Fat transplant: Amazing growth and regeneration of cells and rebirth with the miracle of fat cells

During fat transplantation, adipose tissue is removed from the body and injected into different areas under the skin. The goal of this review article is to look into the efficacy and applicability of fat transplantation in regenerative medicine and rejuvenation, including Nanofat, Microfat, and Millifat.


| INTRODUC TI ON
The effect of regenerating adipose tissue and its intrinsic cells is an evident process. 1 Although fat transfer and transplantation have been established for nearly 30 years, facial fat transplantation has remained pretty consistent in popularity. 2 Fat transplantation has been offered as a viable method for the reconstructive treatment of facial scars, in addition to the beauty of the face and body. 1,2e regenerative function of adipose tissue is related to various factors including multipotent mesenchymal stem/stromal cells (MSCs) or adipose-derived stem cells (ADSCs), enzymes, cytokines, growth factors, and the cellular and molecular mechanisms of these compounds. 1,2cial scars are frequently described as aberrant and can hurt a person's life.Since it is a difficult repair process, using autologous adipose tissue for regenerative repairs of facial scars can be a valuable treatment.The transplanted cells and their secretions may be responsible for this regeneration impact. 1,3The principles of skin fat transplantation and micro lipo-injector techniques with advanced techniques such as Microfat and Nanofat have shown promising results. 2,4,5

| OBJEC TIVE AND ME THOD
The object of this review article is to investigate the effectiveness and application of fat transplantation, including Nanofat, Microfat, and Millifat in regenerative medicine and rejuvenation.To collect data for this purpose, a comprehensive literature search of the National Library of Medicine (PubMed) and Medline databases was conducted for animal/in vitro and human studies published until 2023 using different versions of the following keywords: adipose tissue, fat grafting, nanofat, microfat, millifat, rejuvenation, and regenerative medicine.Relevant articles were independently assessed by two separate reviewers and selected through the assessment of titles and abstracts as well as the reference lists of related articles.Pertinent articles were selected and reviewed.Data on the preparation, isolation, and injection of adipose tissue, clinical applications, and results of injections were collected and evaluated.In this review, we have tried to discuss current opinions about the results of studies investigating the merits of fat transplantation in regenerative medicine.In addition, we designed tables and figures from the general results of these techniques, the method of adipose tissue separation, the preparation of millifat, microfat, and nanofat, and the obtained data.As a result, we can deliver more specific information to the reader.

