April and May on the US east coast = temperature swings = confusing
and sickly weather. This year especially reminds me of the sobering
admonition from the ancient Chinese classic of medicine, <The Yellow
Emperor’s Inner Canon>: “when there is damage from cold in winter,
one suffers from warm diseases in spring (Dong shang yu han, chun bi bing wen)” (see Marta Hanson’s insightful book
on this subject). Seasonality is well known as a central preoccupation
in the Chinese medical tradition: the cosmic resonance of the body and
the larger world according to the quadruple division of the solar year –
the cyclic fluctuation of temperature, directionality of wind, and the
loci of corporal vulnerability that furnished essential cues for a
master practitioner of medicine.
But if etiology in Chinese medicine is classically understood as
seasonal, surely the therapeutics should also follow a seasonal rhythm?
To my surprise, a search for pre-modern monographs that contain the
keyword “four seasons” (sishi) yielded few results. In
addition, they tend to focus on agriculture (which of course also
follows a seasonal rhythm) or popular festivities around the year. I
decide to take a closer look at the latest text that featured “four
seasons” in its title – a title attributed to Qu You (1341-1427), Si shi yi ji (Auspicious
and Inauspicious Deeds in Four Seasons). I thought this text might
teach me something about how a learned scholar approached the notion of
seasonality in the early fifteenth century, and how that might align or
depart from the canonical medical model of seasonality.
The book consists of twelve chapters, each describing the dos and
don’ts for a specific month. I flipped to the chapter on the fourth
month (which corresponded roughly to this present moment in Western
calendar). I learned, to my surprise and delight, a ton of practical
advice with specific recipes: how to properly dry and insulate book and
painting cases before the advent of rainy season; “use eels that have
been sun-dried, burn them inside the house to thwart the thirst of
mosquitoes” (seems appropriate for New Jersey habitat); “wrap your
battle gears along with Sichuanese peppers (huajiao) or powder of Daphne flowers (yuanhua)
to prevent worm damage… wrap windshield collars and earmuffs and store
them in a vat, tightly seal it up, so as the fur will not fall off.”
After the first full moon this month, one “should drink mulberry wine”
to prevent “wind heat” illnesses (see Shigehisa Kuriyama’s discussion of wind in classical Chinese and Greek medicines). The recipe goes as follows:
Use Mulberries, get its juice of three dou (1 dou ~ 18 liter). White Honey four ounces (liang); Butter (suyou) one ounce; raw ginger juice two ounces.
Bring mulberry juice to a boil in a pot, and reduce its volume to three sheng (1 sheng = 1/10 dou), and then add honey, butter, and ginger juice. Add three drachm (qian) of salt and keep boiling till the texture is thick.
Store in porcelain utensils. Each time, take a small cup with wine. This effectively cures various wind-induced illnesses.
Not only does this sound completely delicious and doable to me, I
also realize how recipes like this are in fact completely grounded in
the seasonal rhythm of biological life (I just saw a friend posting the
harvest of fresh mulberries in her backyard in China).
In sum, what Qu You did in this book was to cull from a wide range of
medical and non-medical sources (a rough count yielded over 60
different titles) for hints and tips on how to live according to the seasons.
Some of his references were archaic almanacs that offered divinations
on the most auspicious dates to travel, have sex, trim your nails, or
remove grey hair, as well as dates one should abstain from such
activities. Some were quasi-ethnographic accounts of “customs” (fengsu)
in ancient cities that still lend to a viable reading as practical
guides to festivities. Still others draw from esoteric Daoist literature
on the preservation of vital essence (I have blogged on a related topic
here), a decision on Qu You’s part that raised many eyebrows both during his lifetime as well as centuries later. A Daoist talisman in Qu You, Si shi yi ji (1920 reprint of an 1836 edition).
We must remember that Qu lived through the Ming dynasty’s founder,
Hongwu emperor’s reign (1368-1398) – a period known for its austere
message of moral purity and simplicity. His fourth son, who usurped the
throne shortly after Hongwu’s death to become the Yongle emperor (r.
1404-1424), was not exactly friend of the letters either. Those were not
easy times for a literary aficionado with keen interests in morally
dubious subjects, and yet Qu You continued to compose and comment on
poetry, wrote short stories featuring ghosts and women, and collected
esoteric recipes. He even managed to publish those works, prefacing them
with loud self-defense of his moral stature. Qu eventually got into
trouble, endured decades of exile in the north, and yet again outlived
the Yongle emperor, who threw many a undisciplined scholars like him
into jail, by three years.
Perhaps the seasonal recipes did work well for him after all?
Purpose:
Cumulus cells have a critical role in normal oocyte development and
fertilization. Prunus cerasus is an anthocyanin rich berry and performs
strong antioxidant activity. The present study set to determine if
Prunus cerasus can affect expression of HAS2 (hyaluronan synthase 2) and
progesterone receptor in Cumulus cells and its consequences outcome of
the in vitro fertilization.
