During this webinar, Dr Christos Roukoudis, Gynaecologist & Fertility Specialist at IVF-Life Alicante, discussed the ovarian rejuvenation method, how it works, its benefits and whether it is recommended for older patients.
Dr Roukoudis talked about the PRP and if it’s a good option for patients of advanced reproductive age, so the patients over 40. Dr Roukoudis explained that time is relative, and for all reproductive specialists and anyone who have a desire to have children, it’s the biggest obstacle. Women in general around the globe postpone childbirth until later in life, and due to this fact, they are more likely to face the problem of ovarian insufficiency by the time they are ready to have children. Moreover, the tendency of the unwanted influence of environment and social factors leads to an increase in the incidence of early menopause. As we all know, female age remains the most important and limiting factor of success in both spontaneous conception and assisted reproduction treatment, largely to a loss of ovarian follicles and the impact on the quality of the eggs.
As the graphic shows, from the puberty onwards, the process is faster, and in the time of 40 years, more or less 500 ovulations occur, and in each ovulation, 1000 oocytes are getting recruited, but in each round, only 1 gets selected, the rest get buried inside the ovary. The result is that the live birth rate, especially in women over 40 decreased significantly. Alongside the reduction in the numbers of the oocytes, we also have ageing oocytes, and those are more prone to errors in the DNA synthesis. The cell divisions result in increased aneuploidy rates, so embryos that don’t have the proper amount of chromosomes. A very important role in the decrease in the live birth rate plays other reasons, for example, alternate expression of the immune system with advanced reproductive age.
Our main goal is to enable women to become mothers with their eggs. We know the problems we are dealing with, and it relies on the number of eggs, but also the quality of the eggs. Therefore, we try to overcome this issue, and the solution that has excellent results, especially when we speak about advanced reproductive age, is egg donation. Egg donation has a very high success rate at our clinic (IVF-Life Alicante), after 3 transfers, the live birth rate is around 80%. However, this solution can cause significant psychological, financial, but also religious beliefs for some couples. There are also some legal restrictions in many countries, and for all those reasons, the possibility to restore the variable function is crucially important, and we seek a reliable way to do this.
Nowadays, there are plenty of things proposed, there are several ongoing trials that are investigated to rescue or rejuvenate the oocytes and restore fertility in women with menopause or with ovarian insufficiency. There is an option of in vitro oocyte activation, stem cell therapy, and also others exist that put some ethical point of view on topics, for example, germ cell line manipulation that is not justifiable. The bottom line is that all of those are expensive, complicated methods, and in the end, the success rate is the most important thing, and these are still very low. Therefore, an alternative is proposed, and so the PRP (Platelet-Rich Plasma), which is much cheaper and easier to apply without risks, various studies show that the results are very promising.
How are the platelets achieved? Platelets are small nuclear-cytoplasmic fragments derived from bigger cells, and although, traditionally, age is responsible for stopping bleeding from forming a blood clot, platelets attach to the wall and stop the bleeding. They also provide very important elements for tissue regeneration. Those are growth factors that are inside. There are over 15 000 proteins within the platelets, and growth factors are stored in the form of granules, and those molecules play an important role in healing tissues.
PRP stands for Platelet-Rich Plasma, it’s composed of platelets in a small volume of plasma containing a high concentration of growth factors and other cytokines. It was first used in open-heart surgery with the same mindset, which was to provide some cell regeneration. It’s now widely used in various fields. How does it work? There are several explanations, the first is that the growth factors inside activate the stem cells that are inside the ovary, and they produce new follicles. Another possible explanation is that some amount of dormant follicles are kept in the ovary, and some factors inside the platelets can promote the recruitment of those follicles. The growth factors inside get injected inside the ovary, and they increase the blood perfusion. They influence the hormones locally, and they also provide nutrients to the eggs inside the follicles.
The candidates for the PRP are women of lower ovarian reserve, and the majority are women in menopause, the perimenopause, in advanced reproductive age. Then there are also women with premature ovarian failure (POF), so women that came into menopause before the age of 40.
The PRP process means that we collect the blood, separate the platelets and then take it and are ready to inject it. We apply it inside, in the operating theatre, it is ultrasound-guided. The process is very similar to egg retrieval.
