameness, and digestive and respiratory problems. Data collected for evaluation of daughters included genotypic and phenotypic characteristics at birth, morbidity, reproductive performance, and culling from birth through 305 d in milk of first lactation. Orthogonal contrasts were used to evaluate the effect of the parity of the dam (Prim-NoL vs. Mult-NoCD + Mult-CD) and the effect of clinical disease occurrence in the previous lactation among multiparous dams (Mult-NoCD vs. Mult-CD). Compared with daughters of multiparous cows, daughters of Prim-NoL were lighter at birth (36 vs. 41 kg), had greater genetic merit for production traits (e.g., genomic estimated breeding value for milk yield: 875 vs. 746 kg), were less likely to leave the herd (17 vs. 28%) and to lose pregnancy as a heifer (9 vs. 14%), calved earlier (703 vs. 711 d old), were less likely to have clinical diseases as a first lactation cow (30 vs. 37%), and had reduced performance in the first lactation when considering their genetic merit (e.g., 305-d yield of energy-corrected milk: 11,270 vs. 11,539 kg). Compared with daughters of Mult-NoCD, daughters of Mult-CD were less likely to have digestive problems as a heifer (17 vs. 27%) and clinical disease as a first lactation cow (32 vs. 42%), but were also more likely to leave the herd as a heifer (32 vs. 25%) even though genetic merit for production traits were similar (e.g., genomic estimated breeding value for milk: 744 vs. 749 kg). In conclusion, both parity and health status of the dam in the previous lactation were associated with morbidity, survival, and performance of their daughters from birth through 305 d in milk of the first lactation and might represent factors affecting developmental programming of dairy heifers in utero.

Long-term effects of postpartum clinical disease on milk production, reproduction, and culling of dairy cows.

Two retrospective studies examining data of 7,500 lactating cows from a single herd were performed with the objective of evaluating the long-term effects of clinical disease during the early postpartum period on milk production, reproduction, and culling of dairy cows through 305 days in milk (DIM). In the first study, data regarding health, milk production, reproduction, and culling of 5,085 cows were summarized. Cows were classified according to incidence of clinical problem (metritis, mastitis, lameness, digestive problem, or respiratory problem) during the first 21 DIM (ClinD21). During 305 d of lactation, cows that had ClinD21 produced, on average, 410 kg less milk, 17 kg less fat, and 12 kg less protein compared with cows that did not have ClinD21 (NoClinD21). Although the interval to first breeding was not different between groups of interest, pregnancy rate through 305 DIM was lower in cows that had ClinD21 [adjusted hazard ratio (AHR) = 0.81]. When individual breedings were analyzed, cows that had ClinD21 presented lower rates of pregnancy per breeding for breedings performed before 150 DIM, reduced numbers of calving per breeding for breedings performed before 200 DIM, and greater number of pregnancy losses for all breedings performed through 305 DIM. The rate of culling from calving through 305 DIM was higher in cows that had a single ClinD21 (AHR = 1.79) and in cows that had multiple ClinD21 (AHR = 3.06), which resulted in a greater proportion of cows leaving the herd by 305 DIM (NoClinD21 = 22.6%; single ClinD21 = 35.7%; multiple ClinD21 = 53.8%). In the second study, data regarding postpartum health and 305-d yields of milk, fat, and protein were collected from 2,415 primiparous cows that had genomic testing information. Genomic estimated breeding values (EBV) were used to predict 305-d yields of milk, fat, and protein. Genomic EBV and predicted yields of milk, fat, and protein did not differ between cows that had ClinD21 and those that did not have ClinD21. In contrast, the observed 305-d yields of milk, fat, and protein were reduced by 345, 10, and 10 kg, respectively, in cows that had ClinD21 compared with cows that did not have ClinD21. We conclude that clinical disease diagnosed and treated during the first 21 DIM has long-term effects on lactation performance, reproduction, and culling of dairy cows, which contribute to detrimental consequences of health problems on sustainability of dairy herds. Replication of our studies in multiple herds will be important to confirm our findings in a larger population.

Feed restriction to induce and meloxicam to mitigate potential systemic inflammation in dairy cows before calving.

