- Email: [email protected]
Cryopreserved human skin allografts: Viability and delay of rejection in severely burned patients
616
burns 38 (2012) 614–618
Philip Wright* Wessex Specialist Laser Centre, Salisbury NHS Foundation Trust, Salisbury, Wiltshire, UK *Corresponding author. Tel.: +44 01722 345 520; fax: +44 01722 345 519 E-mail address: [email protected] (P. Wright) 0305-4179/$36.00 # 2012 Elsevier Ltd and ISBI. All rights reserved. doi:10.1016/j.burns.2011.12.024
Letter to the Editor Cryopreserved human skin allografts: Viability and delay of rejection in severely burned patients
Dear Editor, In case of extensive burns, human skin allograft rejection is delayed as a consequence of the immunodepression induced by burn injury. Nevertheless, rejection delay is not only linked to the recipient, but also related to skin preservation modalities. Thus, allografts could be cryopreserved, stored at +4 8C or glycerol-preserved with high density of glycerol. Depending on the modalities, it is possible to divide allografts into two categories: viable (i.e. allografts cryopreserved or stored at +4 8C) and non-viable (i.e. glycerol-preserved allografts). Skin viability can be tested by various methods, including the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, a metabolic assay for which
reproducibility and reliability have been successfully reported [1–3]. A debate exists regarding the importance of using viable, less viable or even non-viable allografts in order to guarantee full function of skin substitute. Currently, there is no established correlation between viability, efficacy of covery, intensity of rejection, and delay of healing in burned patients. Some practitioners consider viability essential for better engraftment or to improve tissue granulation, while others prefer a non-viable product for reasons of storage convenience and cost (storage at ambient temperature versus storage in liquid nitrogen for cryopreserved products). In our tissue bank, human skin allografts are cryopreserved with 15% glycerol. Procurement of skin, cryopreservation procedure and viability assessment have been previously reported [4]. Between July 2006 and September 2008, 400 patients have been admitted in our referral university burn center. Among these patients, 21 were treated with cryopreserved allografts. For 9 of these 21 allografted, retrospective data analysis allowed us to access both to allograft viability and rejection delay after grafting. Viability had been assessed in routine by the tissue bank team using the MTT assay both immediately after harvesting and after thawing (at the same time of the delivery to the burn unit). Rejection delay was assessed by retrospective examination of daily pictures of admitted patients. Clinical data are shown in Table 1. The mean delay of rejection was 13 days which is consistent with the literature. We did not find correlation between viability and rejection delay (Fig. 1), although in 1990, Kearney found, in a mouse study, a strong correlation between the viability of the skin (assessed by Tetrazolium Reductase activity) and the percentage of skin with graft adherence [5]. Many explanations could be offered for our divergent results. First and foremost, low number of patients was included in this preliminary study. Second, graft success is probably not affected only by skin viability. The poor and
Table 1 – Description of the 9 patients. Patients 1 and 8 had been allografted twice. Patient
Gender
1
M
Age year
Agent
TBSA%a (FT TBSA%b)
Allograft total grafted area (cm2)
19
Flame
68 (40)
7.102
Rejection delay (day)
Final outcome (day)
14
Discharge (day 122)
13 14 14 13 12 12 9
Discharge (day Discharge (day Discharge (day Death (day 61) Discharge (day Discharge (day Death (day 58)
16
Discharge (day 97)
13 2 3 4 5 6 7 8
M M M F M M M
50 23 52 54 50 31 57
Flame Flame Flame Scald Flame Flame Flame
75 44 55 42 45 50 90
(75) (44) (50) (24) (25) (25) (60)
10.240 7.981 7.527 5.956 1.361 7.356 22.707
9
M
32
Flame
52 (38)
10.343
129) 108) 83) 37) 152)
10 a b
Percentage of total body surface area. Percentage of full thickness total body surface area.
617
burns 38 (2012) 614–618
Rejection delay (day)
18 16 14 12 10 8 6 4 2 0 0,0
0,5
1,0
1,5
2,0
OD (570 nm)
Fig. 1 – Evaluation of the rejection delay and the viability of the cryopreserved skin (optical density at 570 nm adjusted for weight). Statistical analyses were performed by Spearman test (Rho = S0.22, p = 0.46).
unstable overall health status of the patients, the administration of vasopressors, the local infectious processes within the skin, and the quality of surgical excision are also factors that directly influence whether skin grafts take. Lastly, what is the relevance of the MTT assay relative to the viability in clinical settings?
1.
