Breast enhancement by means of an adipose and stromal cell suspension: Adipofilling®

Q) In the treatment of skin scars caused by acne, is it always necessary to create a suspension of small adipose fragments?
A) No. If there are no subcutaneous tissue deficits, only the cellular suspension is made; this will be injected into the dermis. It is advisable to draw off an abundant amount of fat, as the suspensions created by the Adipopimer can be conserved in 2.5 mL or 1 mL Luer lock syringes in a normal laboratory freezer.

Q) Is supplementation with glucosate solution and insulin always used?
A) Yes. This procedure has become part of our protocol because we have observed that it promotes rooting and facilitates conservation of the suspensions at -32°C.

Q) We have seen the interesting slides of the Adipofilling suspensions, but we have never seen a slide of ultramicrofat, etc. Why do you think that is?
A) Well, I’m sorry to say that I, too, have never seen a slide of the material injected by those who present these methods at congresses and in publications. It’s easy enough to prepare a slide stained with EE, and it’s the first thing that should be done. But then, I’m not really surprised. The methods of preparation utilized today – liposuction with micro-cannulas with tiny holes and filtration with filters that are ever finer, etc – damage the biological material and reduce to a minimum, or even destroy, all the adipocytes. In short, these techniques are destructive and inefficacious. Moreover, these methods are unable to process biological material in large quantities and rapidly while maintaining the maximum cell vitality, which is exactly what Adipofilling does. The histology of the various nanofats, microfats and so on has never been presented because you wouldn’t see anything in the smear.

Q) Has the addition of insulin to the separation liquid of the lipoaspirate processed with the Adipopimer ever elicited phenomena of hypoglycemia?
A) We have only had one case of mild hypoglycemia; that was during Adipofilling of the breasts, when we added 80 IU of insulin to the separation liquid. The symptoms immediately resolved with the administration of two sachets of sugar.

Q) Why do you use mepivacaine and not lidocaine?
A) Lidocaine has a marked toxic effect on pre-adipocytes, while mepivacaine only has modest toxic effects. This explains why its use is advantageous and why prolonged washing of the lipoaspirate is important.

Q) We sometimes hear about “closed systems” of processing fat. What do you think of them?
A) In Adipofilling, the biological material is always in a liquid medium (Ringer or physiological solution); it is never exposed to the air. Moreover, the adipocytes secrete cathelicidin, a potent antimicrobial. We have performed hundreds of Adipofilling procedures, and we have never had any infections. The biological material, especially of small dimensions, is not subject to infection. In short, closed systems are of no use.

Q) What other treatments can be associated to intradermal Adipofilling in the treatment of atrophic scars due to acne?
A) Several: micro-excision of the deepest scars; peeling with acids and mixed peeling with resorcin (Electroporo-Cosmesis); needling; smoothing with high-frequency currents; dermo-abrasion. Naturally, potent biological methods like Adipofilling are the first choice; let’s remember that each Adipofilling session is followed by a visible improvement. Every day, at home, the patient can also apply bionic serums of melatonin and ultra-micronized andrographolide (Korpocare) and mild peeling (glycolic acid, mandelic acid, etc.).

Q) What is the difference between using timedsurgical mixed peeling and intradermal Adipofilling to eliminate lip wrinkles?
A) Timedsurgical mixed peeling is a standardized physico-chemical technique that involves timedsurgical de-epithelialization and the application of a saturated solution of resorcin for about 2 minutes. The resorcin acts on all the geometric points of the de-epithelialized surface, and is extremely efficacious. Subsequently, a crust forms, which remains in place for 1 to 2 weeks. When the crust drops off, the treated area remains slightly reddened for a few months.
By contrast, intradermal Adipofilling is a very simple biological technique. The living adipose and stromal cells that are injected into the dermis exert a potent regenerative action without increasing the volume. If injected beneath the skin of the lip, the suspension elicits a controlled increase in the thickness of the lip, thereby correcting the atrophic appearance typical of the elderly person’s lip. The results of intradermal cellular Adipofilling are already visible after three weeks.
When conserved in 1 mL syringes, the Adipofilling suspension can later be used to correct any areas where the result is incomplete. In conclusion, Adipofilling yields much more rapid results. Timedsurgical mixed peeling is more efficacious but more invasive, nor does it have the volume-enhancing effect that the Adipofilling suspension elicits when injected beneath the dermis.

