EFFECT OF SELECTED FOOD AND MEDICAL PLANTS AND PLANT MOLECULES ON OVARIAN FUNCTIONS
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1 EFFECT OF SELECTED FOOD AND MEDICAL PLANTS AND PLANT MOLECULES ON OVARIAN FUNCTIONS Alexander V. Sirotkin Constantine the Philosopher University and Research Institute of Animal Production, Nitra, Slovakia
2 Food Supply with energy (calories) Supply with structural elements (proteins, lipids, carbohydrates) Supply with regulatory molecules (antioxidants, phytohormones etc.) Hormones Intracellular regulators of cell proliferation, apoptosis, differentiation Physiological state and health
3 FOOD REGULATORY MOLECULES AND REPRODUCTION: - can food plants affect reproductive functions? - what can be endocrine and intracellular mechanisms of such action? - what for plant molecules are responsible for their action? - can some food plants jeopardize or improve reproduction?
4 AIMS OF THE STUDY: to examine - the effect of some food plants (curcuma/turmeric, green tea, rooibos, ginkgo, flaxseed, yucca) and their components - phytoestrogens and antioxidants (resveratrol, quercetin, daidzein, diosgenin) on ovarian cell functions - mechanisms of action: whether these additives work through changes in hormones release, cell proliferation, apoptosis or response to hormonal stimulators - application of some plants to improve reproduction
5 MATERIAL & METHODS A. Species: - pigs - rabbits B. Objects: - living animals - ovarian granulosa cells
6 MATERIAL & METHODS C. Manipulations: Animals Cells Treatments with: - plant extracts (curcumin, green tea, rooibos, ginkgo, flaxseed, yucca) - plant molecules (resveratrol, quercetin, daidzein, diosgenin) - hormones (FSH, IGF-I) + plants and plant molecules Analysis: reproduction, proliferation, apoptosis, hormones release (RIA, Western, imunocytochemistry, RT-PCR a.o.)
7 CAN FOOD PLANTS (curcumin, green tea, rooibos, flaxseed, ginkgo, yucca etc.) DIRECTLY AFFECT OVARIAN CELL FUNCTIONS?
8 Curcumin reduces ovarian cell proliferation: accumulation of PCNA % of cells contained PCNA Curcumin dose added (µg/ml medium) - accumulation of PCNA mrna Relative units (to control values) 1,2 1,1 1,0 0,9 0,8 0,7 0,6 0,5 0, Curcumin dose added (µg/ml medium)
9 Curcumin promotes ovarian cell apoptosis: accumulation of bax % of cells contained bax Curcumin dose added (µg/ml medium) 16 - accumulation of bax mrna Relative units (to control values) Curcumin dose added (µg/ml medium)
10 Curcumin promotes the release of hormones: 3 - progesterone Progesterone release (ng/10 6 cells/ day) Curcumin dose added (µg/ml medium) - testosteron Testosterone release (pg/10 6 cells/day) Curcumin dose added (µg/ml medium)
11 Ovarian cell proliferation (accumulation of PCNA) is inhibited with: 45 - rooibos % of cells contained PCNA ginkgo 20 Rooibos dose added (µg/ml) % of cells contained PC CNA flaxseed Ginkgo dose added (µg/ml) % of cells contained PCNA green tea % oc cells contained PCNA Flaxseed dose added (µg/ml) Green tea AO dose added (µg/ml medium)
12 Ovarian cell apoptosis (accumulation of bax) is promoted with: - rooibos - ginkgo % of cells contained bax Rooibos dose added (µg/ml) ntained bax % of cells con flaxseed Ginkgo dose added (µg/ml) 590 effect of green tea AO on apoptosis of porcine GC % of cells contained bax green tea % of cells contained bax Flaxseed dose added (µg/ml) 45 Green tea AO dose added (µg/ml medium)
13 Ovarian cell leptin release is inhibited with: - rooibos Leptine release (ng/10 6 cells/day) 2,0 1,8 1,6 1,4 1,2 1,0 0,8 0,6 - ginkgo Rooibos dose added (µg.ml -1 ) 2,5 2,0 Leptine release (ng/10 6 cells/day) 1,5 1,0 0,5 0,0 - flaxseed Ginkgo dose added (µg.ml -1 ) 5,5 5,0 Leptine release (ng/10 6 cells/day) 4,5 4,0 3,5 3,0 2,5 2,0 Flaxseed dose added (µg.ml -1 )
14 Ovarian cell progesterone release is inhibited with: 2,6 2,4 2,2 - rooibos - ginkgo Progesterone release (mol/l cells/day) 2,0 1,8 1,6 1,4 1,2 1,0 0,8 0, Rooibos dose added (µg.ml -1 ) Progesterone release (mol/l cells/day) Ginkgo dose added (µg.ml -1 ) is stimulated with: Effect of green tea PP on progesterone secretion by porcine GC - green tea - yucca Progesterone release (ng/10 6 cells/day) Green tea PP dose added (µg/ml medium) Progesteron release (ng/10 6 tissue/day) Yucca dose added (µg/ml medium)
15 FOOD PLANTS CONTAIN PHYTOESTROGENS AND ANTIOXIDANTS - resveratrol, quercetin, diosgenin, daidzein. CAN THE PLANT ACTION BE EXPLAINED BY THE PRESENCE OF THESE MOLECULES? DO PLANT EXTRACTS AND PLANT MOLECULES HAVE SIMILAR ACTION?
