Table 3.

Review of all the clinical studies that have utilised adipose derived stem cells (ADSCs) for cranial facial surgery over the last 10 years

Tissue Replaced	Year and Ref	Study Design	Methods	Outcome	Authors Conclusions
Fat	2008 [84]	3 patients underwent conventional lipoinjection (non-CAL), while 3 patients underwent CAL for facial augmentation.	1. Adipose portion of lipoaspirate digested with 0.075 % collagenase for 30 mins on a shaker at 37 °C.	CAL had a better clinical improvement score than non-CAL to 13 months follow up, although not significant (p = 0.11).	Longer follow up required. Safe and effective treatment.
			2. Mature adipocytes separated from the SVF containing ADSCs by centrifugation (800 g, 10 mins).
			3. The fluid portion was centrifuged (800 g, 5 mins), and the pellets were resuspended in hypotonic water to lyse erythrocytes.
			4. During the processing period, the other half of lipoaspirates was harvested as graft material.
			5. The adipose portion of liposuction aspirates was centrifuged at 700 g for 3 mins without washing. In the non-CAL group, centrifuged fat was injected without SVF supplementation.
			6. In the CAL group, the fresh SVF isolated from both the adipose and the fluid portion was added to the graft material and then put into injection syringe.
Fat	2012 [97]	CAL against traditional soft tissue grafting in 9 patients undergoing facial augmentation with follow-up for 12 weeks.	Similar technique to 2008 [84]	Volume and patient satisfaction was significantly greater for CAL assisted facial augmentation.	No significant adverse effects.
Fat	2012 [98]	5-year history of progressive right facial hemiatrophy, who underwent facial volumetric restoration using CAL.	Same technique as 2008 [84]	At 12 months better volume and symmetry of the frontotemporal region and malar prominence and cheek.	CAL has showed promising results in the long term by decreasing the rate of fat reabsorption.
Fat	2012 [99]	19 year old with dermatoses and contour deformities on her forehead. Total of 10 × 107 cell suspension in a 5-ml agitated with the 180-mL fat graft.	1. Lipoaspirate processed with pure graft system to create a pure fat graft.	Good patient satisfaction and cosmetic effect at 1 year	Prevented multiple surgeries for the patient.
			2. Graft divided into 2 parts, 180 ml spared for reconstruction and 360 ml introduced into the Celution system and processed for 2 h 25 mins.
			3. 10 x107 cell suspension in 5-mL syringe then used.
Fat	2012 [100]	10 patients with Parry-Romberg disease. 5 received ASC and microfat grafts and 5 received microfat grafts only. Follow up 15 months	1. Extracted ADSCs isolated similarly to Yoshimura et al. 2008.	Resorption in this ADSC group was 20.59 % compared to fat only group of 46.81 %.	A microfat graft with simultaneous ADSC injection may be used to treat Parry-Romberg disease without the need for microvascular free flap transfer.
			2. Cell seeded into a culture flask and cultured overnight.
			3. On day 14, patients were injected with secondary fat grafts and test patients simultaneously received 1 × 107 ADSCs.
Fat	2013 [101]	14 patients with craniofacial microsomia were grafted either with supplementation of ADSCs or without supplementation ADSCs.	Similar technique to 2008 [84]	Surviving fat volume at 6 months was 88 % for the experimental group and 54 % for the control group (p = 0.003).	Isolation and supplementation of ADSCs is effective, safe, and superior to conventional lipoinjection for facial recontouring in craniofacial microsomia.
Fat	2013 [102]	38 women who underwent fat transplantation with SVF (n = 26) or fat grafting alone (n = 12).	Similar technique to 2008 [84]	No complications were evidenced during follow-up. Fat survival was higher with SVF (64.8 ± 10.2 %) than fat grafting alone (46.4 ± 9.3 %) (p < 0.01).	Supplementing fat grafts with SVF for cosmetic facial contouring can improve the survival of fat grafts over fat grafting alone.
Bone	2011 [54]	4 patients with calvarial defects received autologous ADSCs seeded in bTCP granules. For 2 patients, a bilaminate technique with resorbable mesh was used.	1. ADSCs were grown under Good Manufacturing Practice for 22 days.	3 months no complications and CT scans ossification was similar to native bone.	The combination of scaffold material such as bTCP and autologous ADSCs constitutes a promising model for reconstruction of human large cranial defects.