Methods:
60 female and 15 male adult mice were used for mating and IVF (in vitro
fertilization). Prunus cerasus extraction was added to the diet of
female mice for 30 days. Ovulation induction and oocytes collection were
done as routine. The cumulus cells were dissected apart, and the
expression of progesterone receptor and HAS2 was detected using RT-PCR
(real-time polymerase chain reaction). Fertilization rate was evaluated
by IVF. All data were analyzed using t-test.
Results:
Data was showed that expression of progesterone receptor and HAS2 in
cumulus cells of mice that received prunus cerasus increased. Moreover,
oocyte fertilization rate also increased significantly.
Conclusion:
Prunus cerasus as an antioxidant natural can become an important
medication for improving oocyte quality and opening new opportunities
for infertility treatment. It is concluded that Prunus cerasus
consumption could improve fertility rate by increasing progesterone
receptor and HAS2 activity in cumulus cells.
Infertility is defined as inability to obtain fertility after one year of unprotected regular intercourse.1-4
It has been reported that about 10% -15% of young couples suffer from
infertility. Of these, 40%-55% are due to female factors, 20%-30% are
due to male factors, and 15%-17% are unexplained infertility.1 Over the years, ART (assisted reproductive technology) has been a method for infertility treatment with different causes.5,6 In order to increase the success rate of fertilization, several herbal and chemical supplemental factors have been used.7 Because of less invasiveness and costliness nature of traditional medicine, recent research has focused on herbal use.8,9 Anthocyanin as secondary metabolite group of flavonoids has attracted much attention.10 Strong antioxidant activity was the best known property of flavonoids.11
Antioxidant activity of anthocyanin is associated with a variety of
health benefits including inflammation, cancer, atherosclerosis, and
diabetes.12
Recent studies have shown that reproductive processes may be influenced
by hypothalamic-pituitary-gonadal axis via scavenging free radicals.13-15
However, interest in antioxidant has recently been intensified because
of their possible effect on egg quality, fertilization, and pregnancy
rates.16,17
Antioxidant therapy has been attended to alleviate infertility, and
dietary antioxidant has been beneficial for female reproductive
disorder.18
Fruits including Sour cherry (Prunus cerasus), containing anthocyanin
and strong antioxidant activity, have attracted much attention.19
Prunus cerasus anthocyanins have a vast range of biochemical and
pharmacological effect, and have been recommended as nutritional
supplement.20
Granulosa cells consist of mural and cumulus cells in preovulatory
follicles. Mural cells are in the follicle wall and cumulus cell in
vicinal of oocyte.21
In the midcycle, increased level of LH result in ovulation by inducing
resumption of meiotic maturation, transformation of granulosa cells, and
expansion of cumulus-oocyte complex. The latter, is associated with
accumulation of hyaluronic acid-rich extra cellular matrix in cumulus
cells.22
During the process of cumulus expansion, the most important genes
involved in the formation of matrix hyaluronic are hyaluronic synthase2
(HAS2).23 HAS2 mRNA is necessary for cumulus expansion and signaling during ovulatory response.24
Cumulus expansion facilitates the fertilization, and the failure of
this process can lead to fertilization problem and is associated with
low potential for implantation and decline in the quality of oocyte.25,26
Paracrine factor under the influence of progesterone is required for
preimplantation embryonic development, but direct or indirect effect of
progesterone is not clear.27
Previous studies in our laboratory showed that the presence of
Progesterone receptor is essential for female fertility and ovulation
control.28,29
In this study, Prunus cerasus effect on progesterone receptor
expression, HAS2 in cumulus cells and the rate of in vitro fertilization
are evaluated.
Sour cherry were collected from the market and authenticated by a
botanist (School of Pharmacy, Tabriz University of Medical Sciences).
The extract, according to World Health Organization (WHO) protocol was
prepared. 500g of Sour cherry was shed-dried powdered and added to 5 Li
of 70°C ethanol (v/v) and left to macerate for 20 hours at room
temperature. The basin was rotated slowly during this time. After
filtration, ethanol at low pressure at 30 °C. The samples were stored
frozen at –20 °C until used.
Study population
A total of 60 female BALB/c mice (4-5weeks) and 15 adult male BALB/c
mice were obtained from the animal house of Tabriz University of Medical
Sciences (TUMS). For a period of 2 weeks prior to the experiment, the
mice were kept in animal room for adapting. 1.5 kg of mice food was
added to 1000ml of Prunus cerasus extract and mixed together completely.
The mixed diet let to dry in 2 days. Female mice were randomly
divided into two equal groups. In the control group, each animal
received diet without Prunus cerasus. In the experimental group, each
animal received 25g diet with Prunus cerasus for 4 weeks. After 4 weeks
for induction of ovulation 10 IU HMG/PMSG (pregnant mare serum
gonadotropin) and after 48 hours, 10 IU hCG (human chorionic
gonadotropin) was injected intraperitoneally.
Oocyte collection
One day post hCG injection, the mice were sacrificed and ovaries were
removed from the female mice. Mice ovaries (n=100) were obtained and
placed in sterile PBS and transported to the Tissue Engineering
Laboratory in two groups. The stromal tissue surrounding the ovaries was
removed, and oocyte were collected under flashing method and cultured
in dishes with Universal IVF culture medium (Origio-Medicult, 10300060)
under sterile mineral oil.
