Heat stress during pregnancy can affect the intrauterine development and modify the expression of key molecules involved in reproductive function in offspring adulthood. This study aimed to evaluate changes in the expression of insulin-like growth factor receptor 1 (IGFR1), insulin receptor (IR) and their downstream signaling pathways in dominant ovarian follicles of Holstein cows born to dams naturally exposed to different temperature-humidity index (THI) conditions during gestation. Cows (n = 26) were evaluated according to their exposure to a THI ≥ 72 during gestation, determined using their birth date or effective service date of their dam. Gestation was divided, based on key events in ovarian development, into two periods (P1 = days 0–150; P2 = day 151 to calving) and three trimesters (T1 = days 0–90; T2 = days 91–180; T3 = day 181 to calving), and the exposure to THI ≥ 72 was calculated in each. Total and phosphorylated expression levels of IGFR1, IR, IRS, PI3K, and AKT were assessed in the dominant follicles of adult offspring. Variables related to prenatal heat stress in P1, T1 and T2 were positively associated with IGFR1 and pIGFR1 expression (P < 0.05). Instead, variables related to prenatal heat stress in P2 were negatively associated with IGFR1 and pIGFR1 expression (P < 0.05). Signaling components such as IRS, pIRS, and pAKT were positively associated with variables related to prenatal heat stress in T2 in ovarian cells of adult offspring. However, PI3K expression in adulthood was negatively associated with variables related to prenatal heat stress in T1 (P < 0.05). These findings indicate that heat stress during pregnancy is associated with persistent alterations in key components of the IGF1/insulin signaling pathway in dominant follicles of adult offspring.

In cattle, low progesterone (P4) concentrations have been linked to anovulation. To study the local effects of P4 on granulosa cells, in the present study we aimed: (a) to characterise the expression of receptors (progesterone receptor (PR), oestrogen receptors α and β (ERα and β), FSH receptor (FSHR) and LH receptor (LHR)), as well as the co-expression of PR and signal transducer and activator of transcription 5 (STAT5) and a cytochrome P450 aromatase A1 (CYP19A1), in a bovine granulosa cell line (BGC-1) sensitive to P4 stimuli; and (b) to evaluate the effects of P4 on cell viability, apoptosis and reactive oxygen species (ROS) production in BGC-1 cells after 12, 24 and 48 h of stimulation. BGC-1 cells expressed CYP19A1, as well as ERα and β, PR, FSHR and LHR. The mean fluorescence intensity of STAT5 and PR and the percentage of simple and double-positive cells (STAT5/PR) were lower in control cells than in P4-stimulated cells after 12, 24, and 48 h of stimulation. Regarding the effects of P4 on cell viability, after 12, 24, and 48 h of stimulation, the percentage of living cells was high and the number of apoptotic and necrotic cells was low. Finally, BGC-1 cells responded to P4 stimulation by stimulating ROS production, without inducing damage. These results support the potential use of P4-stimulated BCG-1 cells to study the mechanisms underlying follicular persistence in cows.

Ovulatory failure in dairy cows can stem from various causes, including the formation of follicular cysts, which disrupt the estrous cycles and extend reproductive intervals, ultimately hindering herd management and reproductive performance. Reproductive dysfunctions are frequently associated with metabolic disorders, commonly observed in these animals. Therefore, this study aimed to evaluate key components of lipid metabolism, primarily related to β-oxidation, including AdipoR1, AdipoR2, AMPK, PPARγ, ObR, ACC1, CPT1, and ACOX1 in ovary, as well as adiponectin, leptin, triglyceride (TG) and non-esterified fatty acid (NEFA) concentrations in plasma and follicular fluid of dairy cows with induced persistent follicles, spontaneous follicular cysts and dominant control follicles. Ovary protein immunoexpression was assessed in situ by immunohistochemistry, plasma and follicular fluid adiponectin and leptin concentrations were quantified by ELISA, and TG and NEFA concentrations were determined by colorimetric methods. The results indicated increased protein expression of adiponectin receptors in follicles of 15 days of persistence, along with an upregulation of CPT1 expression. ACC1 expression was lower in both follicles of 15 days of persistence and in spontaneous cysts than in preovulatory follicles. Furthermore, PPARγ expression was decreased in follicles of 15 days of persistence compared to dominant control follicles. These findings indicate changes in key molecules involved in ovarian lipid metabolism, with a pronounced shift towards β-oxidation pathways and decreased lipid synthesis and lipid storage. Such metabolic disruptions may contribute to impaired ovarian function, potentially leading to ovulatory failure of the preovulatory follicle.