Q) How is it that the Adipofilling suspension can be conserved at the low temperatures of a normal freezer?
The whole world is looking for a cryo-preserver of fat…

A) The Adipopimer, the economical disposable device (Korpo SRL), causes resonance of the biological material, which acquires the property of not freezing; for this reason, it can be conserved at low temperatures for long periods.

Q) How long can the Adipofilling suspension be conserved at -31°C?
A) We have never exceeded one year, but we think it is possible to do so.

Q) Why do your massage the area where liposuction has been carried out?
A) To redistribute the small fragments of fat that remain the subcutaneous tissue and to render the surface of the area uniform.

Q) Why do you advise using mepivacaine rather than lidocaine, which is cheaper?
A) Lidocaine is toxic to the adipocytes and limits the penetration of glucose into the adipose cells.

Q) How do you wash the lipoaspirate?
A) The lipoaspirate is washed with one or two bottles of lactate Ringer or physiological solution in a flask equipped with a tap, until the washing liquid becomes clear. Washing in the flask removes the local anesthetic and epinephrine from the lipoaspirate.

Q) Is the solution of glucose and insulin that you use for “feeding” always the same, or does it vary according to the amount of Adipofilling suspension that is to be injected or conserved at low temperature?
A) For quantities of lipoaspirate up to 100 ml, we use 10 ml of a 5% glucose solution with 30 IU of rapid insulin. For larger quantities, we increase the amount of insulin up to about 60 IU. This solution of 5% glucose and rapid insulin is added to the fragmentation liquid. The insulin exerts its action and is partly eliminated by means of sedimentation or centrifugation at 400 RPM for 3 or 4 minutes.

Q) Have hypoglycemic effects ever been observed?
A) No, but when we use the larger amounts of insulin, we keep some sugar or glucose solution ready.

Q) Can the Adipofilling suspension that is not injected be conserved at a low temperature?
A) Yes, it came as a pleasant surprise to us, too, when we realized that the procedure carried out by the Adipopimer prevented the volumetric and cellular suspensions from freezing, even at a temperature of -31°C!
In practice, the metabolism of the biological material is simply slowed down. The suspensions are conserved in 1 mL syringes that are immediately ready for use. What we have created is an autologous filler that is economical, light and living!

Q) Is it advisable to perform phlebotherapy and Adipofilling at the same time?
A) Yes, the Adipofilling suspension exerts a powerful anti-inflammatory action and restores the integrity of the tissues extremely rapidly. Moreover, two esthetic problems are solved at the same time: excessively dilated veins and the loss of subcutaneous tissue in the back of the hand. In addition, we should not forget the trophic action of the fat on the skin.

Q) So the veins are not obliterated?
A) As is well known, sodium salicylate at a concentration of 12% has a sclerosing effect on the veins of the lower limbs. The veins of the upper limbs, however, are more resistant to the chemical action of sclerosing agents. Our aim is not to close the veins of the hands, but only to reduce their caliber and to strengthen their walls, so that they will not dilate excessively in a hot environment. Our first choice is to use a 6% solution of sodium salicylate in a buffered hydroglycerin vehicle. In rare cases of persistence of venous ectasias, we may use an 8% solution.

Q) Besides the hands, which other difficult areas of aging are corrected in order to complete rejuvenation?
A) Actually, we are very close to achieving overall rejuvenation. The procedures of elastic lifting of the face and neck without dissection help to reposition excess volumes and skin. Adipofilling enables us to increase deficient volumes, especially in the posterior region of the face. Adipofilling also rejuvenates the forehead by making it rounded. Intradermal Adipofilling allows us to treat lip wrinkles and deteriorated skin of the cheeks and neck. By injecting the cellular suspension immediately beneath the skin, we can make fine volumetric corrections safely, for example in the palpebral regions. When injected intradermally, the suspension has no volumetric effect, but exerts a very powerful regenerative action.
Adipolysis enables us to eliminate small excess volumes from the cheeks.
Rejuvenating the appearance of the eyes is also very important. Today, the elastic thread can be used to elongate the eye without acting directly on the canthus, thereby avoiding risky operations with uncertain outcomes. The elastic thread can also shorten a nose-tip that has lengthened with age, thereby yielding both functional and esthetic benefits.
Finally, our bionic serums containing ionized melatonin and andrographolide enhance the beauty of the skin.