Isolation of cumulus cells
The cumulus cells surrounding the oocyte (COCS) were separated from
oocyte using hyaluronidase enzyme. Hyaluronidase enzyme causes the
connection of oocyte and cumulus cells to be released and facilitates
separation. Cumulus cells were transferred to another dish containing
medium. PBS was added for washing and centrifuged 2 times for 5 min.
After centrifuge, the pellet of cumulus cells was centrifuged for 1 h at
-20 °C and then stored at -80 °C until RNA isolation.
In vitro fertilization
Male mice were killed by cervical dislocation, and the sperm were
collected from the cauda epididymis by incubating the pieces of
epididymis in 37°C Co2 incubator for 20 min. The sperm sample was added
to the collected oocyte in the control group and experimental case
group. The fertility success rate was assessed based on the formation of
embryo.
Real time RT – PCR
RNA expression was indicated using real time RT-PCR assay to measure
HAS2 and PGR mRNA. The primers, used in PCR are presented in Table 1.
Internal control gene (GAPDH) was used for normalization of the result.
Total RNA of HAS2 and progesterone receptor were extracted using RNeasy
Micro kit from cumulus cell. The cDNA were synthesized, and primer
design was done. Real time- polymerase chain reaction (RT-PCR) was
performed for gene expression analysis.
All statistical analysis was done using SPSS software version 22 and
independent sample T-test. P values≤0.05 were considered statistically
significant.
Effect of Prunus cerasus on HAS2 and PGR expression in cumulus cells
Expression of HAS2 and PGR mRNA in cumulus cells, isolated from mice
oocyte, was assessed by quantitative real time PCR. There was a
significant increase in HAS2 mRNA in the experimental group compared to
the control (Figure 1). However, the expression level of PGR gene in experimental group was increased compared to the control group (Figure 2).
IVF was performed in both control and experimental groups, and 100
oocytes were used per group. Counting the number of embryos revealed
that in the control group which was not received Prunus cerasus, 77% of
oocytes were developed into embryos, while in the experimental group
receiving Prunus cerasus for 4 weeks, 89% of oocytes were developed into
embryos (P≤0.0001). Developed embryos in the experimental group are
shown in Figure 3.
The results revealed that the addition of Prunus cerasus to laboratory
diet resulted in increase of expression of HAS2 and PGR gene in cumulus
cells and fertility success rate. Since Prunus cerasus contain
significant levels of anthocyanins, strong antioxidant activity can
influence fertility.10
Significant increase in the expression of HAS2 and PGR gene in the case
group compared to the control group could be due to the loss of
oxidative stress produced by antioxidant property of Prunus cerasus,
which can also be justified for increasing fertility rate. Follicular
fluid contains a lot of antioxidants that protect oocyte against free
radicals. The disorder in pro oxidant/antioxidant system in follicular
fluid can damage oocyte′s DNA, and cytoskeleton of membrane can lead to
impaired fertility.30,31 Use of antioxidant rich food prevents progression of diseases related to oxidative stress.32
Free radicals that are indication of oxidative stress in many diseases
are highly reactive, toxic, and short-lived molecules that hurt DNA,
protein, lipid, and carbohydrates within the tissue.32
Oxidative stress specially affects microenvironment of oocyte, which
has a detrimental influence on follicular development, ovulation, oocyte
quality, implantation, and early embryonic development.33In the study of Damar and Eksi and other studies, anthocyanin and antioxidant capacity of Prunus cerasus is evidenced.11,34,35 Showell MG et al.’s study on the effects of antioxidants on fertility was contradictory, needing further investigation.32 Unlike the present study, Jozwick et al. did not report any relationship between free radical marker and pregnancy rate.36
The results of the present study is in agreement with Polak et al. who
state that the concentration of antioxidant in peritoneal fluid of
patients with unexplained infertility was significantly lower than the
its concentration in fertile patients.37
Study of Agawarl et al is in line with the results of current study
demonstrating that pregnancy rates in patients undergoing IVF, using
antioxidant-supplemented media, were higher than standard media without
antioxidants.32
Since studies have shown that antioxidants affect oocyte quality, high
degree of expansion of cumulus cells is directly related to oocyte
quality and expression of HAS2 and PGR gene is necessary for cumulus
expansion. According to the results it could be postulated, it is
antioxidant capacity of prunus cerasus that resulted in increased gene
expression and fertilization rate.
This article is resulted from a research proposal leading to the thesis
of Fatemeh Namvar, MSc student of anatomical sciences and was approved
by research deputy of Tabriz University of Medical Sciences, Tabriz,
Iran.