Most dairy cows experience a transient decrease in feed intake in the 1 to 2 wk before calving, which has been associated with systemic inflammation (SI), indicated by increased blood haptoglobin (Hp) concentration. We aimed to characterize the association between prepartum decrease in feed intake and the onset of SI and, if present, the ability of meloxicam (MEL), a non-steroidal anti-inflammatory drug, to mitigate SI. Holstein cows (n = 45) were assigned to control (n = 13), feed restriction (FR) untreated (FR-U; n = 15), and FR treated with MEL (FR-T; n = 17) groups. Daily feed intake was measured from -22 d from expected parturition until 35 d postpartum. Control cows were fed ad libitum, whereas FR-U and FR-T cows were reduced to 60% of their average intake for 4 consecutive days (-15 to -12 d from expected calving). The FR-T cows received MEL (0.5 mg/kg of body weight) once daily for 4 consecutive days (-13 to -10 d from expected calving). Blood samples were collected -22, -15, -14, -13, -12, -10, -7, -5, -3, 0, 1, 3, 5, 7, 15, 22, and 35 d relative to calving to measure serum concentrations of total calcium, total protein, albumin, globulin, cholesterol, urea, glucose, gamma-glutamyl transferase, aspartate aminotransferase, glutamate dehydrogenase, ß-hydroxybutyrate, nonesterified fatty acids, Hp, and insulin-like growth factor-1. Serum concentrations of lipopolysaccharide-binding protein were measured -22, -15, -14, -13, -12, and -10 d from expected calving. Simplified glucose tolerance tests were performed on -15, -12, -5, 1, and 5 d relative to calving. Mixed linear regression models were used to assess the effects of FR and MEL on each metabolite. The interaction between treatment group and blood sampling day was forced into each model. All models accounted for body condition score, parity, and the cow as a random effect. Nonesterified fatty acids concentrations in both the FR-U and FR-T groups significantly increased from the second until the last day of FR. Feed restriction increased urea concentrations compared with the control group on -14 d but decreased urea concentrations on -10 d from expected calving. Control cows had greater ß-hydroxybutyrate concentrations compared with FR cows on 15, 21, and 35 d postpartum. For all other metabolites, no differences were found. This model of FR produced substantial fat mobilization but based on serum Hp and lipopolysaccharide-binding protein concentrations did not generate measurable SI; therefore, we were unable to evaluate the ability of MEL to mitigate SI.

Squalus bassi sp. nov., a new long-snouted spurdog (Chondrichthyes: Squaliformes: Squalidae) from the Agulhas Bank.

The long-snouted African spurdog Squalus bassi sp. nov. is described based on material collected from the outer shelf and upper continental slope off South Africa and Mozambique. Squalus bassi shares with S. mitsukurii, S. montalbani, S. chloroculus, S. grahami, S. griffini, S. edmundsi, S. quasimodo and S. lobularis a large snout with prenarial length greater than distance between nostrils and upper labial furrows, dermal denticles tricuspidate and rhomboid and elevated number of vertebrae. Squalus bassi can be distinguished from all its congeners by a combination of body and fin colouration, external morphometrics, vertebral counts and shape of dermal denticles. Similar long-snouted congeners from the Indo-Pacific region, including S. montalbani, S. edmundsi and S. lalannei are compared in detail with the new species. This new species has been misidentified as the Japanese S. mitsukurii and the Mediterranean S. blainvillei due to the lack of comparative morphological analyses. The validity of the nominal species S. mitsukurii in the south-eastern Atlantic Ocean and western Indian Ocean is also clarified herein, indicating it has a more restricted geographical distribution in the North Pacific Ocean.

Verification and characterization of chromosome duplication in haploid maize.

Doubled haploid technology has been used by various private companies. However, information regarding chromosome duplication methodologies, particularly those concerning techniques used to identify duplication in cells, is limited. Thus, we analyzed and characterized artificially doubled haploids using microsatellites molecular markers, pollen viability, and flow cytometry techniques. Evaluated material was obtained using two different chromosome duplication protocols in maize seeds considered haploids, resulting from the cross between the haploid inducer line KEMS and 4 hybrids (GNS 3225, GNS 3032, GNS 3264, and DKB 393). Fourteen days after duplication, plant samples were collected and assessed by flow cytometry. Further, the plants were transplanted to a field, and samples were collected for DNA analyses using microsatellite markers. The tassels were collected during anthesis for pollen viability analyses. Haploid, diploid, and mixoploid individuals were detected using flow cytometry, demonstrating that this technique was efficient for identifying doubled haploids. The microsatellites markers were also efficient for confirming the ploidies preselected by flow cytometry and for identifying homozygous individuals. Pollen viability showed a significant difference between the evaluated ploidies when the Alexander and propionic-carmin stains were used. The viability rates between the plodies analyzed show potential for fertilization.