Q) Can the two main suspensions, which have different granulometry, be obtained by activating the Adipopimer for a set number of seconds?
A) To obtain the volumetric suspension, the Adipopimer is activated for 4 seconds; for the regenerative suspension, it is activated for 15 seconds. However, the granulometry obtained must always be checked by tilting the beaker, because the consistency and volume of the adipose lobules may vary from one patient to another. In addition, quantities of more than 100 mL of lipoaspirate may require a longer duration of fragmentation.

Q) Are there any limits to the rejuvenating effect on the skin and to the elimination of wrinkles?
A) This technique is new and its limits are not yet clear. What is clear, though, is that the living adipose and stromal cells injected into the dermis exert the most powerful regenerative action known to date.

Q) What advantages does Adipofilling have over the techniques of nanofat, microfat, etc?
A) Lipoaspiration is quicker; fragmentation of the biological material is atraumatic and rapid; the granulometry (tiny fragments of adipose tissue or a suspension of single cells) can be selected; the vitality of the adipocytes and stromal cells is maintained; and large quantities of lipoaspirate can be processed in a few seconds.
Moreover, it should be borne in mind that small-caliber cannulas damage the adipocytes; indeed, everything (except water) that touches the adipocytes will damage them.

Q) So, doesn’t that mean that the Adipopimer damages the adipocytes?
A) No. The adipocytes and stromal cells are separated by means of the aspiration vortex created by the Adipopimer. A volume of one third of lactate Ringer solution must be added to the washed lipoaspirate; this enables the vortex to separate the adipose tissue first into fragments, and then, after more prolonged fragmentation, into single cells.
Following centrifugation at 400 RPM (17RCF G), the cell suspension is topped by an extremely thin layer of supernatant oil, which demonstrates that the adipocytes remain intact.
Indeed, a smear of the cell suspension will show intact adipocytes. I advise anyone who performs micro-fat and nano-fat techniques to carry out this extremely simple examination. I believe it’s useful to examine under the microscope what we inject. I think Tonnard did this with nanofat obtained from multiple filtration. Nanofat does not contain living adipocytes. Personally, I would never inject it. Injecting a pulp that does not contain adipocytes will not have any significant or lasting effect. Living adipocytes are essential, as they produced numerous substances that favor tissue regeneration.

Q) What can you say about the use of an open system; is there any risk of infection?
A) We’ve always wondered why, in the hundreds of Adipofilling procedures that we have performed in the Department of Plastic and Reconstructive Surgery and the Burns Unit of San Martino regional hospital in Genoa, we have never had a single case of infection. We now know the answer; the adipocytes secrete potent antimicrobial substances, including the peptide cathelicidin.

Q) We’ve seen the extraordinary result of intradermal Adipofilling. After only 20 days, the wrinkles had disappeared and a large area of elastosis and scarring had improved. It looks like a different neck…
A) Intradermal Adipofilling with living cells is extremely efficacious. We now propose to study the changes that occur in skin that is rejuvenated by means of this technique. The adipose and stromal cells injected into the dermis do not exert a volumetric effect; their effect is regenerative. In order to eliminate wrinkles, it is necessary to inject a quantity of cell suspension that extends well beyond the wrinkle itself.
Regenerative Adipofilling is a very important means of rejuvenating our patients’ skin, and opens up a new era. Anyone who understands this can achieve previously unattainable results.

Q) Adipofilling is known to eliminate orthopedic pains…
A) Yes. There’s a whole world yet to discover.

Q) What is the granulometry of the suspension used in Adipofilling of the nose?
A) Small clumps of cells.

Q) How can you see the granulometry of the cellular suspension?
A) After fragmentation, you only need to incline the beaker and see what remains on the glass. Small cell clumps can be clearly seen. If the granulometry needs to be reduced, the suspension can be fragmented further. In this case, a uniform layer made up mainly of single cells will remain on the surface of the glass when the beaker is inclined.

Q) When are single cells used?
A) They are used for more delicate corrections and for the intradermal injection of wrinkles, areas of roughness of the face, stretch marks, etc. Single cells suspension improves the engraftment of the volumetric suspension.