4. Uyar A, Torrealday S, Seli E. Cumulus and granulosa cell markers of oocyte and embryo quality. Fertil Steril. 2013;99(4):979–97. doi: 10.1016/j.fertnstert.2013.01.129. [PMC free article][PubMed][Cross Ref]
5. Bower C, Hansen M. Assisted reproductive technologies and birth outcomes: Overview of recent systematic reviews. Reprod Fertil Dev. 2005;17(3):329–33. [PubMed]
6. Talaulikar
VS, Arulkumaran S. Maternal, perinatal and long-term outcomes after
assisted reproductive techniques (art): Implications for clinical
practice. Eur J Obstet Gynecol Reprod Biol. 2013;170(1):13–9. doi: 10.1016/j.ejogrb.2013.04.014. [PubMed][Cross Ref]
7. Haggarty
P, McCallum H, McBain H, Andrews K, Duthie S, McNeill G. et al. Effect
of b vitamins and genetics on success of in-vitro fertilisation:
Prospective cohort study. Lancet. 2006;367(9521):1513–9. doi: 10.1016/S0140-6736(06)68651-0. [PubMed][Cross Ref]
8. de
Boer HJ, Cotingting C. Medicinal plants for women's healthcare in
southeast asia: A meta-analysis of their traditional use, chemical
constituents, and pharmacology. J Ethnopharmacol. 2014;151(2):747–67. doi: 10.1016/j.jep.2013.11.030. [PubMed][Cross Ref]
9. Ried K, Stuart K. Efficacy of traditional chinese herbal medicine in the management of female infertility: A systematic review. Complement Ther Med. 2011;19(6):319–31. doi: 10.1016/j.ctim.2011.09.003. [PubMed][Cross Ref]
10. Blando F, Gerardi C, Nicoletti I. Sour cherry (prunus cerasus l) anthocyanins as ingredients for functional foods. J Biomed Biotechnol. 2004;2004(5):253–8. doi: 10.1155/S1110724304404136. [PMC free article][PubMed][Cross Ref]
11. Kirakosyan
A, Seymour EM, Noon KR, Llanes DEU, Kaufman PB, Warber SL. et al.
Interactions of antioxidants isolated from tart cherry (Prunus cerasus)
fruits. Food Chem. 2010;122(1):78–83. doi: 10.1016/j.foodchem.2010.02.017. [Cross Ref]
12. Ataie-Jafari
A, Hosseini S, Karimi F, Pajouhi M. Effects of sour cherry juice on
blood glucose and some cardiovascular risk factors improvements in
diabetic women: A pilot study. Nutr Food Sci. 2008;38(4):355–60. doi: 10.1108/00346650810891414. [Cross Ref]
13. Nsonwu-Anyanwu
AC, Charles-Davies MA, Taiwo VO, Li B, Oni AA, Bello FA. Female
reproductive hormones and biomarkers of oxidative stress in genital
chlamydia infection in tubal factor infertility. J Reprod Infertil. 2015;16(2):82–9. [PMC free article][PubMed]
14. Hassanzadeh
K, Roshangar L, Habibi-asl B, Farajnia S, Izadpanah E, Nemati M. et al.
Riluzole prevents morphine-induced apoptosis in rat cerebral cortex. Pharmacol Rep. 2011;63(3):697–707. [PubMed]
15. Roushangar
L, Rad JS. Ultrastructural alterations and occurrence of apoptosis in
developing follicles exposed to low frequency electromagnetic field in
rat ovary. Pak J Biol Sci. 2007;10(24):4413–9. [PubMed]
16. Howatson
G, Bell PG, Tallent J, Middleton B, McHugh MP, Ellis J. Effect of tart
cherry juice (prunus cerasus) on melatonin levels and enhanced sleep
quality. Eur J Nutr. 2012;51(8):909–16. doi: 10.1007/s00394-011-0263-7. [PubMed][Cross Ref]
17. Haidari
F, Jr., Mohammad Shahi M, Keshavarz SA, Rashidi MR. Inhibitory effects
of tart cherry (prunus cerasus) juice on xanthine oxidoreductase
activity and its hypouricemic and antioxidant effects on rats. Malays J Nutr. 2009;15(1):53–64. [PubMed]
18. Visioli F, Hagen TM. Antioxidants to enhance fertility: Role of enos and potential benefits. Pharmacol Res. 2011;64(5):431–7. doi: 10.1016/j.phrs.2011.06.021. [PubMed][Cross Ref]
19. Piccolella
S, Fiorentino A, Pacifico S, D'Abrosca B, Uzzo P, Monaco P. Antioxidant
properties of sour cherries (prunus cerasus l.): Role of colorless
phytochemicals from the methanolic extract of ripe fruits. J Agric Food Chem. 2008;56(6):1928–35. doi: 10.1021/jf0734727. [PubMed][Cross Ref]
20. Bak
I, Lekli I, Juhasz B, Varga E, Varga B, Gesztelyi R. et al. Isolation
and analysis of bioactive constituents of sour cherry (prunus cerasus)
seed kernel: An emerging functional food. J Med Food. 2010;13(4):905–10. doi: 10.1089/jmf.2009.0188. [PubMed][Cross Ref]
21. McKenzie
LJ, Pangas SA, Carson SA, Kovanci E, Cisneros P, Buster JE. et al.
