Biomarker discovery for bovine reproductive health
The strain during the parturition in cows increases the chances of lacerations and creates an opening for environmental microbes to infect uterine tissues leading to endometritis. The uterine infections will mainly disrupt the main functions of the endometrium resulting in dysregulation of the reproductive cycle via prolonging calving to conceptus intervals and causing infertility. In animals, the uterine disease can often be seen in a subclinical form. Due to the lack of clinical signs, more invasive methods are required for the diagnosis of the disease. The most common method for diagnosis of subclinical disease is through cytology by cytobrush, uterine lavage or cytotape sampling which requires skills, laboratory equipment and time. The results gathered by these methods only poorly correlate with each other. Thus, there is a pressing need for devising new diagnostic tools. Moreover, understanding early pathophysiology of uterine infections and the impact of the disease on uterine environment, embryo development and implantation might help us to evaluate the severity of the infertility, and to detect the precursors for developing a chronic illness. Thus, finding ways to prevent the development of subclinical endometritis altogether.
Recent studies have demonstrated that EVs isolated from uterine fluid takes part in preparation of maternal tissues for implantation, in cross-talk between endometrium and blastocyst during embryo development and play regulatory role in immune responses of uterine tissue. However, the effects of bovine intrauterine EVs have not been well-characterized during different phases of the oestrus cycle and in subclinical endometritis. Thus, in this project, we postulate that uterine EVs may change their functional properties based on the phase of oestrous cycle and uterine infections thus they may influence the embryo development and subsequent pregnancy establishment. The proteomic profiling of EVs from different phases of estrous cycle and in subclinical endometritis will be evaluated and based on these signatures, novel markers for diagnosis of subclinical endometritis will be examined. Moreover, based on uterine EVs influencing embryo development, further studies will be carried out to improve the in vitro embryo development for production of high-quality embryos in vitro.
Recent studies have demonstrated that EVs isolated from uterine fluid takes part in preparation of maternal tissues for implantation, in cross-talk between endometrium and blastocyst during embryo development and play regulatory role in immune responses of uterine tissue. However, the effects of bovine intrauterine EVs have not been well-characterized during different phases of the oestrus cycle and in subclinical endometritis. Thus, in this project, we postulate that uterine EVs may change their functional properties based on the phase of oestrous cycle and uterine infections thus they may influence the embryo development and subsequent pregnancy establishment. The proteomic profiling of EVs from different phases of estrous cycle and in subclinical endometritis will be evaluated and based on these signatures, novel markers for diagnosis of subclinical endometritis will be examined. Moreover, based on uterine EVs influencing embryo development, further studies will be carried out to improve the in vitro embryo development for production of high-quality embryos in vitro.
Project background
Parturition is a risky period for the mother and its offspring in all species and cattle is no exception. During this process cows have a risk of physical damage to the birth canal, failure to detach the placenta and often an upsurge in microbial uterine contamination. Inevitably some will develop uterine infections, which will compromise uterine health by dysregulation of inflammatory response to bacteria, viruses, fungi or collaboration between several pathogens. Many different postpartum reproductive diseases are described in literature, but the primary disease-causing long-lasting effects on fertility is endometritis, which occurs 21 days or more after parturition and has no systemic signs of illness. Most problematic form of endometritis is subclinical endometritis as it has no clinical signs and needs more invasive diagnostics (Figure 2). There are several methods proposed to diagnose the disease such as cytology by cytobrush, uterine lavage or cytotape and ultrasonography assessment of uterine fluid amount or endometrial thickness. However, these methods have little agreement with each other. New diagnostic approaches that can be easily implemented into practice are required for early and correct detection of subclinical endometritis, which would lead to proper treatment and increased fertility.
Parturition is a risky period for the mother and its offspring in all species and cattle is no exception. During this process cows have a risk of physical damage to the birth canal, failure to detach the placenta and often an upsurge in microbial uterine contamination. Inevitably some will develop uterine infections, which will compromise uterine health by dysregulation of inflammatory response to bacteria, viruses, fungi or collaboration between several pathogens. Many different postpartum reproductive diseases are described in literature, but the primary disease-causing long-lasting effects on fertility is endometritis, which occurs 21 days or more after parturition and has no systemic signs of illness. Most problematic form of endometritis is subclinical endometritis as it has no clinical signs and needs more invasive diagnostics (Figure 2). There are several methods proposed to diagnose the disease such as cytology by cytobrush, uterine lavage or cytotape and ultrasonography assessment of uterine fluid amount or endometrial thickness. However, these methods have little agreement with each other. New diagnostic approaches that can be easily implemented into practice are required for early and correct detection of subclinical endometritis, which would lead to proper treatment and increased fertility.
