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Clinical use of hormone status determination in canine and feline reproduction Suppawiwat Ponglowhapan, DVM (Hons), M.S., M.Sc, PhD Department of Obstetrics Gynaecology and Reproduction Faculty of Veterinary Science, Chulalongkorn University Introduction • Hormone Hormaein (ภาษากรี ก) = Impetus (a moving force, stimulus) แปลว่า กระตุ้น (excite), ก่อกวน (stir up) หรื อทาให้ เกิดการเปลี่ยนแปลง (to set in motion) • Endocrine The endocrine system is a group of glands that work together and secrete many types of different hormones that regulate the body. Introduction Major endocrine glands. (Male on the left, female on the right.) 1. Pineal gland 2. Pituitary gland 3. Thyroid gland 4. Thymus 5. Adrenal gland 6. Pancreas 7. Ovary 8. Testes Introduction • การควบคุมการทางาน ▫ Autocrine ▫ Paracrine ▫ Endocrine Introduction • Hormone feedback regulatory system ▫ Negative feedback ▫ Positive feedback • Physiological mechanisms of endocrine • Pathologic mechanisms of endocrine ▫ Hormone excess (tumor, exogenous route) ▫ Hormone deficiency (autoimmune, surgery, inflammation) ▫ Hormone resistant (genetic factor) Introduction • สามารถแบ่งฮอร์ โมนออกเป็ น 5 ชนิด 1. Amino acid derivatives; dopamine, catecholamines, thyroid hormone 2. Small neuropeptides; GnRH, TRH, somatostatin, vasopression 3. Large proteins; LH, FSH, insulin, parathyroid hormone 4. Steroid hormones; oestrogen, progesterone, testosterone, cortisol 5. Vitamin derivatives; retinoids Introduction • หน้ าที่หลักของฮอร์ โมน 1. Growth and differentiation 2. Maintenance of homeostasis 3. Reproduction Introduction • สามารถแบ่งฮอร์ โมนออกเป็ น 5 ชนิด 1. 2. 3. 4. 5. Amino acid derivatives; dopamine, catecholamines, thyroid hormone Small neuropeptides; GnRH, TRH, somatostatin, vasopression Large proteins; LH, FSH, thyroid hormone, insulin Steroid hormones; oestrogen, progesterone, testosterone,cortisol Vitamin derivatives; retinoids Reproduction • Progesterone • Oestrogen • Testosterone • Gonadotrophins: LH & FSH • Thyroid hormone Progesterone Definitions 1. A female hormone and the principal progestational hormone that is made mainly by the corpus luteum in the ovary and by the placenta. Progesterone prepares the lining (endometrium) of the uterus to receive and sustain the fertilized egg and so permits pregnancy. 2. The principal progestational hormone liberated by the corpus luteum, adrenal cortex, and placenta, whose function is to prepare the uterus for the reception and development of the fertilized oocyte by inducing transformation of the endometrium from the proliferative to the secretory stage. Progesterone • A sex steroid hormone • CL, adrenal gland, placenta (cat) • Pregnancy, mammary gland development Progesterone ประโยชน์ของการตรวจโปรเจสเตอโรนในสุนขั 1.เพื่อกาหนดวันผสม (optimal breeding time) เพิ่มโอกาสผสมติด เพิ่มจานวนลูกต่อครอก 2.เพื่อตรวจความผิดปกติของการตังท้ ้ อง 3.เพื่อทานายวันคลอด Determination of optimal breeding time in the dog Luteinization of the follicle in the dog Guidelines for interpretation of serum progesterone concentration in the estrous bitch measured by RIA Serum Progesterone (ng/mL) Estimated day of ovulation Best breeding days Estimated parturition from ovulation days 1.0-1.9 + 2 days + 4 days (3-6) + 63 days (62-64) 2.0-3.9 + 1 days + 3 days (2-5) + 63 days (62-64) 4.0-10.0 0 days + 2 days (1-4) + 63 days (62-64) For natural mating or AI using fresh semen, inseminate the day after ovulation For AI using chilled or frozen semen, inseminate 2-4 days after ovulation Predicted time of ovulation based on serum progesterone (n = 50; > 20 breeds) Key points 1.Traditional or classical dates for canine breeding: No one-for-all rule 2.Importance of determining serum progesterone level for canine breeding Determination of optimal breeding time in the dog Determination of optimal breeding time in the dog • LH and ovulation? • Why LH check is less likely to be recommended in practice? ▫ Specificity of LH test ▫ Lab Availability :Thailand? Determination of optimal breeding time in the dog • Is there any in-house LH test kit? YES Determination of optimal breeding time in the dog • Some considerations on the clinical use of LH test kit ▫ Advantages ▫ Price ($230 per box/ 6 tests) ▫ The number of cases ▫ Frequency of the use/ cycle Determination of optimal breeding time in the dog • Unlike other mammals, dogs ovulate when the oestrogen level is declining. • Oestrogen levels can only give us a general idea of when a dog will come into cycle. Vaginal cytology in the cat Determination of optimal breeding time in the dog • Oestrogen and vaginal cytology? basal/ parabasal cell intermediate cell superficial cell Determination of optimal breeding time in the dog • The use of vaginal cytology for canine breeding management ▫ ▫ ▫ ▫ To ensure a dog is in cycle When to start vaginal swab? How often? When to start collecting blood sample for progesterone assay? Recommended workup for determination of optimal breeding time in the bitch • History taking (previous oestrus, first day of vaginal bleeding: Day0) • Physical exam (General and reproductive exam; vaginal discharge, vulvar swelling) • Estrous behaviors • Vaginal smear to confirm the bitch is in estrous stage • Blood collection for progesterone measurement Guidelines for interpretation of serum progesterone concentration in the estrous bitch Serum Progesterone (ng/mL) Estimated day of ovulation Best breeding days Estimated parturition from ovulation days 1.0-1.9 + 2 days + 4 days (3-6) + 63 days (62-64) 2.0-3.9 + 1 days + 3 days (2-5) + 63 days (62-64) 4.0-10.0 0 days + 2 days (1-4) + 63 days (62-64) Key Notes 1.Stress and progesterone hypersecretion 2.Combination of behaviour, vaginal cytology and progesterone 3.Delivery of live puppies can not be guaranteed. Progesterone ประโยชน์ของการตรวจโปรเจสเตอโรนในสุนขั 1.เพื่อกาหนดวันผสม (optimal breeding time) เพิ่มโอกาสผสมติด เพิ่มจานวนลูกต่อครอก 2.เพื่อตรวจความผิดปกติของการตัง้ ท้ อง 3.เพื่อทานายวันคลอด Pregnancy monitoring Hypoluteoidism in the bitch • • • • Hypoluteoidism = luteal insufficiency Progesterone levels > 2.0-2.5 ng/mL are necessary to support pregnancy. Incidence? Cause? ▫ Insufficient relaxin low prolactin ▫ Endometrial infection/inflammation low progesterone PGF2 alpha luteolysis Hypoluteoidism in the bitch • Treatment ▫ Medroxyprogesterone acetate (MPA) 0.1 mg/kg PO sid ▫ Altrenogest 0.088 mg/kg PO sid ▫ Progesterone in oil 1-2 mg/kg IM every other day NOTE Discontinued 2-3 days prior to the expected due date Progesterone ประโยชน์ของการตรวจโปรเจสเตอโรนในสุนขั 1.เพื่อกาหนดวันผสม (optimal breeding time) เพิ่มโอกาสผสมติด เพิ่มจานวนลูกต่อครอก 2.เพื่อตรวจความผิดปกติของการตังท้ ้ อง 3.เพื่อทานายวันคลอด Prediction of parturition date กำไร • 3 Oct 2009 • Golden Retriever, 2 yrs • 54 days of pregnancy • Severe bloody vaginal discharge • General health status is good • No signs of parturition • X-ray: 3 fetal skeletons (not yet full-term) • U/S: one fetus with slow heat beat (< 180 bpm) ▫ Predicted parturition date by U/S (biparietal diameters) = -7 days กำไร • 6 Oct 2009 • Abortion: one fetus • Progesterone level 1.9 ng/mL • 7 Oct 2009 at night • Deliver 2 living puppies Oestrogen and testosterone • Research purpose • Clinical use is for diagnosis of the presence of gonadal tissue by GnRH stimulating test Ovarian remnant syndrome oestrogen Bilateral abdominal cryptorchidism testosterone Ovarian remnant syndrome • How to diagnose wait till the bitch/queen shows oestrus signs GnRH stimulation test • Adverse effects ▫ Owner ▫ Animal Bilateral abdominal cryptorchidism • How to diagnose ▫ Dog: GnRH stimulation test ▫ Cat: penile spine • Adverse effects Before castration After castration ▫ Testicular tumour (high incidence) Testicular tumour Ovarian remnant syndrome • Exner et al., 2008. Theriogenology. Estradiol measurement after GnRH stimulation as a method to diagnose the presence of ovaries in the female domestic cats • GnRH analogue buserelin (Receptal, Intervet) 0.4 microgram/kg IM