Human cumulus granulosa cell gene expression: A predictor of
fertilization and embryo selection in women undergoing ivf. Hum Reprod. 2004;19(12):2869–74. doi: 10.1093/humrep/deh535. [PubMed][Cross Ref]
22. Curry TE, Jr. Adamts1 and versican: Partners in ovulation and fertilization. Biol Reprod. 2010;83(4):505–6. doi: 10.1095/biolreprod.110.087056. [PubMed][Cross Ref]
23. Sugiura
K, Su YQ, Eppig JJ. Targeted suppression of has2 mrna in mouse cumulus
cell-oocyte complexes by adenovirus-mediated short-hairpin rna
expression. Mol Reprod Dev. 2009;76(6):537–47. doi: 10.1002/mrd.20971. [PMC free article][PubMed][Cross Ref]
24. Adriaenssens
T, Segers I, Wathlet S, Smitz J. The cumulus cell gene expression
profile of oocytes with different nuclear maturity and potential for
blastocyst formation. J Assist Reprod Genet. 2011;28(1):31–40. doi: 10.1007/s10815-010-9481-9. [PMC free article][PubMed][Cross Ref]
25. Gebhardt
KM, Feil DK, Dunning KR, Lane M, Russell DL. Human cumulus cell gene
expression as a biomarker of pregnancy outcome after single embryo
transfer. Fertil Steril. 2011;96(1):47–52 e2. doi: 10.1016/j.fertnstert.2011.04.033. [PubMed][Cross Ref]
26. Hong
SJ, Chiu PC, Lee KF, Tse JY, Ho PC, Yeung WS. Cumulus cells and their
extracellular matrix affect the quality of the spermatozoa penetrating
the cumulus mass. Fertil Steril. 2009;92(3):971–8. doi: 10.1016/j.fertnstert.2008.07.1760. [PubMed][Cross Ref]
29. Kim J, Bagchi IC, Bagchi MK. Control of ovulation in mice by progesterone receptor-regulated gene networks. Mol Hum Reprod. 2009;15(12):821–8. doi: 10.1093/molehr/gap082. [PMC free article][PubMed][Cross Ref]
30. Agarwal A, Gupta S, Sharma R. Oxidative stress and its implications in female infertility - a clinician's perspective. Reprod Biomed Online. 2005;11(5):641–50. [PubMed]
31. Agarwal A, Allamaneni SS. Role of free radicals in female reproductive diseases and assisted reproduction. Reprod Biomed Online. 2004;9(3):338–47. [PubMed]
32. Meghashri S, Kumar HV, Gopal S. Antioxidant properties of a novel flavonoid from leaves of Leucas aspera. Food Chem. 2010;122(1):105–10. doi: 10.1016/j.foodchem.2010.02.023. [Cross Ref]
33. Bahadori
MH, Ghasemian F, Ramezani M, Asgari Z. Melatonin effect during
different maturation stages of oocyte and subsequent embryo development
in mice. Iran J Reprod Med. 2013;11(1):11–8. [PMC free article][PubMed]
34. Damar I, Eksi A. Antioxidant capacity and anthocyanin profile of sour cherry (prunus cerasus l.) juice. Food Chem. 2012;135(4):2910–4. doi: 10.1016/j.foodchem.2012.07.032. [PubMed][Cross Ref]
35. Wang H, Nair MG, Strasburg GM, Booren AM, Gray JI. Antioxidant polyphenols from tart cherries (prunus cerasus) J Agric Food Chem. 1999;47(3):840–4. [PubMed]
36. Pasqualotto
EB, Agarwal A, Sharma RK, Izzo VM, Pinotti JA, Joshi NJ. et al. Effect
of oxidative stress in follicular fluid on the outcome of assisted
reproductive procedures. Fertil Steril. 2004;81(4):973–6. doi: 10.1016/j.fertnstert.2003.11.021. [PubMed][Cross Ref]
37. Polak
G, Koziol-Montewka M, Gogacz M, Blaszkowska I, Kotarski J. Total
antioxidant status of peritoneal fluid in infertile women. Eur J Obstet Gynecol Reprod Biol. 2001;94(2):261–3. [PubMed]
Articles from Advanced Pharmaceutical Bulletin are provided here courtesy of Tabriz University of Medical Sciences
Received: 15 December 2016 / Accepted: 4 February 2017 / Published: 9 February 2017
Abstract
:
Objective: To highlight a natural
approach to coexisting oligomenorrhea, subfertility, luteal phase
insufficiency and multiple fibroids cohesively when in vitro
fertilisation (IVF) has failed. Case Presentation: A 43-year-old
woman with diminished ovarian reserve and multiple uterine fibroids had
previously been advised to discontinue IVF treatment. According to
Chinese Medicine diagnosis, herbal formulae were prescribed for
improving age-related ovarian insufficiency as well as to control the
growth of fibroids. After 4 months of treatment, the patient’s menstrual
cycle became regular and plasma progesterone one week after ovulation
increased from 10.9 nmol/L to 44.9 nmol/L. After 6 months, she achieved a
natural conception, resulting in a live birth of a healthy infant at an
estimated gestational age of 40 weeks. Conclusions: The
successful treatment with Chinese Herbal Medicine for this case
highlights a natural therapy to manage infertility due to ovarian
insufficiency and multiple fibroids after unsuccessful IVF outcome.