| AD IP OS E TISSUE AND G R AF TING TECHNI Q U E S
Adipose tissue has received a lot of attention due to its simplicity of separation by liposuction, autologous use, and biocompatibility.
Autologous fat grafts are employed in a variety of applications, including regenerative medicine, scar repair, facial rejuvenation, and volume restoration. 6,7In recent years, researchers and doctors have employed various protocols, manipulations, and substances such as platelet-rich plasma (PRP) to improve performance while decreasing immunological and inflammatory responses. 7,8Microvascular fragments, stromal vascular fraction (SVF), stem cells, microfat, and nanofat can be obtained by preparing and manipulating fat tissue. 4,7ese products are used in clinical and research settings to boost angiogenesis and tissue repair.Macrofat, microfat, and nanofat are utilized in plastic surgery for cosmetic and regenerative purposes.
Because of its high concentration of growth factors and stem cells, Nanofat is employed as a remodeling susbstance, whereas Microfat and Macrofat are typically used as filler material. 4,7e two main components of fat tissue are mature adipose cells and SVF. 9 The patient can be treated in vitro with SVF and cultivated ADSCs.10 According to recent studies, adipose tissue contains the greatest number of adult stem cells in the body.3 The use of nanofat is one of the sources that may easily and cheaply separate SVF and ASCs, expanding the therapeutic range and the usage of fat tissue transplantation.7,11,12 SVF and matrix are composed of various cells, including ADSCs, vascular smooth muscle cells, MSCs, pericytes, fibroblasts, endothelial cells, hematopoietic cells, and other immune cells.8][19][20][21] There are two general ways to obtain SVF: the first method is a chemical protocol that involves enzymatic digestion, and the second method is a physical protocol that involves mechanical separation from connective tissue.22 Collagenase is commonly used for enzymatic digestion, and cells are separated by filtration or centrifugation.22,23 The most important goal in all these stages is to keep the stem cells and stromal cell niche alive, which has numerous therapeutic effects for SVF. 22The precise mechanism of SVF's therapeutic actions is unknown; it could be through numerous pathways or be specific to tissue conditions.22,24 Adipose tissue contains ASCs, which play an important role in tissue regeneration and rejuvenation.ASCs secrete a variety of factors, including vascular endothelial growth factor (VEGF), Transforming growth factor (TGF)-β, hepatocyte growth factor (HGF), fibroblast growth factors (FGF), platelet-derived growth factor (PDGF), placental growth factor (PlGF), stromal cell-derived factor 1 (SDF-1), interleukins (IL)-8, and leptin which cause angiogenesis, increase endothelial cell proliferation, decrease endothelial cell apoptosis, proliferation and migration of dermal fibroblasts and epidermal keratinocytes, increase collagen and fibronectin production, which are ECM proteins, and regulate collagen synthesis.16 Furthermore, ASCs can boost the release of anti-fibrotic factors like TGF-β3 while inhibiting profibrotic factors like TGF-β1 and IL-6.These properties have enhanced the usage of nanofat in regenerative and anti-aging applications.16 According to a recent meta-analysis, 25 the efficacy of cell-assisted lipo-transfer (CAL) was investigated, and fat survival was significantly higher in 16 investigations. CL produces higher adverse effects than standard techniques, regardless of injection site or injection amount (less than 100 mL).3,25 Cell survival may be impacted by the location of the adipose tissue in the donor body, the method of removal, the cannula, the pressure used, or the capability of injecting the complete solution while under anesthesia, 3 and the transplant volume is determined by the injection site prior to tissue collection.26

| Preoperative preparation
Before undergoing fat transplantation, the patient should consult with a specialist and outline his/her expectations.The conditions should be explained to him/her by the specialist.It is also recommended that the patient take antibiotics a day before surgery.Furthermore, fixed-focused ultrasound treatment can be performed a few days to a week before surgery to promote the probable release of endogenous angiogenic growth factors. 27,28The patient can select between general anesthesia and local anesthesia for surgery and fat removal, but in most cases, local anesthesia is sufficient. 27

| Liposuction
Liposuction is a type of surgery in which fat is removed from particular parts of the body using suction.Fat removal is done from any place with excess subcutaneous fat as well as the patient's favorite areas if there is enough fat.As indicated in Figure 1, specialists normally recommend donating from the abdomen, however, hips or thighs may be used too. 1,29matrix metalloproteinases (MMP)-9 levels in abdominal lipoaspirates are higher in studies.However, the amount of SVF and ASC cells did not differ significantly between areas. 1,29e removal operation alone takes 45-60 min.Under sterile conditions, harvesting begins with a 14G needle and a 60 mL syringe.
After inserting a 12-hole cannula with a 2.5 mm diameter opening into a 14G needle hole, 120 cc of fat is removed. 27

| Processing and emulsion
After harvesting and draining the swollen fluid, the fat is washed with a lactate ringer to reduce blood contamination.Although there is still debate on this, systems such as pure graft and centrifuges are not necessary for the preparation of fat for transplantation. 27,30,31 shown in Figure 1, after complete gravitational separation, the fat content is divided into three layers: the oily layer on the upper, fat in the middle (low volume and high numbers of SVF cells), and the watery layer below.The oily and watery layers are separated and removed, leaving only the refined fat. 1,27,30The remaining fat is transferred to another syringe and processed into Microfat and Nanofat. 27On the remaining fat, two filtration stages are done.
The first step is to transfer fat from a full 10 mL syringe to an empty 10 mL syringe using a 2.4 mm female Luer-Lok connector (Milifat).
The same operation is done 30 times more intensely with a 1.2 mm Luer-Lok connector (Microfat). 1,32This is known as emulsification,