			2. 15 x106 cells of passage 3 and 4 were subsequently combined with 60 mL of bTCP granules for 48 h before the operation.
Bone	2004 [55]	Calvarial defect of 7-year old using fibrin glue and ADSCs from the iliac bone.	1. ADSCs from the gluteal region during harvesting of the bone graft from iliac crest.	No complications and union at 3 months.	Further studies, both in vitro and in vivo, are needed to turn this first case into a reproducible and reliable treatment regimen in craniofacial bone reconstruction.
			2. Processing and isolation for 2 h.
			3.10 ml of the prepared solution of ADSCs was evenly applied to the cancellous bone grafts.
			4.To keep the cells in place, 8 ml of autologous (obtained preoperatively by plasmapheresis and cryoprecipitation) fibrin glue was applied using a spray adapter.
Bone	2009 [61]	Maxillary reconstruction following hemimaxillectomy due to a large keratocyst. ADSCs were expanded for 14 days prior to be seeded on a titanium cage with bTCP.	1. First operation was used to extract the ADSCs, which was then expanded for 14 days.	After 8 months, the flap had developed mature bone structures and was placed in the area without complication.	This is the first clinical case where ectopic bone was produced using autologous ADSCs in microvascular reconstruction surgery.
			2. Prior to combining the cells with beta-TCP, the beta-TCP was incubated for 48 h in basal medium containing 12 mg rhBMP-2
			3. Following the incubation, the media containing rhBMP-2 was discarded. Subsequently, to allow cell attachment, approximately 13 x106 cells were combined with 60 ml of bTCP granules 48 h prior to the operation.
			4. In the second operation a titanium cage filled with ADSCs and bTCP was inserted into the left rectus abdominis muscle.
			5. The rectus abdominis free flap was then raised 9 mths later to open the cage, disconnect the vessels and then the flap was placed in the maxillary defect.
Bone	2010 [60]	14-year-old adolescent boy with Treacher Collins syndrome whose bilateral orbitozygomatic defects were treated with engineered bone made from a combination of human bone allograft, ADSCs, BMP-2, and periosteal grafts.	1. 28 mm of fresh lipoaspirate from the abdomen was pipetted onto the bone allograft	The reconstruction remained stable during a 6-month follow-up, biopsy of the engineered bone showed health, lamellar bone.	The combination of ADSCs, BMP-2, bone allograft, and periosteum may provide an alternative method to both osteocutaneous free flaps and large structural allografts with less morbidity and improved long-term results.
			2. After this, each construct was covered with recombinant human BMP-2 on a collagen sponge.
			3.Lastly, periosteal grafts from the patient’s left femur were sewn into position over the bilateral bony constructs.
Bone	2012 [58]	All patients with jaw defects were reconstructed with ADSCs, resorbable scaffolds, and growth factor as required. Vascularized soft tissue beds were prepared for ectopic bone formation and later microvascular translocation as indicated.	Same technique as 2009 [61]	23 ADSCs seeded resorbable scaffolds combined with rhBMP-2 were successfully implanted to reconstruct jaws except for three failures (one infection and two cases of inadequate bone formation).	ADSC-aided reconstruction of large defects remains challenging as it takes longer and has a higher cost than the conventional standard immediate reconstruction but results are encouraging.
Bone	2013 [59]	10 cm anterior mandibular ameloblastoma resected and repaired using β-TCP granules, recombinant BMP-2, and Good Manufacturing ADSCs.	Similar technique to 2009 [61]	After 10 months dental implants could be implanted and prosthodontic rehabilitation was completed.	ADSCs in combination with β-TCP and BMP-2 good option for mandibular defects without the need for ectopic bone formation and allowing rehabilitation with dental implants.

Key; ADSCs adipose derived stem cells, bTCP beta tricalcium phosphate, CAL cell assisted lipotransfer, SVF stromal vascular fraction