Q) When you wash and handle the fat, why do you use glassware?
A) Because glass is safe. Remember that fat tends to wear away plastic materials, leading to the release of substances, which I prefer to avoid as far as possible.

Q) How long after esthetic rhinoplasty can Adipofilling of the nose be performed?
A) After one or two years, noses that have undergone rhinoplasty normally become skeletonized, taking on a typical artificial appearance. Adipofilling eliminates this artificial look and corrects the defects that are manifested after this period of time.

Q) Treatment of this case of chronic radiodermatitis and bone exposure unequivocally demonstrates the enormous regenerative power of Adipofilling, which derives both from the dimensions of the cells and from the union of living adipose and stromal cells. Do you think that this effect can be achieved by means of the recent techniques of fat preparation?
A) Unfortunately, the recent techniques of fat preparation – filtration through micro-filters, nanofat, microfat, aspiration through cannulas with micro-perforations, working the lipoaspirate with small metal spheres, etc – tend to destroy the adipocytes, either partly or completely. Naturally, the rubbing and squeezing caused by these irrational techniques also damages the stromal cells. To function correctly, adipose and stromal cells must be living and be used together, in order to exert both their volumetric effects and their regenerative effects. In Adipofilling, the lipoaspirate is fragmented according to the laws of granulometry; this keeps the adipocytes and stromal cells alive. Adipofilling rapidly yields large or small amounts of cell clumps or individual cells that can be injected anywhere, even into the dermis. The above-mentioned recent techniques do not consider the preservation and the function of the biological material. But this doesn’t surprise me anymore. We need only think, for example, of what has happened in phlebology; allegedly “validated” techniques, such as sclerotherapy and phlebectomy, are implemented worldwide – and yet they are destructive. Indeed, they treat a three-dimensional pathology in a two-dimensional manner. In other words, they treat the effect (which is also the “escape valve” for hemodynamic hypertension) of this pathology and not its cause; in short, they ignore the physiopathology of the most common disease on earth!

Q) Once the lipoaspirate has been washed and fragmented, how is it enriched with stem cells?
A) The portion of the lipoaspirated material that will not be injected is fragmented for a longer time. This yields a suspension of single cells. Centrifugation is carried out at a higher speed (500 RPM) and for a longer time (5 minutes). After centrifugation, the stromal cells make up the lowest layer of the material centrifuged; this layer is white in color, and is situated immediately above the water. The stromal cells are then added to the fragmented lipoaspirate that is to be injected, and remixed by means of two syringes and a connecting tube.

Q) In this case of radiation ulcer, was the fragmented lipoaspirate enriched?
A) No.

Q) This patient seems to be very thin. Was it difficult to obtain a sufficient quantity of fat to perform Adipofilling®?
A) Even if the patient is very thin, it is always possible to aspirate 100 mL of fat from one or both trochanters; this will become 60 mL of cell suspension, which is sufficient for these facial corrections.

Q) Which regions is it best to take the fat from?
A) All regions are suitable, but in some the fat needs to be evaluated; for example, the knees are not always suitable because the adipose tissue may be too fibrous.

Q) Does hardening ever occur?
A) After injecting the suspension, we normally massage the area gently to distribute the cells. Some areas may display hardening for a few hours, but the cells nevertheless find their proper location and the tissues become soft again.

Q) Is anesthesia of the receiving area carried out?
A) Only at the entry points of the needle or micro-cannula.

Q) How is the micro-cannula inserted?
A) The skin is pierced with an 18 G needle, and the cannula is inserted through this aperture.

Q) How is the Adipofilling® implanted?
A) As Adipofilling® is a cell suspension, it can be uniformly distributed as if it were a “volumetric revitalization” agent. We avoid multiple insertions of the needle or cannula as this causes needless fibrosis, the effects of which nevertheless disappear in a relatively short time.

Q) Which is the longest phase of this procedure?
A) Washing the lipoaspirate. First of all, I advise using mepivacaine as an anesthetic, because lidocaine prevents glucose from penetrating into the adipose cell. Next, I advise using an efficacious amount of epinephrine and waiting for a sufficiently long time before carrying out liposuction. There should preferably be no blood in the lipoaspirate. These small quantities of fat can be washed with lactate Ringer solution in the same syringe used for aspiration.
Fragmentation with the Adipopimer® (Korpo, Italy) takes only a few seconds.