F I G U R E 1
Steps to prepare different types of fat for transplantation.(1) Harvesting fat from the abdomen or thighs.At this stage, this adipose tissue can be used to prepare adipose tissue derivatives.To prepare milifat, microfat, or nanofat: (2) After centrifugation, the upper and lower layers are discarded for purification, and the middle layer, which is the fat tissue and cells, is separated to prepare milifat, micro fat, or nano fat.(3) Then the fat obtained passes emulsify through a 2.4 mm connector, 30 times, to produce milifat (4) For micro fat, the fat obtained passes emulsify through a 1.2 mm connector, 30 times (5) If nano fat is needed, the fatty tissue should be passed through the 400-600 μm emulsifier once.To prepare stromal vascular fraction (SVF): (6) The separated fat tissue is washed.( 7) Physical protocol or chemical protocol is used to prepare SVF. ( 8) After centrifugation, the different layers are separated and the SVF pallet is obtained.The nanofat, milifat, microfat, SVF, and ADSCs obtained can be used for injection for rejuvenation and regeneration.and it is necessary for the destruction of all fat cells and the formation of very small cell particles. 1,33[34][35]

| Preparation and injection
Extracted fats to anatomical locations are done according to the size of Millifat (2-2.5 mm), Microfat (1 mm) Nanofat (500 μm and less). 18,27Usually, the injection begins in the deepest area of the injection site, particularly the face, and advances to the surface.As a result, Millifat, Microfat, and Nanofat are used for injection from the bottom layer to the top layer, respectively (shown in Table 1). 27,36r example, for facial injections maximum of 12 puncture sites are made in the face with 18G and this process can be repeated with all three sizes of fat graft. 27e emulsified fat was pressed through a 0.5 mm pore filter and then transferred to 1 mL sterile syringes for injection. 1,2The resulting fluid is injected into the scar or dermis tissue with sharp needles 24G, 25G, or 27G respectively.As shown in Table 2, the needle diameter depends on the quality and firmness of the scar tissue. 2,33,35rigottomy is a treatment in which fibrous bands are tightened and attached to the scar to deeper levels before being mechanically removed and replaced with Microfat.This treatment is used for diminishing shrinking scars.In the rigottomy procedure, a centrifugal solution is injected into the scar site and subsequently injected through an incisional cannula beneath the scar. 1,32

| Millifat
To make millifat, fat is passed between two 20 mL syringes through an emulsifier with a 2.4 mm diameter. 27,33Millifat can be injected into patients, particularly in the suborbicular oculi fat (SOOF), for structural augmentation in the temporal regions, the deep fat compartments of the cheek, the buccal fat pad, medial and prezygomatic, on periosteal in the orbit, the lateral eyebrow region, the pyriform region, the mandible, columella, nasal bridge, the chin, and the lips. 27,33

| Microfat
To make microfat, we can use 3 mL syringes with 0.7 mm 2-inch injection cannulas.A 23-25G cannula is sufficient to generate entrance sites for crossing. 33The top-down approach is utilized using a 0.7 mm Microfat injection cannula and approximately 10-16 mL of Microfat on each side of the face (a total of 20-32 mL of Microfat).
To prevent overfilling, it is injected deep along the periosteum to overfill these areas. 2,27The Microfat is superficially grafted to the skin surrounding the mouth, the needle is positioned on top of the nasal muscle, the upper lip and mouth incisions are employed, and the regular marionette cutting. 34It should be noted that microfat is injected subcutaneously using a 21G cannula.