Cystic ovarian disease (COD), characterized by the presence of persistent follicles, is a major cause of subfertility in dairy cows. This study aimed to evaluate the expression of receptors and enzymes involved in ketone body metabolism, cholesterol regulation, and steroidogenesis within ovarian follicular cells at different stages of persistence. The study was conducted in a model of follicular persistence induced by prolonged progesterone administration in dairy cows, and in cows diagnosed with spontaneous COD. The protein levels of key components, including HMG-CoA reductase, mitochondrial HMG-CoA (mHMG-CoA) synthase, SCOT, LDL-R, SRB-1, CYP17A1, CYP19A1, StAR, and 3βHSD, was assessed in follicles through immunohistochemistry. Additionally, total cholesterol, HDL cholesterol, LDL cholesterol concentrations in follicular fluid and plasma were measured using a biochemical autoanalyzer, while β-hydroxybutyrate (BHB) levels were evaluated with reactive strips. Results showed that protein levels of SRB-1 and LDL-R in granulosa cells was higher in cows in late stages of follicular persistence and COD cows than in the control group (P < 0.05). In contrast, mHMG-CoA synthase, HMG-CoA reductase and SCOT revealed an opposite pattern (P < 0.05). In granulosa cells, CYP19A1 levels were lower in follicles with 5 days of persistence than in control follicles and 3βHSD levels were higher in late stages of persistence than in controls. These alterations evidenced an imbalance in relevant components of lipid metabolism and steroidogenesis. Changes observed in late persistence or cyst would be a consequence of follicular persistence contributing to subfertility in cattle.

In dairy herds, it is expected that cows will be cycling and the uterus will be ready for a new conception before the fourth week postpartum. However, an alteration in the endometrial remodeling can delay conception, increasing the parturition-to-conception interval, and consequently decreasing the reproductive performance. The endometrial matrix has a relevant participation in the processes of postpartum uterine remodeling. In this sense, the matrix metalloprotease (MMP) system and its inhibitors (TIMPs) are directly involved in the proteolytic degradation of the matrix and their action is related to the concentration of steroid hormones. The aim of this study was to evaluate the protein expression of MMP2, MMP14, MMP9, and their inhibitors, TIMP1 and TIMP2, in the luminal epithelium, glandular epithelium and stroma of endometrial biopsies from dairy cows, at 60 days in milk. Together, the results obtained provide evidence about the expression of MMP2, MMP14 and MMP9, and their inhibitors, TIMP1 and TIMP2, in the postpartum uterus of dairy cows, and about how the balance in their expression could be associated with the achievement of pregnancy. The high protein expression of MMP2, MMP14 and TIMP1 in dairy cows with short parturition-to-conception interval could be important for uterine remodeling and early conception in dairy cows. In addition, the imbalance observed in the MMP9/TIMP1 ratio could be generating an excess of gelatinase activity in the endometrium, causing a delayed conception.

The cytokine context present in the reproductive tract of cows is closely involved in normal uterine functions, including the estrous cycle and the establishment and maintenance of pregnancy. However, the roles of some cytokines in the uterus, and their relation with reproductive performance remain to be elucidated. Thus, this study aimed to examine the protein expression of several cytokines such as TNFα, IL-6, IL-8, IFNγ, IL-4, and TGF-β3 in endometrial biopsies previous to conception, to evaluate the possible association with delayed conception in dairy cows. Protein expression levels were evaluated by immunohistochemistry. Results showed that the protein expression levels of TNFα, IL-6, IL-4 and TGF-β3 were not associated with the parturition-conception interval, whereas the high protein expression levels of IFNγ were associated with the parturition-conception interval. Finally, the low protein expression of IL-8 showed a statistical tendency to be associated with delayed conception. This is the first report about the protein expression of IFN-γ in the endometrium of dairy cows and also, this cytokine could enhance the favorable conditions to achieve an early pregnancy.