16 Resveratrol 55 - inhibits proliferation (accumulation of PCNA) % of cells contained PCNA ng/ml 100ng/ml 1000ng/m Resveratrol dose added (µg.ml -1 medium) promotes apoptosis % of cells contained bax (accumulation of bax) 45 Resveratrol dose added (µg.ml -1 medium) Inhibits release of progesterone Progesterone release (ng.10-6 cells.day -1 ) Resveratrol dose added (µg.ml medium -1 )
17 Quercetin 55 - inhibits proliferation (accumulation of PCNA) % cells contained PCNA Quercetin dose added (µg.ml -1 ) - promotes apoptosis (accumulation of bax) % cells contained bax Quercetin dose added (µg.ml -1 )
18 Diosgenin : promotes proliferation (accumulation of PCNA) % of cells containing PCNA A A A 30 Diosgenin dose added (µg/ml) 90 - promotes apoptosis (accumulation of bax) % of cells containing Bax Diosgenin dose added (µg/ml)
19 Daidzein : 70 1A - promotes proliferation (accumulation of PCNA) % of cells containing PCNA a a Daidzein (µm) 80 1B - promotes apoptosis (accumulation of bax) x % of cells containing bax a a 30 Daidzein (µm) - Inhibits release of progesterone P4 secretion pg/10 6 cells/day C a a a 4 Daidzein (µm)
20 CAN PLANTS AND PLANT MOLECULES AFFECT OVARIAN RESPONSE TO HORMONAL STIMULATORS?
21 Daidzein blocks the stimulatory effect of FSH on : - proliferation PCNA 36 kda (accumulation of PCNA) 0 0,01 0,1 1 FSH dose added (IU/ml) 0 0,01 0,1 1 FSH dose added (IU/ml) + daidzein( µm) 25 2C a - release of progesterone P4 secretion pg/10 6 cells/d day b FSH b FSH + daidzein a b 0 0,00 0,01 0,10 1,00 FSH dose added (IU/ml medium) 25 2E - release of leptin Leptin secretion ng/10 6 cells/day b a FSH a ab a ab FSH + Daidzein -5 0,0 0,1 1,0 10,0 FSH dose added (ng/ml medium)
22 Quercetin blocks the stimulatory action of FSH on ovarian cell proliferation (accumulation of PCNA) FSH % cells contained PCNA 60 FSH and quercetin FSH and quercetin dose added (µg.ml -1 )
23 Resveratrol blocks the stimulatory effect of FSH on the release of: progesterone P4 secreti ion ng/10 6 cells/day B FSH + PF 046 FSH B A 20 0,00 0,01 0,10 1,00 FSH dose added (IU/ml medium) testosteron T secretion pg/10 6 cells/day B FSH FSH + PF 046 B A B A B 0 0,00 0,01 0,10 1,00 FSH dose added (IU/ml medium)
24 Curcumin blocks the anti-apoptotic effect of IGF-I 60 IGF-I + curcumin % of cells containe ed bax 30 B IGF-I alone B B 20 AB 10 IGF-I dose added (ng/ml medium)
25 Green tea blocks the stimulatory effect of IGF-I on progesterone release 30 Progesterone release (ng/10 6 ce ells/day) IGF-I + green tea PP IGF-I alone AB A AB 9 IGF-I dose added (ng/ml medium)