Analysis of the stomatognathic system functions and mastication muscles in children affected by molar hypomineralization (MH): a cross-sectional study.

PURPOSE: To analyze the functions of the stomatognathic system in children with or without molar-incisor hypomineralization (MIH). METHODS: For this cross-sectional study, 72 children aged 6-12 years were recruited and divided in two groups: with MIH (G1) and without MIH (G2). T-SCAN was used to verify the distribution of occlusal contacts, gnathodynamometer to measure maximum molar bite force, and Iowa Oral Pressure Instrument (IOPI) to assess the strength of facial expression muscles. The t test and paired t test (p ≤ 0.05) were used for statistical comparisons. RESULTS: The molars affected by MIH exhibited lower distribution of occlusal forces (p < 0.001) and lower maximum molar bite force (p < 0.05) compared to the molars in the control group. However, there was no difference between the MIH-affected sides compared to the unaffected side, nor between the molars affected by MIH and their antagonists (p > 0.05). There were no differences in the forces of the facial expression muscles between the groups. CONCLUSIONS: These findings suggest that MIH significantly impacts occlusal force distribution and bite force, but not facial expression musculature.

Effects of 1 or 2 treatments with prostaglandin F2α on subclinical endometritis and fertility in lactating dairy cows inseminated by timed artificial insemination.

The objectives of the current study were to investigate the efficacy of PGF2α as a therapy to reduce the prevalence of subclinical endometritis and improve pregnancy per artificial insemination (P/AI) in cows subjected to a timed artificial insemination (AI) program. A total of 1,342 lactating Holstein dairy cows were allocated randomly at 25 ± 3 d in milk (DIM) to remain as untreated controls (control, n=454) or to receive a single PGF2α treatment at 39 ± 3 DIM (1PGF, n=474) or 2 treatments with PGF(α at 25 ± 3 and 39 ± 3 DIM (2PGF, n=414). All cows were enrolled in the double Ovsynch program at 48 ± 3 DIM and were inseminated at 75 ± 3 DIM. A subset of 357 cows had uterine samples collected for cytological examination at 25 ± 3, 32 ± 3, and 46 ± 3 DIM to determine the percentage of polymorphonuclear leukocytes (PMNL). Subclinical endometritis was defined by the presence of ≥ 5% PMNL. Vaginal discharge score was evaluated at 25 ± 3 DIM and used to define the prevalence of purulent vaginal discharge. Body condition score was assessed at 25 ± 3 DIM. Pregnancy was diagnosed 32 d after AI and reconfirmed 28 d later. At 32 ± 3 DIM, the prevalence of subclinical endometritis was reduced by treatment with PGF2α at 25 ± 3 DIM in 2PGF (control=23.5% vs. 1PGF=28.3% vs. 2PGF=16.7%); however, this benefit disappeared at 46 ± 3 DIM, and 14% of the cows remained with subclinical endometritis. One or 2 treatments with PGF2α did not influence P/AI on d 32 or 60 after timed AI, which averaged 39.9 and 35.2%. Similarly, treatment with PGF2α had no effect on pregnancy loss between 32 and 60 d of gestation (11.9%). Cows diagnosed with both purulent vaginal discharge and subclinical endometritis had the lowest P/AI and the highest pregnancy loss compared with those diagnosed with only 1 of the 2 diseases or compared with cows having no diagnosis of uterine diseases. Interestingly, subclinical endometritis depressed P/AI and increased pregnancy loss only when it persisted until 46 DIM. On d 32 after AI, cows not diagnosed with subclinical endometritis and those that resolved subclinical endometritis by 46 DIM had greater P/AI than those that remained with subclinical endometritis at 46 DIM (45.4 and 40.0 vs. 25.0%, respectively). Similar to P/AI, cows not diagnosed with subclinical endometritis and those that resolved subclinical endometritis by 46 DIM had less pregnancy loss than those with subclinical endometritis at 46 DIM (9.6 and 13.5 vs. 43.9%, respectively). One or 2 treatments with PGF2α before initiation of the timed AI program were unable to improve uterine health, P/AI, and maintenance of pregnancy in lactating dairy cows. Cows diagnosed with both purulent vaginal discharge and subclinical endometritis had the greatest depressions in measures of fertility at first AI, particularly when subclinical endometritis persisted in the early postpartum period.

A new tetra-primer ARMS-PCR for genotyping bovine kappa-casein polymorphisms.