Failure of ovulation can lead to follicular persistence, one of the main components of the pathogenesis of cystic ovarian disease (COD) in dairy cattle. Follicular persistence causes the permanence of a functional follicular structure in the ovary, which alters the cyclicity of the female and causes infertility. The aim of the present study was to evaluate the expression of estrogen receptors (ESR) 1 and 2, and the coregulatory proteins NCOA1, NRIP1 and LCOR by immunohistochemistry, in antral and preovulatory/persistent follicles in a model of follicular persistence induced by low levels of progesterone, to detect incipient changes during COD development, on the expected day of ovulation (P0) and after 5 (P5), 10 (P10) and 15 (P15) days of follicular persistence. Twenty-five Holstein cows were used, which were distributed in 5 groups: control group (n = 5), group P0 (n = 5), group P5 (n = 5), group P10 (n = 5), group P15 (n = 5). ESR1 expression was lower in antral follicles of the P5 (theca), P10 and P15 (theca and granulosa) groups relative to the control group (p < 0.05), and also lower in granulosa cells of persistent follicles of the P5, P10 and P15 groups than in dominant follicles of the control group (p < 0.05), without differences in theca cells. ESR2 expression showed no differences between groups. The ESR1:ESR2 balance favored ESR2 expression along the development of persistent follicles, as from 5 days of persistence (p < 0.05). NCOA1 expression was higher in granulosa cells of both antral and persistent follicles from the P0 group relative to the P5 and P10 groups, but showed no differences with the control and P15 groups (p < 0.05). Theca cells of antral and persistent follicles showed higher expression in the P0 and P15 groups in relation to the control, P5 and P10 groups (p < 0.05). No differences were detected for NRIP1 in antral, dominant and persistent follicles between groups. LCOR expression showed a decrease in granulosa cells of antral follicles from all persistence groups relative to the control group (p < 0.05). In theca cells, antral follicles of the P10 group showed lower LCOR expression than the control group (p < 0.05). LCOR expression was similar for dominant and persistent follicles. Considering that the ESR1:ESR2 balance favored ESR2 expression along the development of persistent follicles, as well as the decreased LCOR and NCOA1 expression, we may assume that, at the early stages of persistence, there is a negative regulation of ESR transcription. This coincides with the effects of estrogens through ESR on proliferation and apoptosis among other processes that favor follicular persistence. The results obtained provide relevant information in the knowledge of local events during the development of follicular persistence that could explain the failures in the reversion of the disease through hormonal treatments and the high recurrence rates reported for COD. In addition, it contributes to the study and identification of possible therapeutic targets, for the design of new treatments.

This study aimed to analyze the possible relationship between days to conception and different oxidative stress (OS) biomarkers and liver functional parameters in multiparous dairy cows. Besides, a fast reliable method for the accurate measurement of malondialdehyde (MDA) by liquid chromatography–tandem mass spectrometry was developed in several matrices. During lactation, the days to conception of 28 cows were determined for a retrospective study. According to this parameter, cows were divided into two groups: high and low days to conception (HDC and LDC, respectively). Blood, urine and liver biopsies were sampled 21 days before the expected calving date, and 7 and 21 days after calving. The method developed for MDA was validated according to international requirements. The lower limit of quantification was 0.25 µmol/L for plasma and urine and 10.00 µmol/L for liver tissue. No differences between groups were observed in the systemic concentration of non-esterified fatty acids, β-hydroxybutyric acid and liver triacylglycerol content (P > 0.05). Cholesterol concentration was higher in the LDC than in the HDC group (P < 0.05). Plasma 3-nitrotyrosine (3-NT) concentration was lower in the LDC than in the HDC group on day 21 post-calving (P < 0.05). Superoxide dismutase activity was higher in the LDC than in the HDC group (P < 0.05). Particularly, in the liver, 3-NT and MDA concentrations were lower in the LDC than in the HDC group (P < 0.05). These results allow inferring that the amelioration of OS biomarkers in plasma and liver could be related to a better reproductive performance of dairy cows.