The most important thing in evaluating if treatment, especially experimental treatment, could be useful is to perform studies and see if the results are encouraging or show the opposite. The first study is from a group in Turkey, they showed the results from women who came into menopause before the age of 40. In this group, around 311 women between 24-and 40 were recruited, and those women were injected with PRP. Around 7% were able to conceive spontaneously, and around 64% developed new follicles, although they were not able to see follicles before. Around 28% had no antral follicles, so the PRP didn’t work at all.
The next study was from the same group in Turkey, presented in 2022, where they recruited around 510 women between the age range 30 and 45 who were diagnosed with a poor ovarian reserve (POR). They saw that after the PRP injection, around 4% conceived spontaneously, around 93% had some eggs, so they performed an IVF, and from that 93%, they saw that almost 312, it was 66% generated embryos and had a transfer. Around 70% of those achieved a pregnancy, and 11% had a live birth. The most important part is this 11.4%. The most important thing with those studies is to understand how many of those from 11% would achieve the pregnancy without the PRP. If the PRP was a real game-changer.
In another study, there were 83 women recruited, and those women had a low ovarian reserve. The majority of the women were over 40, and they applied PRP. 46 women received PRP, and the other 37 functioned as a control group, so they didn’t receive anything. When they compared the results, they saw that 26% versus 5% in the control group had a clinical pregnancy. Regarding a higher AMH, ovarian reserve marker, the control group with the PRP had a higher percentage, it was 24% against 5%. Keep in mind that AMH has also natural fluctuations, you can measure it twice, and you can see a difference from cycle to cycle, especially if we have an AMH at 0.1, then it could be 0.18 without doing anything. The most important thing from this study is that there was no difference in the rates of first-trimester miscarriage and live birth between those 2 groups. The impact of PRP in this study was not high.
In the next study, PRP was also performed. There was a group of around 80 patients, and among those 80 patients, 51 had an increased AMH. This study showed if we can see higher ovarian reserve markers, especially AMH, after the application of the PRP after 4 weeks. It was the first study that showed a significant increase. This study, in general, proved that PRP seems to provide some better results and can increase ovarian reserve.
Another study involved 38 women between 31 and 45, all of them had a low ovarian reserve and 2 unsuccessful attempts to receive the oocytes through IVF. They did a single PRP treatment, and the studies showed a significant improvement in the hormone levels. Some of them had a much higher increase from 0.08 to 1 nanograms, and they mentioned 10 pregnancies, and 4 were conceived naturally of 6 healthy babies delivered and born in total.
However, since PRP is still an experimental therapy, there is no standard protocol. This is very important to highlight because this fact explains the different outcomes in the various studies, there is a different time of application in those studies, a difference in the concentration of the platelets, in some studies, they inject the PRP more often, so also the technique is different, in some studies they use laparoscopy as well to apply it better, at least this is what they claim.
At IVF-Life clinic, another study is being performed, and the duration is set for 6 years. It was started in April 2021, we don’t have anything to announce so far, but in this study, we aim to demonstrate the efficiency and the safety of the intraovarian administration of the autologous conditioned serum (AAS) for the ovarian tissue regeneration.
The candidates were women between 20-50 years old and had to meet one of the criteria:
The exclusion criteria are:
The difference between this and other studies is that the administration of autologous conditioned serum (AAS) that we obtain from the platelet-rich plasma gives us only the growth factors without other cell particles. We will be able to achieve a high concentration inside of the ovary. The study will include the clinics where IVF-Life is present, so in all of the clinics. We can do a multicentric study. Then the objectives will be to improve the reserve markers and their quality to restore the menstruation cycle and the natural conception and improve the outcomes of a cycle.
The application is done at the beginning of a menstruation cycle in patients with regular cycles. First, we do priming, for example, the DHEA and CoQ10 for 6 to 8 months, then we stimulate, during the stimulation, during the egg retrieval, we apply the PRP. If there are eggs there, we recover them, fertilize and let them develop. If we can do a transfer, we do a transfer, if not, we wait for the menstruation, control the next cycles, see if we have a suitable cycle for stimulation, we don’t wait for the PRP to kick in because PRP needs at least 4 to 6 weeks at least, but it can also be after 6 months that we see a result.
We wait and look for each cycle to see if we have a good cycle for stimulation but to see an effect we need 3 months, and then there could be, for example, a second stimulation. After 20 to 40 days after the application, we measure the hormones, do a scan to see the morphology of the ovary and decide the further steps. Those could be with a positive result, we don’t wait, we start with the stimulation, and if we don’t have a positive result, we control it month per month for the next six months.