Keywords:
Chinese herbal medicine; diminished ovarian reserve; infertility; in vitro fertilization; uterus fibrosis
1. Introduction
Advanced maternal age contributes to subfertility due to diminished ovarian reserve (DOR) and decreased oocyte quality [1]. Reduced fertility potential may also be attributable to uterine fibroids which are more prevalent in women aged over 30 [2] depending on their size and location, which can affect embryo implantation [3].
Indeed, women with DOR and fibroids have a significantly low fertility
rate despite advanced in vitro fertilization (IVF) techniques [4].
Given the limited treatment options after failure in IVF with DOR,
these women often seek alternative therapy, such as Traditional Chinese
medicine (TCM) to reserve adoption or using donor’s egg as the last
option.
Chinese medicinal herbs have been used
for more than two thousand years to treat gynaecological disorders
including infertility. In the last decades, experimental and clinical
studies have shown that these herbal ingredients can regulate
gonadotropin-releasing hormone and ovarian sex hormone levels to induce
ovulation and promote blood flow to the ovaries to improve ovarian
reserve [5].
TCM is unique as it applies a formula with multiple natural ingredients
that are capable of counteracting complex endocrine and reproductive
disorders. Here, we present a successful live birth with Chinese herbal
medicine (CHM) treatment to a 43-year-old nulliparous woman with DOR and
uterus fibroids after failed conception from IVF.
2. Case Presentation
A
43-year-old woman visited our clinic for CHM treatment of infertility.
She reached menarche at 13 years of age and at this time her menstrual
cycle was regular with a normal menstrual flow. Oral contraceptive pills
were taken from age 21–23 and she had no previous pregnancies prior to
our treatment. She started trying to conceive at age 40 and her husband,
aged 41, had no remarkable health problems and normal semen analysis as
defined by the World Health Organization 2010 criteria [6].
After two years of trying to achieve natural conception, she was
referred to a fertility specialist for IVF treatment. Prior to the
initiation of treatment, a pelvic ultrasound showed that the right ovary
was not visible and the left ovary measured 2.7 mL with only 3 antral
follicles and both fallopian tubes were clearly patent. The uterus was
bulky measuring at 103 × 125 × 68 mm with multiple fibroids including
two larger subserosal fibroids. Serum anti-mullerian hormone (AMH) level
of <3 pmol/L was within the <25% percentile for correlated age.
Hormone testing on day 5 of her menstrual cycle identified follicle
stimulating hormone (FSH) levels of 1.9 IU/L (normal range 1.5–10),
luteinising hormone (LH) levels of 2.8 IU/L (normal range 2.0–12),
Oestradiol (E2) levels of 913 pmol/L (normal range <320) and
Progesterone (P) levels of <0.5 nmol/L (normal range 0.3–4.0).
According to the ESHRE criteria this patient had suspected poor ovarian
response [7].
Therefore, controlled ovarian hyperstimulation (COH) with daily FSH
injections from day 3 of her cycle was applied for oocyte retrieval,
however, only one mature oocyte was collected, and she failed to
conceive after fertilized embryo transferring. The fertility specialist
therefore advised her to seek egg donation or cease IVF. The patient
consequently sought out treatment at our Chinese Medicine clinic in hope
of achieving fertility.
At the time the
patient visited the clinic her menstrual cycles ranged between 24 and 42
days. According to the diagnosis, basic formula containing 10 herbs was
prescribed (Table 1),
aiming to (1) improve diminished ovarian function and regulate the
menstrual cycle and (2) restrain the growth of uterus fibroids which
corresponds to tonifying vital essence and regulating and nourishing
blood in TCM.
Table 1.
Description of Chinese Herbal Formula and Possible Pharmacological Effects [8,9].
The herbal mixture was decocted into 500
mL and divided into two drinks daily. Ovulation was monitored by basal
body temperature (BBT) and ovulation prediction kit. Plasma progesterone
was measured one week of ovulation revealed by BBT chart and LH surge.
Following initial consult and first month treatment with CHM, the
patient underwent hormone testing one week after ovulation: P was 10.9
nmol/L, LH was 0.8 IU/L (normal range 2.0–12), FSH was 2.0 IU/L (normal
range 1.5–10), E2 was 184 pmol/L (normal range 125–1300). After 12 weeks
of treatment, the patient’s menstrual cycle became regular with cycle
lengthen between 30 and 35 days. Pelvic ultrasound on day 9 of cycle
indicated ovaries of both sides were normal in size and uterine size was
reduced with measuring 84 × 62 × 56 mm as well as decreased sizes of
intramural and subserosal fibroids. A blood test was also performed at
day 23 of her menstrual cycle (about one week after ovulation) and
revealed improvement of progesterone levels from 10.9 up to 28.1 nmol/L.