Conflict of interest
*Corresponding author at: Service de chirurgie ge´ne´rale, plastique et ambulatoire, AP-HP Hoˆtel-Dieu, 1 place parvis Notre Dame, 75004 Paris, France. Tel.: +33 1 42 34 80 63; fax: +33 1 42 34 82 21 E-mail addresses: [email protected] [email protected] (S. Gaucher) 0305-4179/$36.00 # 2012 Elsevier Ltd and ISBI. All rights reserved. doi:10.1016/j.burns.2011.12.025
None.
references
Letter to the Editor [1] Klein MB, Shaw D, Barese S, Chapo GA, Cuono CB. A reliable and cost-effective in vitro assay of skin viability for skin banks and burn centers. J Burn Care Rehabil 1996;17:565–70. [2] Bravo D, Rigley TH, Gibran N, Strong DM, Newman-Gage. Effect of storage and preservation methods on viability in transplantable human skin allografts. Burns 2000;26: 367–78. [3] Castagnoli C, Alotto D, Cambieri I, Casimiri R, Aluffi M, Stella M, et al. Evaluation of donor skin viability: fresh and cryopreserved skin using tetrazolium assay. Burns 2003;29:759–67. [4] Gaucher S, Elie C, Ve´rola O, Jarraya M. Viability of cryopreserved human skin allografts: effects of transport media and cryoprotectant. Cell Tissue Bank February 2011 [Epub ahead of print]. [5] Kearney JN, Wheldon LA, Gowland G. Cryopreservation of skin using a murine model: validation of prognostic viability assay. Cryobiology 1990;27:24–30.
Sonia Gauchera,b,c,* Universite´ Paris Descartes, Paris Sorbonne Cite´, 75006 Paris, France b Service de chirurgie ge´ne´rale, plastique et ambulatoire, AP-HP Hoˆtel-Dieu, 75004 Paris, France c Service des bruˆle´s, AP-HP Hoˆpital Cochin, 75014 Paris, France
a
Mohamed Jarraya Banque des Tissus Humains, AP-HP Hoˆpital Saint Louis, 75010 Paris, France
A call for evidence: Timing of surgery in burns Dear Editor, Despite the progress in burn care, there is still no worldwide consensus on the best timing of surgery in burns. In some countries, for example the United Kingdom (UK), early excision and grafting is well accepted, while in other countries, like the Netherlands, delayed excision and grafting is the usual approach. This difference was the topic of a ‘‘battle’’ between two burn surgeons: A. Kay, Royal Centre for Defence Medicine (UK) and G.I.J.M. Beerthuizen, Martini Hospital Groningen (The Netherlands), organised at the European Burn Association Congress in The Hague on September 24th, 2011. Kay presented his vision: early excision and grafting, why not? In his opinion, early excision in massive deep burns should be performed at admission when the burns are scrubbed for proper assessment and escharotomies might be performed. Kay stated that it would be illogical to stop at this point and not remove the dead tissue that stimulates the ongoing inflammatory response. He referred to a metaanalysis [1] in which several studies, published since the introduction of early excision and grafting in the 1970s, are combined. The authors of this review concluded that early excision and grafting is beneficial in reducing mortality and
burns 38 (2012) 614–618
Philip Wright* Wessex Specialist Laser Centre, Salisbury NHS Foundation Trust, Salisbury, Wiltshire, UK *Corresponding author. Tel.: +44 01722 345 520; fax: +44 01722 345 519 E-mail address: [email protected] (P. Wright) 0305-4179/$36.00 # 2012 Elsevier Ltd and ISBI. All rights reserved. doi:10.1016/j.burns.2011.12.024
Letter to the Editor Cryopreserved human skin allografts: Viability and delay of rejection in severely burned patients
Dear Editor, In case of extensive burns, human skin allograft rejection is delayed as a consequence of the immunodepression induced by burn injury. Nevertheless, rejection delay is not only linked to the recipient, but also related to skin preservation modalities. Thus, allografts could be cryopreserved, stored at +4 8C or glycerol-preserved with high density of glycerol. Depending on the modalities, it is possible to divide allografts into two categories: viable (i.e. allografts cryopreserved or stored at +4 8C) and non-viable (i.e. glycerol-preserved allografts). Skin viability can be tested by various methods, including the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) assay, a metabolic assay for which
reproducibility and reliability have been successfully reported [1–3]. A debate exists regarding the importance of using viable, less viable or even non-viable allografts in order to guarantee full function of skin substitute. Currently, there is no established correlation between viability, efficacy of covery, intensity of rejection, and delay of healing in burned patients. Some practitioners consider viability essential for better engraftment or to improve tissue granulation, while others prefer a non-viable product for reasons of storage convenience and cost (storage at ambient temperature versus storage in liquid nitrogen for cryopreserved products). In our tissue bank, human skin allografts are cryopreserved with 15% glycerol. Procurement of skin, cryopreservation procedure and viability assessment have been previously reported [4]. Between July 2006 and September 2008, 400 patients have been admitted in our referral university burn center. Among these patients, 21 were treated with cryopreserved allografts. For 9 of these 21 allografted, retrospective data analysis allowed us to access both to allograft viability and rejection delay after grafting. Viability had been assessed in routine by the tissue bank team using the MTT assay both immediately after harvesting and after thawing (at the same time of the delivery to the burn unit). Rejection delay was assessed by retrospective examination of daily pictures of admitted patients. Clinical data are shown in Table 1. The mean delay of rejection was 13 days which is consistent with the literature. We did not find correlation between viability and rejection delay (Fig. 1), although in 1990, Kearney found, in a mouse study, a strong correlation between the viability of the skin (assessed by Tetrazolium Reductase activity) and the percentage of skin with graft adherence [5]. Many explanations could be offered for our divergent results. First and foremost, low number of patients was included in this preliminary study. Second, graft success is probably not affected only by skin viability. The poor and
Table 1 – Description of the 9 patients. Patients 1 and 8 had been allografted twice. Patient
Gender
1
M
Age year
Agent
TBSA%a (FT TBSA%b)
Allograft total grafted area (cm2)
19
Flame
68 (40)
7.102
Rejection delay (day)
Final outcome (day)
14
Discharge (day 122)
13 14 14 13 12 12 9
Discharge (day Discharge (day Discharge (day Death (day 61) Discharge (day Discharge (day Death (day 58)
16
Discharge (day 97)
13 2 3 4 5 6 7 8
M M M F M M M
50 23 52 54 50 31 57
Flame Flame Flame Scald Flame Flame Flame
75 44 55 42 45 50 90
(75) (44) (50) (24) (25) (25) (60)
10.240 7.981 7.527 5.956 1.361 7.356 22.707
9
M
32
Flame
52 (38)
10.343
129) 108) 83) 37) 152)
10 a b
Percentage of total body surface area. Percentage of full thickness total body surface area.