It’s very important to say that the method still remains experimental. This concerns more the patients over 40 and who come to the clinic with the desire to become pregnant. These patients usually have not only reduced or low reserve, but the quality of those oocytes is also low.
Other parameters in advanced age influence the success of a potential pregnancy, for example, the immune system. Therefore, for those couples with a very long journey behind them, with several cycles who are clinging to any potential hope of success with their own eggs, for all those reasons, it’s particularly important to empathize that with or without PRP, we still have a very low probability of success. This must be clear to the couple, we must communicate very clearly that the success of the reports presents in only a very small number of patients.
I’m eagerly awaiting the evaluation of the results and hopefully they will allow us to draw 100% right conclusions regarding the importance of PRP and also to refine this technique even more with time. I’m pretty sure that this is the next trend.
It depends on the case, I rarely applied it without stimulation because I want to have big ovaries so that I’m able to apply it properly and also have better results. However, it depends, for example, if a woman is in menopause, we can do it whenever, if a woman has cycles, then I would stimulate a bit.
There is no protocol about it, we measure it only to see where we are and where we get, so it’s not like an existing protocol about how high the FSH must be or how high the AMH should be.
We talk about a low reserve, so every cycle counts. I don’t care if PRP, in the end, will start working, it could be the case that it doesn’t work. I don’t put all my money on the hope in the PRP. If I see a cycle, I monitor the cycles, I see the next cycle, and if follicles are there, I start to stimulate. With PRP, we see more often that there may be 1-2 follicles. It’s not like we’re going to suddenly get 20 follicles, or the ovary will become 20 years younger. We can achieve a few more follicles in the long run, but time is the most important thing here, and we don’t want to lose time and lose the potential chance if it’s there.
You can use it more times, this is also what the study showed, it’s an individual decision, I personally would look into the case, I talk to my patients, and we decide if it would make sense or not to apply it again. I start first once and see how the response is, and then check the results. My goal is not to collect eggs, my goal is to have success and have a baby as soon as possible.
Yes, you can do that. There was one study where a woman in menopause had PRP done, and she then conceived naturally. I have seen that the cycle was normal for some months after the application, so she had menstruation. In some women with perimenopause symptoms like flashing, those symptoms were gone. Therefore, it’s another potential use of the PRP, and it could also be to treat those kinds of symptoms.
Not really, because PRP is gained from your own blood, so it’s like reacting allergic to your own growth factors, and I have never seen it or heard it in any studies. If I have difficulties performing it, if I have a woman that suffers from very high obesity, then it’s quite difficult to apply it and also if I see that the ovary is very difficult to access, then we must think about how we’re going to apply it, perhaps we can do it via laparoscopy. If the woman has like 100 core diseases and then maybe it’s not the best candidate for PRP, those are individual decisions, and we need to see each case and if the patient is suited for therapy.
It depends on is the answer, it could be very fast after 4 weeks to see more follicles, but it could be also the case after 6-7 months, and this is what I see more often. I see a few follicles more after 6-7 months. PRP starts working, or we’re going to see the results, normally if we know the physiology of how the ovary and the recruitment of the follicles work, after 3 months. It’s because the eggs that you have now in this cycle were recruited 3 months before. If I apply it now, then in 3 months, technically, I will see the impact of the PRP then.
The only study that is quite reliable regarding the human growth hormone described the application, and this is how I do it during IVF treatment is every second day 0.1 millilitre 5.8 or 3 milligrams.
I test the immune system, which is a very important player for success, in delivering a healthy embryo. We know that the immune response during our life changes, especially regarding pregnancy after the age of 40, the response is different from some years ago. Many of my colleagues claim that the reason for a miscarriage is that the embryo is not okay, but it’s not true, we have seen that PGT-A is not the solution for all of our problems because other problems are in place. The alteration of the immune system must be treated individually with the proper guidance of the immunologists.
It makes sense, but not to all the patients. We look at how the various white blood cells that physiologically live inside your lining and, in some cases, in your blood also behave, how the populations are if some population of white blood cells is over-represented, and then we see if you’re a candidate for IVIG or other medication. They’re various things, but it is all customized, we will not do it to every patient.
It depends, the nature doesn’t work 0 or 100%. There is a grey area, it’s not like one woman produces only aneuploid embryos, it’s also a big trap of PGT-A testing. If the question has more to do with a low fertilization rate or a low maturation rate, it could be yes, we can have a benefit out of PRP. If you want, share your case with me, and I will give you an individualized opinion about if PRP would make sense. A low blastocyst rate is also a matter of debate, a woman with a low egg reserve doesn’t benefit from a blastocyst transfer.