Before ovulation, the ovary exhibits signs of local inflammation. However, the effects of adrenocorticotropin (ACTH) on the complexity of this inflammatory response are not yet well described. Thus, the aim of this study was to evaluate the effects of ACTH administered to dairy cows during the preovulatory period on the local distribution of different subsets of leukocytes infiltrated in the ovary, along with the gene expression of relevant chemokines (C-C motif chemokine ligand-2 (CCL2), C-X-C motif chemokine ligand-8 (CXCL8), CCL25 and CXCL1) involved in leukocyte chemotaxis and blood perfusion on the follicular wall of dominant follicles. Also, the direct effect of ACTH on chemokine gene expression was addressed in cultured antral follicular walls. For this purpose, both an in vivo and an in vitro experiment were performed. For the in vivo experiment, exogenous ACTH (100 IU) was administered intramuscularly to Holstein cows (n = 12) during proestrus every 12 h for four days before ovulation, when ovariectomy was performed (day 18). Daily ovarian Doppler ultrasonography was used to evaluate the percentage of irrigated area, the pulsatility index and the resistance index in the dominant follicles. The distribution of monocytes-macrophages (CD14), T- (CD2) and B-lymphocytes (CD79a) and granulocytes (CH138A) in the ovary was analyzed by immunohistochemistry. In follicular wall samples, gene expression of CCL2, CXCL8, CXCL1 and CCL25 was evaluated, whereas IL-17A expression was analyzed by Western blot. The total number of CD14, CD79a and CD2 infiltrated cells was lower in the ACTH-treated group than in the control group (p < 0.05). Chemokine gene expression showed lower mRNA of CCL2, CCL25 and CXCL1 (p < 0.05) in the ACTH-treated group. Meanwhile, IL-17A protein expression and hemodynamic parameters were similar between groups (p > 0.05). In the in vitro assay, antral follicular walls were stimulated with ACTH to corroborate the gene expression profile of chemokines. mRNA expression of CCL2 tended to be lower in the stimulated follicular walls (p = 0.092). Our results suggest that exogenous ACTH stimulus during the preovulatory period reduces the number of infiltrated leukocytes in the bovine ovary and this could be due to a lower chemotaxis capacity of the ovary.

Glucocorticoids (GCs) act through their receptor (GR) as regulators in different biological processes such as reproduction. In the absence of GCs, the GR remains inactive in the cytoplasm by associating with heat shock proteins (HSPs), which act as molecular chaperones, among which the most relevant are HSP90 and HSP70. Cytoplasmic GC-activated GR mediates non-genomic effects, interacting with members of signaling pathways such as PI3K/Akt, which participates in several metabolic processes, including the insulin signaling pathway. The aim of the present study was to evaluate possible associations between the cytoplasmic GR and the main intermediates of the insulin signaling pathway and HSP90 and HSP70 in ovaries of dairy cows. To this end, the protein expression of cytoplasmic GR, key members of the insulin signaling pathway, and HSPs was evaluated in ovarian preovulatory follicles of non-lactating Holstein cows in proestrus. Positive associations were observed between protein expression of GR and HSP90, IRS1, pIRS1, PI3K and pAkt (p < 0.05; β > 0) in granulosa cells of dominant follicles of dairy cows. Instead, in theca cells, no associations were observed between protein expression of GR and members of the insulin signaling pathway or HSPs. These data provide evidence of the possible association between the non-genomic mechanisms of action of the GR and the insulin signaling pathway in the bovine ovary.