From week 13, the formula was slightly modified according to the follicular phase by adding Radix Polygoni multiflori (He Shou Wu) 10 g to nourish blood, and in the luteal phase by adding Radix Dipsaci (Xu Duan) 12 g and Cortex Eucommiae ulmoidis (Du Zhong) 9 g to assist in embryo implantation. Anti-fibrosis herb, Prunellae vulgaris and Rhizoma Curcuma phaeocaulis
had been ceased after 4 months of treatment. After 5 months of taking
CHM, although AMH was at 1.3 pmol/L, plasma progesterone levels after
one week of ovulation went from 10.9 nmol/L to 44.9 nmol/L, suggesting
that the quality of oocyte and function of corpus luteum had been
improved. After 6 months of treatment with CHM, she achieved natural
conception, resulting in a live birth of a healthy female infant
weighing 3350 g at an estimated gestational age of 40 weeks through
caesarean section. No modifications to the lifestyle of the patient were
made during this period.
3. Discussion
The
conventional approaches to sterility include ovulation induction,
intrauterine insemination and IVF. In this case, woman with advanced
maternal age, IVF may be the most successful technique to achieve
conception. However, because of the DOR, this patient presented a very
poor response to COH, resulting in unsuccessful IVF. Consequently, the
patient turned to CHM. Our treatment achieved pregnancy and a healthy
live birth with Chinese herbal formulae containing herbs supporting the
folliculogenesis cycle, invigorating blood, improving microcirculation,
and resolving masses to benefit both ovary and uterus.
Chinese
herbal medicines have long been used for the treatment of infertility.
Numerous studies demonstrated that CHM could regulate the
gonadotropin-releasing hormone (GnRH) to induce ovulation and improve
the uterus blood flow and menstrual changes of endometrium [10,11].
An advantage of CHM treatment is that it utilizes individualized
formulas tailored to a patient’s condition. In this case, non-invasive
treatment with CHM targeted two major causes underlying her infertility
simultaneously, including poor quality of oocyte due to diminished
ovarian reserve and uterine fibroids. The successful pregnancy
eventually achieved through improvement of uterus environment and
enhanced ovarian function.
It is generally
accepted that submucousal fibroids but not subserosal fibroids have a
negative impact on fertility by the virtue of their involvement in the
endometrial cavity. This patient had multiple large subserosal and
intramural fibroids of equal proportions. It has also been confirmed
that myomas have higher levels of aromatase converting estrogens to
estradiol, resulting in an imbalance of estradiol and progesterone,
which is detrimental to embryo implantation and contributes to
miscarriage [3].
In this infertility case, uterine fibroids have been considered as a
treatment target because the estradiol levels of 913 pmol/L (normal
range <320) was significantly high at follicular phase and there was
also evidence of a bulky uterus caused by multiple fibroids. The large
subserosal and intramural fibroids affect the endometrium homogeneous
and may have negative impact on the embryo implantation on her natural
conception and during IVF procedure. It is also a concern that these
fibroids may aggressively grow during the pregnancy because of the
ovarian hormone surge, subsequently affecting foetus development.
The herbal formula used in this case is able to balance yin and yang,
nourish blood and invigorate blood circulation from a TCM perspective.
In medical science, the CHM formula may restore ovarian function through
improving blood flow to reproductive organs, regulating hormone
secretions, lowering systemic inflammation as well as having anti-tumour
properties. Among of them, Radix Salvia miltiorrhizae, Radix Paeoniae lactiflorae, Spica Prunellae vulgaris and Rhizoma Curcuma phaeocaulis
have strong effects on blood stasis which is involved in mass
(fibroids) formation in the reproductive system. Pharmacological studies
have shown that these herbs reduce inflammation and inhibit neoplasia [12,13]. Meanwhile, the combination of Radix Rehmannia glutinosa, Semen Cuscutae chinensis, Fructus mori, Fructus lycii, Radix Morindae officinalis are commonly used in infertility due to ovarian factors, such as PCOS and primary ovarian insufficiency [13,14,15,16]. However, the precise mechanisms of Chinese herbs remain unclear with further research warranted.
After
one month of treatment, day 6 oestradiol levels of 930 pmol/L reduced
to 387 pmol/L. At the month five of the treatment, plasma progesterone
from 10.9 nmol/L raised to 44.9 nmol/L, indicating improved ovarian
function. Meanwhile, the size of the fibroids had shrunk with an overall
reduced size of the uterus. These results indicate that CHM treatment
inhibited fibroid growth and improved ovarian function, and may suggest
enhanced quality of oocyte, evidenced by regularity of menstrual cycle,
mid-cycle ovulation and elevated progesterone levels at luteal phase.
Evidence suggests that oocyte quality profoundly affects fertilisation
and embryo development [17].
The treatment achieved this woman’s natural pregnancy, suggesting the
improvement of oocyte quality. The limitation in this case observation
is that other assessments for oocyte quality such as morphological
character of oocyte and corpus luteum and other biomarkers, such as
mitochondrial status and glucose-6-phosphate dehydrogenase 1 activity
and apoptosis of follicular cells, were not measured.