Q) Is the Adipopimer® that we see the market version?
A) Yes, it is the most recent version, which is the best suited to creating the ideal suspension. Correction of this facial atrophy was achieved with the prototypes that enables us to produce today’s market version.

Q) Can the suspension be frozen?
A) Yes, but we prefer to use it fresh.

Q) Is the result permanent?
A) In the face, the cells present after two months are permanent. Naturally, if the patient puts on weight, the grafted cells will also increase in volume; if the patient loses weight, they will decrease in volume. After Adipofilling® the patient must eat properly without going on a diet.

Q) Is rooting always satisfactory?
A) By performing the technique correctly, we have always obtained good results, without complications. The volume enhancement that we seek to achieve is biological; in other words, it involves living cells. We are not interested in achieving increased volume due to surgical artifacts of traumatic fibrosis or to the insertion of adipose lobules containing apoptotic cells.
The first Adipofilling® procedure may seem disappointing, but it plays an important role in preparing for the subsequent cell graft.

Q) Centrifuged Adipofilling offers the extraordinary possibility of obtaining stromal cells in a few minutes. When are these used?
A) The stromal cells in the fragmented material that is not injected are separated from the adipocytes by means of centrifugation at 300 RPM for about 5 minutes. These cells are remixed with the cell suspension that is to be injected. They are chiefly used for regenerative purposes in several fields, for example in the treatment of radio-dermitis or, by means of intradermal injections, to improve stretch marks on the skin.

Q) Do you realize that Adipofilling and Adipopimer will revolutionize the fat transfer?
A) Sure

Q) What is the difference between Adipofilling and lipofilling?
A) It’s a question of size. Adipofilling is cellular, while lipofilling is lobular. A cell suspension can be injected immediately beneath the dermis, and even into the dermis; by contrast, the superficial injection of lobular fat can cause problems. With Adipofilling, the stem cells are distributed uniformly in the tissues, while with lipofilling the stem cells remain within the structure of the adipose lobule. With Adipofilling, the suspension to be injected can easily be enriched with stem cells, which makes for greater efficacy in terms of both volume enhancement and regeneration. With Adipofilling, the adipose material is drawn through a 4 mm or 3 mm cannula; lipofilling uses finer cannulas, which, in addition to slowing down the process of extracting the fat, cause considerable damage to the lipo-aspirate.

Q) If the cell suspension is centrifuged at 500 rpm for 5 minutes, the stromal cells are isolated; what happens if these cells are not remixed with the adipocytes?
A) The stromal cells, which contain the stem cells, play an important role in the survival and rooting of the cell graft. Grafting only adipose cells does not yield the same results.

Q) Here, we can see signs of a physical action around the navel…
A) Another operator burned this patient with an apparatus called “plasma scintillator” or “electron-flow scintillator”. My mother, Dr. Spolidoro, used similar equipment over 70 years ago! As I’ve already said, it’s very unlikely that a physical means can equal, let alone improve upon, the biological action of Adipofilling, the results of which increase exponentially as sessions are repeated. The result seen in this video publication was achieved in only one session of Adipofilling!

Q) Can facial atrophy (PRS) also be treated with Adipofilling?
A) Yes, and the results are very good.

Q) What other pathologies have been treated with Adipofilling?
A) We have treated cases of radio-dermatitis, even with exposure of the bone; in elderly patients, we have prepared the scalp for the trouble-free creation of the skin flaps needed for reconstruction following destruction by cancer; we have injected Adipofilling immediately beneath the scars left by extensive burns; we have used it in the rectum, labia majora and vagina, in the lips of patients with scleroderma, in the sequelae of poliomyelitis, in scoliosis, etc.

Q) What are the most common aesthetic applications?
A) Volumetric enhancement of the lips and of the face in general, including the forehead; in the hands; in the breasts, sometimes after breast coning by means of the elastic thread (Elasticum® Korpo, Genova, Italy); in all forms of subcutaneous and muscular tissue loss; beneath stretch marks, owing to its potent biological action; in the nasal pyramid, where it eliminates the appearance of the “remodeled nose”; on the face and body in general, including the scalp.