The alteration of signaling molecules involved in the general metabolism of animals can negatively influence reproduction. In dairy cattle, the development of follicular cysts and the subsequent appearance of ovarian cystic disease (COD) often lead to decreased reproductive efficiency in the herd. The objective of this review is to summarize the contribution of relevant metabolic and nutritional sensors to the development of COD in dairy cows. In particular, we focus on the study of alterations of the insulin signaling pathway, adiponectin, and other sensors and metabolites relevant to ovarian functionality, which may be related to the development of follicular persistence and follicular formation of cysts in dairy cattle. The results of these studies support the hypothesis that systemic factors could alter the local scenario in the follicle, generating an adverse microenvironment for the resumption of ovarian activity and possibly leading to the persistence of follicles and to the development and recurrence of COD.

The cytokines of the interleukin-1 (IL-1) family are closely involved in the resolution of inflammation in cows with metritis and endometritis. However, little is known about the role of these cytokines beyond uterine regression in the absence of disease, especially around conception. Thus, the aim of this study was to examine the gene and protein expression of IL-1α, IL-1β, IL-1RI, IL-1RII and IL-1Ra in endometrial biopsies previous to conception, to evaluate the possible association of these cytokines with delayed conception in dairy cows. Gene and protein expression levels were evaluated by real-time PCR and immunohistochemistry, respectively. The gene expression levels of cytokines were not associated with the duration of the period to conception following parturition. However, high protein expression of IL-1β and low protein expression of IL-1Ra were significantly associated with early conception. These results suggest that an imbalanced protein expression of IL-1β and IL-1Ra in the endometrium of dairy cows could be part of the maternal immune response mechanism necessary to propitiate early conception and probably to maintain pregnancy.

The regular consumption of citrus fruits by humans has been associated with lower incidence of chronic-degenerative diseases, especially those mediated by free radicals. Most of the health-promoting properties of citrus fruits derive from their antioxidant content of carotenoids and ascorbic acid (ASC). In the current work we have investigated the scavenging (against hydroxyl radical) and quenching capacities (against singlet oxygen) of four different carotenoid extracts of citrus fruits in the presence or absence of ASC (μM range) in organic solvent, aqueous solution, micelles and in an innovative biomimicking liposomal system of animal cell membrane (AML). The fruits of four varieties of citrus were selected for their distinctive carotenoid composition (liquid chromatography characterization): ‘Nadorcott’ mandarin and the sweet oranges ‘Valencia late’, ‘Ruby Valencia’ and ‘Pinalate’ mutant. The quenching activity of citrus carotenoids strongly depended on the biological assemblage: freely diffusible in organic solvent, ‘Ruby Valencia’ carotenoids (containing lycopene) showed the highest quenching activity, whereas ‘Nadorcott’ mandarin extracts, rich in β-cryptoxanthin, prevailed in micellar systems. Interestingly, the addition of 10 μM ASC significantly increased the quenching activity of all citrus extracts in micelles: ‘Valencia’ orange (+53%), ‘Pinalate’ (+87%), ‘Ruby’ (4-fold higher) and ‘Nadorcott’ mandarins (+20%). Accurate C11-BODIPY581/591 fluorescence assays showed solid scavenging activities of all citrus extracts against AML oxidation: ‘Valencia’ (−61%), ‘Pinalate’ (−58%) and ‘Ruby’ oranges (−29%), and ‘Nadorcott’ mandarins (−70%). Indeed, all four citrus extracts tested here have balanced antioxidant properties; extracts from the ‘Nadorcott’ mandarin slightly prevailed overall, due, at least in part, to its high content of β-cryptoxanthin. This study depicts some of the antioxidant interactions between citrus fruit carotenoids and ascorbic acid in models of animal cell membranes and reinforces the contribution of them in promoting health benefits for humans.