Application
Removing scars and wounds from surgery or accident 32 Filling old hands with prominent veins 41 Filling the gaps and voids under the skin and deep facial lines 27,34 Treatment of AGA and strengthening and hair growth 55

Durability
Long duration of about 4 to 5 years or more 41 Abbreviations: AGA, Androgenic Alopecia; SVF, stromal vascular fraction.Wrinkles and damage caused by the operation may decrease up to 1 year after the operation.Over time, the graft volume may decrease between 50% and 90%, so the patient needs counseling at specific times after the operation.[40]

| Hand rejuvenation
Due to their greater visibility, hands are crucial for beauty. 412][43] As shown in Figure 2, the distribution of fat on the back of the hand is divided into three layers: superficial, intermediate, and deep. 44,45Fascial planes separate the laminae, and the extensor retinaculum connects the fascial layers where they merge.Tendons run deep into the hand, while veins run superficially to the extensor retinaculum. 41Surface layer joints start with a depth of less than 1 mm and the distribution of fat in this layer is uneven.This layer has 8-12 septal adhesions on the surface that contain small blood vessels. 44,46The middle layer consists of blood vessels and nerves, and the dorsal layer of the dorsal veins is of medium diameter. 47The beauty of the hand is related to the ectomorphic body type, and long/slender hands are the most desirable type. 41e amount of hand tissue varies in degrees.Grade 0 refers to normal and smooth hands, grade 1 to mild atrophy with rhythm, grade 2 to moderate atrophy with exposed veins, and grade 3 to severe atrophy of exposed tendons 47.Nonsurgical treatment can resolve grade 1 volume reduction, however surgery is the best option for grades 2 and 3. 41,42 Principles of fat transfer according to the types of hands with low pressure, low speed, and low volume are done through multiple tunnels and multiple levels. 42,48,49Throughout the fat transfer, emphasis should be paid to evenly distributing small amounts of fat at the desired levels under low pressure. 40,41Incision sites for fat grafting differ depending on the technique. 42Although, in general, incisions in other webspace can allow good access to the deeper layer and the distal area of the back of the hand while minimizing scarring. 41To reduce the number of scars, an incision between the third and fourth metacarpals is recommended by some experts. 41,45ere is a lot of controversy over which laminates are ideal for fat deposition.There is a lot of agreement about the safety and efficacy of the injection only to the surface layer.In this layer, there is no important structure and the deeper vessels and tendons are covered by the fat graft 41,42,45,49 (shown in Figure 2).Due to the presence of dorsal veins in the middle lamina, some experts recommend that preventing intravascular fat deposition is required. 42However, some advocate for fat grafting to maximize the effectiveness of the treatment without serious safety concerns. 45,49In general, some experts recommend deep layer transplantation to correct concavity between metacarpal bones, while deep fat transplantation is suggested in some cases. 41,42,45,49The preferred method for fat delivery is cannulas with sizes between 14G and 17 G.Cannulas bigger than 14G increase the possibility of central necrosis and vascular issues. 40,42,45,49,50perts agreed in their latest recommendations that 15-30 mL of fat per back transplant are moderate, depending on the technique and amount of correction required for the transfer. 40,42,45,49For harvesting and donating, most Specialists employ classic obesity of the flanks, periumbilical, or middle thighs. 51As an example, the inner thigh is recommended as an ideal location due to superior graft resection and less patient discomfort. 41,45t grafting to the hand is usually used as an effective treatment to reduce the volume and subsequent protrusion of blood vessels and tendons due to aging. 40,42,49The most common problem is edema.It has been assumed that edema is a natural aspect of the healing process following a fat transplant and should not be regarded as a real problem. 45,48For immediate management, pressure dressings can be used, which are useful for a few days after the operation.
Other issues that resolve on their own include ecchymosis, which may be caused by vascular damage, and paresthesia, which may be caused by damage to tiny nerve structures. 40,42Furthermore, antibiotic prophylaxis with cephalosporin is generally used to reduce the infection risk. 40t transplantation is one of the hand rejuvenation techniques with fillers such as hyaluronic acid and poly-l-lactic acid to restore tissue volume. 41Synthetic fillers are approved by the US Food and Drug Administration (FDA) for safety and effectiveness.The distinction between these fillers and fat grafts is that they represent no risk or adverse effects to the patient in exchange for the same tissue volume recovery result. 41On the other hand, the treatment time of these fillers is temporary and depends on the products. 41In general rejuvenation techniques may not restore tissue volume well, but they are effective in changing the beauty of the back surface of the hand. 52,53