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Figure 2. Diagnosis of subclinical endometritis and the possible outcomes of the disease. Subclinical endometritis in bovine can be diagnosed 21 days postpartum in clinically healthy cows. One of the most common methods for diagnosing subclinical endometritis is with endometrial cytology, for example carried out with a cytobrush technique. The disease is determined with high proportion of neutrophils in the cytology slide. Subclinical endometritis can result in lower pregnancy rate, prolonged calving-conceptus intervals, increased services per conceptus, and postpartum anestrous, which all can lead to culling of the cow. |
EVs are lipid bilayer-delimited particles derived from almost all types of cells containing different biomolecules (e.g. proteins, lipids, nucleic acid), distributing in biological fluids (e.g. blood, uterine fluid) and taken up by cells of the same or different type. Conveying molecules from one cell or tissue to another EVs can exert their biological roles, influencing several normal physiological and pathological conditions by either direct interaction with cell surface receptors or by transmission of their contents by endocytosis, phagocytosis or fusion with the membrane of the target cells. Recent studies have demonstrated that EVs isolated from uterine fluid take part in preparation of maternal tissues for implantation, in cross-talk between endometrium and blastocyst during embryo development and play regulatory role in immune responses of uterine tissue (Figure 3). However, the effects of bovine intrauterine EVs have not been well-characterized during different phases of oestrus cycle and in subclinical endometritis.
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Figure 3. Uterine inflammatory response regulated by extracellular vesicles (EVs). EVs are secreted by pathogens and endometrial cells. These pathogen EVs contain different biomolecules, which are uptaken by endometrial cells can induce an inflammatory response in the endometrium. Inflamed endometrial cells produced EVs, which reflect their physiological condition, can be uptaken by other endometrial cells and influence the physiological condition of recipient endometrial cells. |
Endometritis in Human is the inflammation of the endometrium. Symptoms may include fever, lower abdominal pain, and abnormal vaginal bleeding or discharge. It is the most common cause of infection after childbirth. It is also part of spectrum of diseases that make up pelvic inflammatory disease. The clinical symptoms between Human and cattle endometritis may differ. However, the lessons learned in our research in cattle will be very important to decipher the mechanisms of immunity in the female reproductive tract in different species including Human.
Project objectives
Characterization of uterine fluid EVs profiles of cows with potential application as biomarkers of uterine health.
Characterization of uterine fluid EVs profiles of cows with potential application as biomarkers of uterine health.
- Developing an optimum method for EVs purification from cow uterine fluid.
- Characterization of EVs profiles in bovine uterine fluid.
- Comparison of EVs profiles in healthy cattle during different time points in the oestrus cycle and in cattle suffering from subclinical endometritis.
- Evaluation of bovine uterine fluid EVs acquired from healthy cows and cows with endometritis impact on embryo growth.
- Evaluation of potential use of EVs as biomarkers for diagnosis of subclinical endometritis.
Integration with EMU expertise, research strategy and development
The department of clinical veterinary medicine (CVM) (vl.emu.ee/en/structure/chair-of-clinical-veterinary-medicine/) within EMU research is structured around different areas of disease research and investigation including bovine reproductive physiology and diseases. The current research project is well in line with a long line of research projects in this department to decipher and understand different aspects of endometritis in cattle. Furthermore, this project is well in line with the research project I and will help understanding different means of host-pathogen interactions and strengthening these line of research projects in EMU.
The department of clinical veterinary medicine (CVM) (vl.emu.ee/en/structure/chair-of-clinical-veterinary-medicine/) within EMU research is structured around different areas of disease research and investigation including bovine reproductive physiology and diseases. The current research project is well in line with a long line of research projects in this department to decipher and understand different aspects of endometritis in cattle. Furthermore, this project is well in line with the research project I and will help understanding different means of host-pathogen interactions and strengthening these line of research projects in EMU.
Project team
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Johanna Piibor
Institute of Veterinary Medicine and Animal Sciences Student |
Alireza Fazeli
Institute of Veterinary Medicine and Animal Sciences Supervisor |
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Andres Valdmann
Institute of Veterinary Medicine and Animal Sciences Supervisor |
Suranga Kodithuwakku
Institute of Veterinary Medicine and Animal Sciences Investigator |
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Rallage Keerthi Dissanayake
Institute of Veterinary Medicine and Animal Sciences Invetigator |
Ants Kavak
Institute of Veterinary Medicine and Animal Sciences Supervisor |