In the present work we aimed to study the effects of parenteral vitamin and mineral supplementation on hepatic fatty acid metabolism as well as on the oxidative stress biomarkers in biological samples of transition cows. The supplemented group (SG, n = 11) received a subcutaneous injection of 5 mL of vitamin A palmitate 35 mg/mL, vitamin E acetate 50 mg/mL plus other injection of 5 mL of copper edetate 10 mg/mL, zinc edetate 40 mg/mL, manganese edetate 10 mg/mL, and sodium selenite 5 mg/mL on days − 60, − 30, and 7 (± 3) relative to calving. The control group (CG, n = 11) received two subcutaneous injections of 5 mL of 9 mg/mL sodium chloride at the same times of the SG. Blood, urine, and liver biopsies were sampled 21 (± 3) days before the expected calving date and 7 and 21 (± 3) days after calving. Results revealed that supplemented animals had higher glutation peroxidase (GSH-Px) activity, lower and higher concentration of 3-nitrotyrosine (3-NT) in the liver and plasma, respectively, higher expression of the mitochondrial beta-oxidation enzyme carnitine palmitoyltransferase 1 in the liver, and lower content of hepatic triacylglycerol, mirroring plasma liver function parameters. No differences between groups were found in the superoxide dismutase activity, MDA concentrations, the protein abundance of peroxisomal acyl-CoA oxidase 1, diacylglycerol O-acyltransferase 1, and peroxisome proliferator-activated receptor alpha. These results suggest that the vitamin and mineral supplementation provided to dairy cows had a beneficial effect on GSH-Px activity, hepatic 3-NT concentration, and on the metabolic adaptation during the peripartum period.

Background:Hemolytic uremic syndrome associated with Shiga-toxin produced by Escherichia coli is a serious worldwide foodborne disease. Nowadays, no treatment is available, only supportive care can be provided, and 50 % of the patients require a period of dialysis. Recently, a therapy based on Neutralizing Equine Anti Shiga Toxin (NEAST) antibodies has been developed. NEAST is composed of F(ab’)2 fragments from equine immunoglobulins.Objective:The purpose of this study was to develop an ELISA to measure serum concentrations of NEAST in mice and rabbits, and to validate it according to international recommendations. The validated method was further used to analyze the NEAST PK during preclinical studies.Methods:A sandwich ELISA was developed, the performance of the calibration curve was assessed, and it was validated based on the parameters as accuracy, precision, specificity, selectivity, stability of the analyte, and dilutional linearity.Results:This immunoassay was specific, sensitive, accurate and precise in a dynamic range from 7.81 to 500 ng/mL and from 15.63 to 500 ng/mL for mice and rabbits, respectively. This method was successfully applied to PK studies of NEAST after intravenous administration.conclusion:The results obtained are expected for a robust ELISA used for macromolecule analysis. Since NEAST is an equine F(ab′)2, this immunoassay would serve for the evaluation of the PK profile of any biological product based on molecules with similar characteristics. This immunoassay may be useful for current and future preclinical trials conducted for registration purposes.

In dairy cows, reproductive diseases such as cystic ovarian disease (COD) represent a major problem that impacts on dairy production. It has been postulated that the insulin-like growth factor (IGF) system may contribute to follicular persistence and development of COD. Thus, the aim of the present study was to analyze relevant members of the IGF system in a critical period immediately after the expected time of ovulation, to obtain information about their role in follicular persistence in dairy cows. Proteins of the IGF system were evaluated at 0 (expected day of ovulation), 5, 10 and 15 days of follicular persistence to determine whether the changes previously detected in cows with COD occur early in COD pathogenesis. The serum concentration of IGF1 was higher in cows with 10 and 15 days of follicular persistence than in control cows. IGF1 expression in granulosa cells was similar in the follicles analyzed. In contrast, in theca cells, persistent follicles of days 5 and 10 showed the lowest IGF1 expression. IGF binding protein (IGFBP) 2 and 3 expression was lower in persistent follicles than in dominant follicles of the control group. Although IGF receptor (IGFR) 1 expression was similar in the groups analyzed, p-IGFR1 expression was significantly higher in dominant follicles of the control group than in persistent follicles. These data suggest alterations in the IGF system at the early stages of follicular persistence. The evidences obtained allow supporting that the IGF system could plays a key role in dairy cattle reproduction.