Q) Is Adipofilling performed by means of a disposable instrument?
A) The instrument is extremely economical. Moreover, using a disposable instrument ensures proper preparation of the lipoaspirate, sterility and the absence of cross-contamination.

Q) Does performing Lipostructure still make sense?
A) To be honest, in my opinion, it has never made sense.

Q) In Adipofilling®, cell survival is better than in other methods of transferring adipose tissue. Is this due to the fact that the filler is a suspension of cells?
A) Yes, individual cells survive better than lobules and are much more easily nourished. Fragmenting the adipose lobules causes far less damage to the cells than centrifugation at 3000 rpm, which is sheer madness.

Q) Why?
A) First of all, it is not clear what is meant by 3000 rpm. We should use the term “atmospheres”. In any case, we can suppose that 3000 rpm corresponds to about 1500 atm. This treatment causes a portion of the adipose cells to burst (a lot of oil is seen in the supernatant). Moreover, on electronic microscopy, the apparently whole adipocytes display numerous micro-vacuoles, which is by no means a good sign. By contrast, after mechanical fragmentation, a minimal amount of oily supernatant is seen and the size of the adipocytes (100,000 nm) safeguards them against damage. Anyone who is familiar with granulometry is well aware of this.

Q) Is Adipofilling® of the breast an ambulatory procedure? How long does it take?
A) Yes. Adipofilling® is an ambulatory procedure. It only takes a few seconds to transform the lobular fat into a cellular suspension with the Adipopimer®; washing the lipoaspirate takes longer.

Q) Why does the lipoaspirate need to be washed?
A) Washing removes any anaesthetic, epinephrine and blood that may be present. To reduce the washing time, an anaesthetic solution of 1 mg of epinephrine in 500 ml of lactate Ringer solution can be prepared; the operator then waits 10 minutes after infiltration. This limits the amount of blood in the lipoaspirate, thereby reducing the washing time; indeed, the presence of blood in the lipoaspirate necessitates more prolonged washing.

Q) To make up the anaesthetic solution is it necessary to use mepivacaine?
A) Studies conducted by diabetologists have shown that lidocaine prevents glucose from entering the adipose cells. However, after thorough washing, we have not observed substantial clinical differences between mepivacaine and lidocaine, however we use mepivacaine.

Q) How much can breast volume be increased by?
A) A considerable increase in breast volume can be achieved, though this depends on the amount of adipose tissue that the patient has and, naturally, on the number of operations performed. If the patient has little superfluous adipose tissue, the volumetric increase will be modest. However, the available adipose tissue can be used to correct any asymmetry and to improve the shape of the breast, for example by widening the base or filling areas where the volume is insufficient.

Q) In Adipofilling®, can the cellular suspension be enriched with stem cells?
A) Yes, quite easily. Stem cells can be isolated by centrifuging a portion of the cellular suspension for five minutes at the lowest speed of the centrifuge. A thin white layer of stromal cells will be seen to form at the distal end of the syringe; these cells contain the stem cells.

Q) Once the stem cells have been isolated, what has to be done?
A) The layer that forms at the end of the syringe has to be mixed with the non-centrifuged cellular suspension; this is done by means of a connecting tube. The stromal cells obtained by means of minimum-velocity centrifugation are used, together with the adipocytes, for small corrections, for example corrections of the nose.

Q) Are stem cells often added?
A) We have never used them in breast procedures. We do not think it is advantageous to stimulate the breast tissue excessively.

Q) How is the washing of the lobular fat lipoaspirate carried out?
A) An erlenmeyer flask equipped with a tap is used and the lobules are washed until the Ringer solution becomes colourless.

Q) What are the “tricks of the trade” in this operation?
A) The fat must be drawn off through a large-calibre cannula at a low aspiration pressure; it must then be thoroughly washed, carefully fragmented with the Adipopimer® and uniformly spread in the subcutaneous tissue. The Adipopimer® is used only once; in this way, ideal fragmentation is always obtained.

Q) Why does the fat have to be drawn off through a large-calibre cannula?
A) A large-calibre cannula causes less damage to the adipose cells than a small-calibre cannula


Korpo thanks Medical Video Journal CRPUB.ORG open access for the material placed at our disposal.