Kappa-casein (κ-casein) is one of the most abundant milk proteins. Its main function is to avoid the aggregation of casein micelles, keeping them, and therefore calcium phosphate, in pockets in solution. In bovines, a κ-casein functional polymorphism has been associated with fat, calcium, and protein milk contents and faster curd contraction in cheese production. Quicker curd contraction reduces the loss of milk solids, enhancing cheese yield. This polymorphism induces a double amino acid substitution (Thr136Ile and Ala148Asp). The polymorphism is normally detected by PCR-RFLP, which is a laborious method. An interesting methodological alternative is the tetra-primer amplification refractory mutation system PCR (tetra-primer ARMS-PCR). A tetra-primer ARMS-PCR for the detection of this κ-casein polymorphism has been described. However, specificity was not achieved, probably due to problems with primer design. We developed a new tetra-primer ARMS-PCR for the detection of the κ-casein polymorphism. This new method was validated in a double-blind test, by comparison with the results obtained for 50 Guzerá bulls formerly genotyped by PCR-RFLP. This new method achieved 100% sensitivity and specificity. We conclude that this method is a useful, cost-efficient alternative for the detection of functional κ-casein polymorphisms.

A gene (RPGR) with homology to the RCC1 guanine nucleotide exchange factor is mutated in X-linked retinitis pigmentosa (RP3).

X-linked retinitis pigmentosa (xlRP) is a severe progressive retinal degeneration which affects about 1 in 25,000 of the population. The most common form of xlRP, RP3, has been localised to the interval between CYBB and OTC in Xp21.1 by linkage analysis and deletion mapping. Identification of microdeletions within this region has now led to the positional cloning of a gene, RPGR, that spans 60 kg of genomic DNA and is ubiquitously expressed. The predicted 90 kD protein contains in its N-terminal half a tandem repeat structure highly similar to RCC1 (regulator of chromosome condensation), suggesting an interaction with a small GTPase. The C-terminal half contains a domain, rich in acidic residues, and ends in a potential isoprenylation anchorage site. The two intragenic deletions, two nonsense and three missense mutations within conserved domains provide evidence that RPGR (retinitis pigmentosa GTPase regulator) is the RP3 gene.

Association of dental hypersensitivity and anxiety in children with molar-incisor hypomineralisation (MIH).

AIM: The objective of this study was to investigate whether dental hypersensitivity and dental fear were linked to the presence and severity of MIH. METHODS: For this cross-sectional study, 1830 students between the ages of 6 and 12 years were recruited from four randomly selected schools. The Children's Fear Survey Schedule-Dental Subscale questionnaire was used to assess dental anxiety and fear. The children's self-reported dental hypersensitivity resulting from MIH was evaluated using the Wong-Baker Facial Scale and the Visual Analog Scale (VAS). RESULTS: MIH was correlated with tooth hypersensitivity, particularly in severe cases. Dental fear was present in 17.4% of the children with MIH, but it was not associated with dental hypersensitivity, gender, or age. CONCLUSION: No association was found between dental fear and dental hypersensitivity in children with MIH.

Epidemiological, clinical and laboratory aspects of Angiostrongylus cantonensis infection: an integrative review.

This integrative literature review study analyzes the findings of the last 5 years of the parasite Angiostrongylus cantonensis. It is known that this nematode is found in the pulmonary arteries of rats, where it remains as a definitive host. From mice, the cycle spreads to animals, such as snails, which in contact with humans can trigger the disease. In humans, the parasite causes several neurological, abdominal manifestations and mainly meningitis. Based on the review of studies, its epidemiology shows worldwide distribution, although there are endemic cases for this parasite, such as asian countries. Laboratory findings generally showed altered CSF with turbidity, increased protein and eosinophilia, which generated meningeal signs in the patient, moreover MRI exams showed multiple alterations. Rare findings of the nematode in the eyeball, lung and signs of peritoneal inflammation were reported, which requires further studies to understand the whole pathophysiology. Finally, conservative treatment based on anthelmintics and anti-inflammatories brought good responses, although there are reports of deaths, which demonstrates the importance in the prevention and therapy of this disease.

Influence of Arg72Pro polymorphisms of TP53 on the response of buccal cells to radiotherapy.