EL SARS-CoV-2 es el agente etiológico responsable de la enfermedad COVID-19 que inició una pandemia desde finales de 2019. Existen distintos modelos animales para esta enfermedad siendo el modelo de ratón transgénico K18-hACE2, originalmente desarrollado para el estudio del SARS-CoV-1, de gran relevancia en el contexto actual. Considerando que los ratones se comercializan como hemicigotas y dada la ausencia de ratones homocigotas para el transgén K18-hACE2, el objetivo de este trabajo fue generar una colonia de ratones homocigotas K18-hACE2-Tg/Tg bajo condiciones ambientales controladas. Para la F1 se utilizaron 10 ratones hemicigotas (7 hembras y 3 machos), de 6-8 semanas de edad, provenientes de The Jackson Laboratory (USA). De la F1 se identificaron mediante genotipificación por PCR 78 animales: 62 hemicigotas (Tg/0) y 16 salvajes (0/0). Aquellos animales identificados como transgénicos se cruzaron con ratones C57BL/6J para seleccionar los parentales homocigotas que generaron 100% de hemicigotas en la F2. De 224 crías analizadas, se obtuvieron 7 parentales homocigotas K18-hACE2-Tg/Tg que fueron utilizados para la colonia fundación de ratones homocigotas K18-hACE2-Tg/Tg. El modelo de ratón homocigota K18-hACE2-Tg/Tg desarrollado en este trabajo podrá ser empleado para el estudio de la patogénesis de la enfermedad y la evaluación de posibles terapéuticos contra el SARS-CoV-2

The epidermis of mammals is characterized by having a stratum granulosum that produces an orthokeratotic stratum corneum, different from the typical reptilian parakeratotic stratum. Nonetheless, some mammals show distinct degrees of parakeratosis in epidermal regions with few or no pilose follicles (e.g., areas subjacent to cornified scales). With respect to the epidermis and the development of cornified scales in the Dasypodidae, previous studies have supported the presence of a continuous stratum granulosum without any variations during ontogeny. This condition, in which the cornified scales develop without a loss of the stratum granulosum, was interpreted as primitive for eutherians. The present contribution expands the knowledge on the epidermis of Chaetophractus vellerosus in distinct ontogenetic stages in order to determine whether the cornified scales show the same developmental pattern as in other eutherians. The presence of a stratum granulosum in C. vellerosus neonates and its reduction in more advanced ontogenetic stages, in direct relationship with cornified scale development, supports the hypothesis that the partial parakeratosis in the xenarthran integument is secondary, as in other eutherians, and can be interpreted as a derived character state.

CONTEXT:Dairy cattle experience stressful environmental situations that affect production. Heat stress during gestation can influence the intrauterine development of offspring, resulting in long-term damage that can affect the reproductive life of the adult offspring.AIM:The aim of the present study was to evaluate changes in the expression and regulation of steroid hormone receptors in the ovary of Holstein cows gestated under different temperature-humidity index (THI) during their in utero development.METHODS:Animals were classified by their exposure to temperature-humidity index (THI) ≥72 during their development in utero according to date of birth or date of effective service of their mother. This study was not carried out under controlled conditions, but the conditions to which the cows were naturally exposed during their development were considered retrospectively, controlling the variables in the statistical analyses (age as a covariate, dairy farm as a random factor). Gestation was divided into two periods (P1=days 0-150; and P2=day 151 to calving) and three trimesters (T1=days 0-90; T2=days 91-180; and T3=day 181 to calving), and the exposure to THI ≥72 was calculated in each one. The following characteristics were evaluated: gene expression of estrogen receptor (ESR) 1, ESR2 and progesterone receptor (PGR), CpG methylation in the 5'UTR of ESR1 and ESR2, and protein expression of ESR1, ESR2, PGR and coregulatory proteins in the dominant follicles of daughter cows in adulthood.KEY RESULTS:We found associations between heat stress variables during gestation and the methylation status of CpG sites in the 5'UTR of ESR1 and ESR2 in dominant follicles. Results also showed association between exposure to high THI values during intrauterine development and expression of ESR1, ESR2 and PGR and coregulatory proteins in dominant follicles of adult cows.CONCLUSIONS:These results provide novel information about the impact of prenatal heat stress on molecular aspects at the ovary level in the offspring, during their adult life, which probably impacts the reproductive aspects of the herd.