| Hair growth
Alopecia is a condition caused by the gradual reduction of hair loss due to various abnormal reasons such as the endocrine system, genetic factors, and stress.In androgenetic alopecia (AGA), immune cells produce an inflammatory response against hair follicles that eventually causes thinning hair, loss of hair follicles, and hair loss. 54,55Despite the known treatments to date (approved treatment by the FDA only delays AGA progression), there is no effective modality to inhibit disease progression.[58] Hieronymus Stevens et al. 59 used the combination of SVF and PRP in the treatment of AGA.To prevent pathological damage to the damaged tissue of the host and to maintain proper signaling conditions, the SVF can be combined with PRP. 221][62] The results of this study showed that a single-dose injection of SVF-PRP can increase hair density in AGA.
In another study, Ghazala Butt et al. 55 divided patients into two groups: the PRP group (only PRP was injected) and the SVF-PRP group (adipose-derived SVF mixed with PRP).PRP and SVF-PRP were injected into the affected areas of the scalp in AGA patients. 55,59,63The results of this study confirmed that SVFenriched PRP treatment is more effective than PRP alone.Studies show that SVF-enriched therapies play a significant role in the treatment of alopecia.However, despite the limitations of this study, SVF enrichment may be a promising way to treat AGA in both men and women. 55 this light, Mohammad Ali Nilforoushzadeh et al. 63 suggested that the use of scalp injection of adipose tissue can elicit a positive effect on hair growth in trauma alopecia.The results of studies show the effectiveness of using adipose tissue in the treatment of alopecia and studies can increase the effectiveness of these treatments for all types of alopecia by determining the optimal treatment regimen.

| Scar treatment
Scarring is the pathological reaction of repair and recovery following tissue trauma, and it can have an impact on a person's appearance. 64thods for treating scars include intraregional injection of corticosteroids, 5-fluorouracil, bleomycin, silicone sheets, laser, and surgery.However, these methods have different effects and can cause discoloration, high recurrence rates, skin atrophy, and pain.As first described by Neuber in 1893 and later modified by Coleman, autologous transplant is another modality to treat scarring. 65According to this method, the presence of MSCs can improve scar tissue quality.
Because of its changing properties, this approach can be used for more than just filling the volume. 65tologous ingredient injection with clinical and pathological improvement is a treatment technique for treating the characteristics of the scar and its symptoms and assisting the scar to return. 66r example, in mammoplasty, which is often used to decrease the breast, a long-lasting but low-resolution scar is generated, which is one of the main causes of dissatisfaction with this surgery.Previous research has shown that treating immature scars with a combination of macromolecules can successfully reduce the growth of hypertrophic scars. 67Furthermore, autologous primary transplantation can be employed in any part of the face to treat adult scars and fibroids in terms of skin texture, color, volume, softness, and pattern quality. 65crofat and nanofat can be used to treat atrophic acne scars. 68,69an Yu et al. 70 discovered that fetuin-A is secreted from fibroblast cells and can suppress the fibrotic characteristics of hypertrophic scar fibroblasts.It was also discovered that the etv4 transcription factor can be an essential factor in the anti-fibrotic effect of micrograft in hypertrophic scars.Nanofat grafting has been proven in histological studies of wound tissue to improve rejuvenation in fine-line scars by speeding and enhancing collagen synthesis. 71 general, based on the results of various studies, fat transplantation is the treatment of choice for most scars and acts as an independent or adjunctive method.