Polymorphisms in the TP53 gene codon 72 (Arg72Pro) influence apoptosis induction and DNA damage repair. We evaluated how variants of protein p53 (p53Arg and p53Pro) affect cell death and DNA damage repair by analyzing the frequencies of karyorrhexis and micronuclei. There were significant differences in the frequency of karyorrhexis between the three p53 genotypes (Arg/Arg, Arg/Pro, and Pro/Pro), between samples taken before and after radiotherapy, and between patients and controls. The frequency of micronucleated cells increased significantly after radiotherapy. There were no significant differences in the micronucleus frequency in healthy tissues of these patients compared to controls, or in the comparisons between the three genotypes. We conclude that Arg72Pro polymorphism influences cell apoptotic capacity. This is the first study investigating karyorrhexis and micronuclei, as indicators of apoptosis after radiotherapy, and how these indicators are influenced by the TP53 polymorphism Arg72Pro.

Modulation of inflammation by anti-TNF α mAb-dendrimer nanoparticles loaded in tyramine-modified gellan gum hydrogels in a cartilage-on-a-chip model.

Rheumatoid arthritis (RA) is an autoimmune and chronic inflammatory disease characterized by joint inflammation. Since the inflammatory condition plays an important role in the disease process, it is important to develop and test new therapeutic approaches that specifically target and treat joint inflammation. In this study, a human 3D inflammatory cartilage-on-a-chip model was established to test the therapeutic efficacy of anti-TNFα mAb-CS/PAMAM dendrimer NPs loaded-Tyramine-Gellan Gum in the treatment of inflammation. The results showed that the proposed therapeutic approach applied to the human monocyte cell line (THP-1) and human chondrogenic primary cells (hCH) cell-based inflammation system revealed an anti-inflammatory capacity that increased over 14 days. It was also possible to observe that Coll type II was highly expressed by inflamed hCH upon the culture with anti-TNF α mAb-CS/PAMAM dendrimer NPs, indicating that the hCH cells were able maintain their biological function. The developed preclinical model allowed us to provide more robust data on the potential therapeutic effect of anti-TNF α mAb-CS/PAMAM dendrimer NPs loaded-Ty-GG hydrogel in a physiologically relevant model.

Variations in genotype-phenotype correlations in phenylketonuria patients.

Phenylalanine hydroxylase deficiency is a trait inherited in an autosomal recessive pattern; the associated phenotype varies considerably. This variation is mainly due to the considerable allelic heterogeneity in the phenylalanine hydroxylase enzyme locus. We examined the genotype-phenotype correlation in 54 phenylketonuria (PKU) patients from Minas Gerais, Brazil. Two systems were used. The first was a phenotype prediction system based on arbitrary values (AV) attributed to each mutation and the second was a correlation analysis. An AV was assigned to each mutation: AV = 1 for classical PKU mutation; AV = 2 for moderate PKU mutation; AV = 4 for mild PKU mutation, and AV = 8 for non-PKU hyperphenylalaninemia mutation. The observed phenotype for AV analysis was the clinical diagnosis established by the overloading phenylalanine test. Among the 51 PKU patients that we analyzed based on this trait, in 51% the predicted phenotype did not match the observed phenotype; the highest degree of concordance was found in patients with null/null genotypes. The genotype was observed to be a good predictor of the clinical course of the patients and significant correlations were found between phenylalanine values at first interview and predicted residual activity, genotype and arbitrary value sum.

Dendrimer nanoparticles for colorectal cancer applications.

Cancer nanotechnology is a prolific field of research, where nanotools are employed to diagnose and treat cancer with unprecedented precision. Targeted drug delivery is fundamental for more efficient cancer treatments. For this, nanoparticles have been extensively used during the past few years in order to improve the specificity, selectivity and controlled release of drug delivery. It holds potential in minimizing systemic toxicity through the development of functionalized particles for targeted treatment. Among all the type of nanoparticles, dendrimers display several advantages, which make them ideal candidates for improved and targeted drug delivery in cancer research. Dendrimers can transport large amounts of drug into specific areas. In addition, they can be employed for monitoring the progress of the treatment process, with an unprecedented theranostic capability. Special emphasis is given to colorectal cancer and to the preferred employed strategies for producing drug-loaded/functionalized NPs for cancer therapy in the past few years.

Effect of anti-inflammatory treatment on systemic inflammation, immune function, and endometrial health in postpartum dairy cows.