DHEA is helpful, there are plenty of studies, I give 75 milligrams, so one pill contains 25 milligrams, and I give 3. I give it for like 6 to 8 weeks, after that, I start with the stimulation in patients with a low ovarian reserve together with DHEA, I give also CoQ10, not the expensive stuff, you don’t need expensive stuff. After 6 to 8 weeks, we start the simulation.
About the NADH, there is no study, but if you want to take it, it’s inside many compounds, you can take those, it will do any harm. About the upper age limit for the PRP application, I would not recommend it at 50, to be honest, I would rather recommend thinking about egg donation, which would be a better option.
The woman comes before the application of the PRP, we drain the blood, we do all the process, the centrifugation, everything that I showed you and then we apply it inside the ovaries. It’s from the blood of each patient, it is not like you’re going to have something from Walmart, it’s from each patient with their own blood growth factors.
It can be done. However, we need a rather high amount of blood to drain the PRP, I tend to do it one at a time. However, yes, technically, we can combine it.
I would not give it that way because you don’t achieve what you want, you can’t achieve the higher concentration in your ovaries. PRP via intravenous infusions will not be helpful.
There is no data regarding this, but it would be like a blood transfusion in a way because you would take blood from a different person. Therefore, you would carry some additional risks. I would not expect a better result because you still have the growth factors, and there are the platelets, so you don’t have a lack of quality with age and your platelets, but there is no study about it, and I would not do it. The PRP’s an experimental treatment, and that would be even more experimental.
The idea is quite the same, so the growth factors inside will have an impact on the lining, and the endometrium can be thicker. We also have a protocol if we have like therapy hormone-resistant endometrium to apply PRP, and in some cases, we see an improvement, yes, and we do this.
It’s more the quantity. The influence on the quality is much lower in comparison to quantity. Therefore, it’s more of a matter of quantity.
Some people criticize the PRP, and I’m also not 100% convinced, and they say that by the poking inside your ovaries, maybe you trigger some process inside the ovaries, and the ovaries start to work more. This is what all those studies must clarify. I would never take a woman to poke in the ovaries like this without applying anything. Another field of medicine has shown some importance, and it is to apply it in the ovary and to have better results, so yes, but I’m not 100% sure if the poking of the ovaries is possibly the main hero here in the end.
It’s a complicated topic, and as I said before nature doesn’t work 0 or 100%. This is a problem with the whole PGT-A testing, many women are traumatized that they have done a cycle and have 5 embryos that were tested, and all of them are aneuploid. The conclusion is that they can only produce chromosomally abnormal embryos, but it’s not the case. For example, if a karyotype was performed before, and we had a look at the chromosomes, and everything was okay, there is one reason more that a woman doesn’t produce only unhealthy embryos.
Coming back to your question, the eggs could have an abnormal amount of chromosomes, the PRP cannot have a direct impact on those eggs. However, there is a chance that some more eggs will appear, and in those eggs, the chromosomes will be okay.
I don’t exactly know the weight, we do a scan and check if the ovaries are accessible and then decide if it’s possible or not.
I think so yes, Asherman’s syndrome is a very difficult diagnosis because it’s therapy-resistant. In some cases, there is a very low amount of functional endometrial tissue, so PRP is then one of the main assets that we can use to try to improve the endometrium lining, but in many cases, we’re not able to do it. A stem cells therapy would also be a possible approach.
It’s an option, you also can try to conceive naturally, so whatever you want. In some studies, some natural conceptions were mentioned, so technically, you could conceive naturally.
There is no reliable data on how PRP can increase the chances, but we applied it inside the cavity, we applied it in the endometrium, especially in cases where we have implantation failures, so it’s something that we can consider doing. For example, if we have a lining of 5 millimetres, I don’t tend to give all the medication or hormones, estrogens in the highest doses in the world, which increase the risks of having thrombosis. I try to apply the PRP to see if we can achieve a better lining, so it has also its place.
I rather inject it inside the cavity, we can also do flushes, but I’m not a fan of it, to be honest. I rather apply it with a small needle in the hysteroscopy, I search for the areas where the lining is thin, and then I apply it, so it’s controlled and very organized, not only flashes.
No, I don’t think so, because here you deliver growth factors, and with the mitochondrial injections, it’s also genetic material. It’s like DNA, so it’s something different, both of them are injections, but it’s different.