| COMPLIC ATIONS AND CHALLENG E S
Fat transplantation has disadvantages as well.Aesthetic irregularity or asymmetry is a common issue with fat injection.Another typical issue is restricted fat masses caused by fat accumulation in the superficial layers or necrosis.However, Lipodissolve, direct open excision, and suction lipectomy can be used to treat these patients. 34,39ong other existing problems, we can mention fat embolism, nerve injury, hematoma, and infection.Some of these risks, such as infections, are extremely unlikely.The use of preventive treatments such as the injection of 1% lidocaine with 1:200000 epinephrine to prevent embolic events is suggested.The lip injections as the last injection site are recommended to prevent contamination with normal flora, the use of antibiotics to reduce the possibility of infection, and the use of acyclovir for patients with cold sores to reduce these problems also are recommended. 39,72Other problems with these treatments include the non-permanence of the effects of some of these treatments, the existence of less dangerous and simpler alternatives, different treatment results, and the existence of different treatment protocols and preparation methods. 41,42,45,48his characteristic allows them to be grafted to multiple locations and exploit their properties.Some properties of fat tissue boost its efficiency, such as an increase in stem cells, growth factors, cytokines, and substances that aid in repair, regeneration, and rejuvenation.In this article, different methods of rejuvenation and clinical uses of adipose tissue were investigated.This approach may be more effective when paired with a greater number of stem cells, such as SVF or PRP.

4. 4 . 3 | 37 4. 5 |
NanofatDepending on the site, several approaches are used to inject nano fat.A 23G needle is used to insert the microcannula, which is the smallest feasible cannula.Injections with 25G needles are easier because of their hardness, which allows them to be used subcutaneously. 2,27A 30G needle is used to inject microstructures into the target areas of the skin The usual volume of Nanofat is 8-10 mL on each side of the face, or a total of 16-20 mL. 2,18Since Nanofat regenerates tissue and improves skin quality in wounds, it has the potential to diminish the appearance of superficial wrinkles, such as those around the eyes or lips.Importantly, the Nanofat is commonly used to improve the overall quality of the skin. 1,2,27Nanofat has a strong vascularization capability and can be employed to increase tissue vascularization and as a potential source of functioning microvessel segments.Post-operation The patient requires special care following surgery and fat tissue transplantation.Antibacterial lotions and heparin cream can be administered locally to prevent infection and ecchymosis, respectively.Up to 6 weeks after the operation, the patient may experience some degrees of bruising and swelling.Deep massage, tapping, pressure, and the use of corticosteroids should be avoided to improve the function of fat tissue and prevent fat migration and necrosis.

F I G U R E 2
Different hand layers and suitable places for injection.The hand is generally divided into three layers: superficial, middle, and deep.The laminae are separated by the facial plates, and where the facial layers meet, they are closely bound by the extensor retinaculum.Despite the many differences in the appropriate injection layer in terms of safety and performance, there are many agreements for injection only to the surface layer because there is no important structure in it.For injection, cannulas with sizes between 14 and 17 G are used as standard.The amount recommended by experts according to the technique used and the desired amount of correction is 15-30 mL of fat for the back of each hand.

7 |
CON CLUS IONRejuvenation, beauty, and cell growth power restoration have long been studied, and this subject has recently gained increased attention.Most individuals care about looking younger, without wrinkles and deep and sunken areas in their faces, or the beauty of their hands.Many people, on the other hand, are bothered by the position and appearance of stitches and wounds for a variety of reasons.Furthermore, people are frequently looking for a technique to repair their hair growth due to daily stress and genetics.As a result, numerous approaches have been created to overcome these problems, and the transplantation of fat and its stem cells with long-term effects, fewer complications, natural, and effective has garnered a lot of attention.Fat cells have a lot of energy storage and a lengthy lifespan.
Variety of different fat transplant methods for rejuvenation.An overview of liposuction procedures and types.
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