Systemic inflammation (SI) is increasingly studied in several species because it may be central in many metabolic disturbances and be a risk factor for clinical disease. This proof-of-concept study evaluated the effects of the anti-inflammatory drug meloxicam on markers of SI and energy metabolism, polymorphonuclear neutrophil (PMN) function, and endometritis in clinically healthy postpartum dairy cows. Cows received meloxicam (0.5 mg/kg of body weight; n = 20) once daily for 4 days (10-13 days postpartum) or were untreated (n = 22). Blood samples were collected -7, 1, 3, 5, 7, 10, 11, 12, 13, 14, 18, 21, 28, and 35 days relative to calving to measure serum concentrations of metabolic and inflammatory markers. Function of peripheral blood PMN were evaluated at 5, 10, 14, and 21, and proportion of PMN in endometrial cytology were performed at 5, 10, 14, 21, 28 and 35 days postpartum. Meloxicam decreased serum haptoglobin from the second until the last day of treatment, and improved indicators of energy metabolism (lesser ß-hydroxybutyrate and greater insulin-like growth factor-1 during treatment, and greater glucose at the end of treatment than control cows). This improved PMN function at 14 days postpartum, but the endometrial inflammatory status was not affected.

Technical note: A new and cost-effective method for detection of the bovine acyl-CoA:diacylglycerol acyltransferase 1 K232A polymorphism in cattle.

A new, quick, and inexpensive method for detecting the bovine acyl-CoA:diacylglycerol acyltransferase1 (DGAT1) polymorphism (K232A) through tetra-primer amplification refractory mutation system by PCR (ARMS-PCR) was developed in the present investigation. The DGAT1 gene was recently identified as underlying variation in milk production traits. To date, PCR techniques such as PCR-RFLP have been used for detecting the DGAT1 K232A polymorphism, despite being expensive and laborious. The method proposed here, a tetra-primer ARMS-PCR, showed 100% sensitivity and specificity when compared with PCR-RFLP results obtained in a sample of 80 animals tested in a double-blind system. Our results suggest that the use of tetra-primer ARMS-PCR for DGAT1 K232A polymorphism genotyping could greatly reduce costs providing information for both research purposes and for dairy cattle breeders who perform DGAT1 genotyping for gene-assisted selection.

Genetic diversity and population structure in Brazilian Mangalarga Marchador horses.

One hundred and fifteen unrelated Mangalarga Marchador horses were sampled from three geographically distinct regions of Minas Gerais State, Brazil (South, Southeast, and Northeast) and tested for 10 microsatellite loci. Genetic diversity and population structure parameters were estimated with ARLEQUIN 3.0, CERVUS 2.0, POPGENE 1.31, GENEPOP on the web, STRUCTURE 2.0, and SPAGEDI 1.2 software packages. Under Hardy-Weinberg assumptions, seven markers were at equilibrium (LEX014, LEX017, LEX019, SGCV23, TKY321, VHL20, and VIASH39), while two (ASB3 and LEX031) presented significant homozygote excess. Seventy-four alleles were identified in these nine markers, with a mean of 8.22 alleles. Mean heterozygosity was 0.637 and polymorphism information content was 0.662. Markers ASB3, LEX019, SGCV23, TKY321, and VHL20 were highly informative (PIC >0.7) and may be useful for eventual expansion of parentage test panels. The F(ST) value (0.0562) indicated relatively little geographical structure. However, based on a Bayesian-based cluster analysis under a three-cluster model, 94% of the 115 individuals were correctly assigned to the subpopulations from where they were sampled. Mean pairwise f was relatively high (0.11), and in spite of the efforts towards non-consanguineous sampling, 1% of the pairs of individuals shared over 50% of the alleles. These results strongly suggest that the population is genetically structured. Under a conservation genetics approach, two strategies are recommended: avoidance of crosses between highly endogamic individuals and stimulation of crosses between individuals from those regions for which low genetic flow was identified.

Colorectal tumor-on-a-chip system: A 3D tool for precision onco-nanomedicine.

Awareness that traditional two-dimensional (2D) in vitro and nonrepresentative animal models may not completely emulate the 3D hierarchical complexity of tissues and organs is on the rise. Therefore, posterior translation into successful clinical application is compromised. To address this dearth, on-chip biomimetic microenvironments powered by microfluidic technologies are being developed to better capture the complexity of in vivo pathophysiology. Here, we describe a "tumor-on-a-chip" model for assessment of precision nanomedicine delivery on which we validate the efficacy of drug-loaded nanoparticles in a gradient fashion. The model validation was performed by viability studies integrated with live imaging to confirm the dose-response effect of cells exposed to the CMCht/PAMAM nanoparticle gradient. This platform also enables the analysis at the gene expression level, where a down-regulation of all the studied genes (MMP-1, Caspase-3, and Ki-67) was observed. This tumor-on-chip model represents an important development in the use of precision nanomedicine toward personalized treatment.