Infertility
is a life-altering burden and the disorder affects a couple’s emotional
health and wellness. Given the limited treatment options after failure
in IVF, women with DOR who intend to get pregnant have to rely on
assisted reproductive technology using donor eggs. The procedure is
costly and only has a live birth rate of ~30% [4]
and contributes to psychological distress. The results of this case
demonstrate that CHM is capable of targeting multiple reproductive
abnormalities involved in infertility. More specifically, herbal
medicine can improve ovarian function and prevent inevitable miscarriage
related to luteal phase defect and multiple fibroids. The successful
treatment with CHM for this case highlights the potential of a natural
approach to coexisting oligomenorrhea, subfertility, luteal phase
insufficiency and multiple fibroids cohesively. CHM therapy offers a
hope for aged women who have failed IVF cycles and decide to pursue
parenthood with their own oocytes. The repeatability of CHM on
infertility should be warranted through rigorously designed clinical
trials.
Acknowledgments
We are thankful to our patient for her willingness to use her anonymized data for this publication.
Nelson, L.M. Clinical practice. Primary ovarian insufficiency. N. Engl. J. Med.2009, 360, 606–614. [Google Scholar] [CrossRef] [PubMed]
Zhang, C.; Xu, X. Advancement in the treatment of diminished ovarian reserve by traditional Chinese and Western medicine. Exp. Ther. Med.2016, 11, 1173–1176. [Google Scholar] [CrossRef] [PubMed]
Coope,
T.G.; Noonan, E.; von Eckardstein, S.; Auger, J.; Baker, G.; Behre,
H.M.; Haugen, T.B.; Kruger, T.; Wang, C.; Mbizvo, M.T.; et al. World
Health Organization Reference Values for Human Semen Characteristics. Hum. Reprod. Update2010, 16, 231–245. [Google Scholar] [CrossRef] [PubMed]
Ferraretti,
A.P.; La Marca, A.; Fauser, B.C.; Tarlatzis, B.; Nargund, G.;
Gianaroli, L.; ESHRE Working Group on Poor Ovarian Response Definition.
ESHRE consensus on the definition of ‘poor response’ to ovarian
stimulation for in vitro fertilization: The Bologna criteria. Hum. Reprod.2011, 26, 1616–1624. [Google Scholar] [CrossRef] [PubMed]
Chen, J.K.; Chen, T.T. Chinese Medical Herbology and Pharmacology, 1st ed.; Art of Medicine Press Inc.: City of Industry, CA, USA, 2001. [Google Scholar]
Shen, Y.J. Pharmacology of Chinese Herbal Medicine, 1st ed.; People’s Medical Publishing House: Beijing, China, 2000. [Google Scholar]
Huang, S.T.; Chen, A.P. Traditional Chinese medicine and infertility. Curr. Opin. Obstet. Gynecol.2008, 20, 211–215. [Google Scholar] [CrossRef] [PubMed]
Lee,
S.H.; Kwak, S.C.; Kim, D.K.; Park, S.W.; Kim, H.S.; Kim, Y.S.; Kim, Y.;
Lee, D.; Lee, J.W.; Lee, C.G.; et al. Effects of Huang Bai (Phellodendri Cortex) and Three Other Herbs on GnRH and GH Levels in GT1–7 and GH3 Cells. Evid. Based Complement. Alternat. Med.2016, 2016, 9389028. [Google Scholar] [CrossRef] [PubMed]
Chen,
X.X.; Lin, W.L.; Yeung, W.F.; Song, T.H.; Lao, L.X.; Zhang, Y.B.; Meng,
W. Quality and safety control of Tumor-Shrinking Decoction (TSD), a
Chinese herbal preparation for the treatment of uterine fibroids. Biotechnol. Appl. Biochem.2015. [Google Scholar] [CrossRef] [PubMed]
Hu,
Q.; Noor, M.; Wong, Y.F.; Hylands, P.J.; Simmonds, M.S.; Xu, Q.; Jiang,
D.; Hendry, B.M.; Xu, Q. In vitro anti-fibrotic activities of herbal
compounds and herbs. Nephrol. Dial. Transplant.2009, 24, 3033–3041. [Google Scholar] [CrossRef] [PubMed]
Hung,
Y.C.; Kao, C.W.; Lin, C.C.; Liao, Y.N.; Wu, B.Y.; Hung, I.L.; Hu, W.L.
Chinese Herbal Products for Female Infertility in Taiwan: A
Population-Based Cohort Study. Medicine (Baltimore)2016, 95, e3075. [Google Scholar] [CrossRef] [PubMed]
Ong, M.; Peng, J.; Jin, X.L.; Qu, X. Chinese Herbal Medicine for the Optimal Management of Polycystic Ovary Syndrome. Am. J. Chin.2017, in press. [Google Scholar]
Chao, S.L.; Huang, L.W.; Yen, H.R. Pregnancy in premature ovarian failure after therapy using Chinese herbal medicine. Chang. Gung Med. J.2003, 26, 449–452. [Google Scholar] [PubMed]