26 CAN PLANT PRODUCTS BE USED FOR IMPROVEMENT HORMONAL AND REPRODUCTIVE STATE?
27 Addition of yucca can promote hormones release by cultured rabbit ovarian tissue: - progesterone - testosteron - estradiol
28 Feeding with yucca increases rabbit plasma level of - oxytocin - prostaglandin F - prostaglandin E
29 Feeding with yucca increases rabbit Effect of yucca on conception rate (%) on rabbit does - conception rate Conception rate (average) Yucca dose added (g/100kg food) Effect of yucca on kindling rate (%) on rabbit does - kindling rate Kindling rate (average) Yucca dose added (g/100kg food)
30 CONCLUSIONS: 1. Food plants curcuma/turmeric, green tea, rooibos, ginkgo, flaxseed can directly suppress ovarian cell functions via - inhibition of proliferation, - promotion of apoptosis, - change in steroid and peptide hormone release - blockage of response to upstream hormonal stimulators (FSH, IGF-I).
31 CONCLUSIONS: 2. Phytoestrogens and antioxidants resveratrol and quercetin act similar to curcuma, green tea, rooibos, ginkgo and flaxseed. Can be active component of food plants?
32 CONCLUSIONS: 3. Phytoestrogens and antioxidants daidzein and diosgenin have both stimulatory and inhibitory effect on ovarian cell functions: - promote proliferation - promote apoptosis - suppress progesterone release - suppress the response to FSH Not responsible for the main food plant effect?
33 CONCLUSIONS: 4. Yucca promotes - steroid and peptide hormones release both in vivo and in vitro - fertility (conception and kindling rate). Can be useful for improvement reproduction?
34 COLLABORATION A.Kadasi, A. Stochmalova, A. Alexa, M. Foldesiova, A. Balazi, P. Chrenek (Slovakia) R. Grossmann (Germany) J. Kotwica (Poland)
35 THANK YOU!
36 Some effects of nutrition on physiological state Low calories diet - delays puberty - prolongs duration of life - activates immune system - prevents and inhibits infectious diseases depressions metabolic disorders reproductive disorders and infertility cancer
37 Some effects of nutrition on physiological state Phytoestrogens -stimulate puberty -Inhibite some metabolic (osteoporosis) and psychic (depression, agressivity) diseases -promote inflammation -promote or suppress reproductive disorders (meno- and andropause, infertility, ovulatory cycles) -promote or suppress cancer
38 Results RESULTS Results Inhibitedted effect of resveratrol and rapamycin on the expression of PCNA % of cells contained PCNA % of cells contained PCNA Resveratrol dose added (µg.ml -1 medium) Rapamycin dose added (µg.ml -1 medium)
39 RESULTS Results Stimulated effect of resveratrol and rapamycin on the expression of bax. % of cells contained bax % of cells contained bax Resveratrol dose added (µg.ml -1 medium) Rapamycin dose added (µg.ml -1 medium)
40 RESULTS Results Inhibited effect of resveratrol and rapamycin on progesterone release ) Progesterone release (ng.10-6 cells.day ) Progesterone release (ng.10-6 cells.day Resveratrol dose added (µg.ml medium -1 ) Rapamycin dose added (µg.ml -1 medium)
41 RESULTS Stimulated effect of resveratrol Results on testosterone release. Effect of rapamycin on testosterone release was not found day -1 ) Testosterone release (pg.10-6 cells Col 1 vs Col 2 Testosterone release (pg.10-6 cells s.day -1 ) Col 1 vs Col Resveratrol dose added (µg.ml -1 medium) Rapamycin dose added (µg.ml -1 medium)
42 Resveratrol : - inhibits the release of progesterone P4 secretion ng/10 6 cells/day ,0 0,1 1,0 10,0 Resveratrol dose added (µg/ml medium) promotes the release of testosteron T secretion pg/10 6 cells/day ,0 0,1 1,0 10,0 Resveratrol dose added (µg/ml medium)
43 Resveratrol inhibits proliferation (accumulation of PCNA) of ovarian cells 5 júcich PCNA % buniek obsahuj ng/ml 100 ng/ml 1000 ng/ml ng/ml Dávka resveratrolu - 36 K 0ng/ml 100ng/ml 1000ng/ml
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