617
burns 38 (2012) 614–618
Rejection delay (day)
18 16 14 12 10 8 6 4 2 0 0,0
0,5
1,0
1,5
2,0
OD (570 nm)
Fig. 1 – Evaluation of the rejection delay and the viability of the cryopreserved skin (optical density at 570 nm adjusted for weight). Statistical analyses were performed by Spearman test (Rho = S0.22, p = 0.46).
unstable overall health status of the patients, the administration of vasopressors, the local infectious processes within the skin, and the quality of surgical excision are also factors that directly influence whether skin grafts take. Lastly, what is the relevance of the MTT assay relative to the viability in clinical settings?
1.
Conflict of interest
*Corresponding author at: Service de chirurgie ge´ne´rale, plastique et ambulatoire, AP-HP Hoˆtel-Dieu, 1 place parvis Notre Dame, 75004 Paris, France. Tel.: +33 1 42 34 80 63; fax: +33 1 42 34 82 21 E-mail addresses: [email protected] [email protected] (S. Gaucher) 0305-4179/$36.00 # 2012 Elsevier Ltd and ISBI. All rights reserved. doi:10.1016/j.burns.2011.12.025
None.
references
Letter to the Editor [1] Klein MB, Shaw D, Barese S, Chapo GA, Cuono CB. A reliable and cost-effective in vitro assay of skin viability for skin banks and burn centers. J Burn Care Rehabil 1996;17:565–70. [2] Bravo D, Rigley TH, Gibran N, Strong DM, Newman-Gage. Effect of storage and preservation methods on viability in transplantable human skin allografts. Burns 2000;26: 367–78. [3] Castagnoli C, Alotto D, Cambieri I, Casimiri R, Aluffi M, Stella M, et al. Evaluation of donor skin viability: fresh and cryopreserved skin using tetrazolium assay. Burns 2003;29:759–67. [4] Gaucher S, Elie C, Ve´rola O, Jarraya M. Viability of cryopreserved human skin allografts: effects of transport media and cryoprotectant. Cell Tissue Bank February 2011 [Epub ahead of print]. [5] Kearney JN, Wheldon LA, Gowland G. Cryopreservation of skin using a murine model: validation of prognostic viability assay. Cryobiology 1990;27:24–30.
Sonia Gauchera,b,c,* Universite´ Paris Descartes, Paris Sorbonne Cite´, 75006 Paris, France b Service de chirurgie ge´ne´rale, plastique et ambulatoire, AP-HP Hoˆtel-Dieu, 75004 Paris, France c Service des bruˆle´s, AP-HP Hoˆpital Cochin, 75014 Paris, France
a
Mohamed Jarraya Banque des Tissus Humains, AP-HP Hoˆpital Saint Louis, 75010 Paris, France
A call for evidence: Timing of surgery in burns Dear Editor, Despite the progress in burn care, there is still no worldwide consensus on the best timing of surgery in burns. In some countries, for example the United Kingdom (UK), early excision and grafting is well accepted, while in other countries, like the Netherlands, delayed excision and grafting is the usual approach. This difference was the topic of a ‘‘battle’’ between two burn surgeons: A. Kay, Royal Centre for Defence Medicine (UK) and G.I.J.M. Beerthuizen, Martini Hospital Groningen (The Netherlands), organised at the European Burn Association Congress in The Hague on September 24th, 2011. Kay presented his vision: early excision and grafting, why not? In his opinion, early excision in massive deep burns should be performed at admission when the burns are scrubbed for proper assessment and escharotomies might be performed. Kay stated that it would be illogical to stop at this point and not remove the dead tissue that stimulates the ongoing inflammatory response. He referred to a metaanalysis [1] in which several studies, published since the introduction of early excision and grafting in the 1970s, are combined. The authors of this review concluded that early excision and grafting is beneficial in reducing mortality and