Autologous Skin Fibroblast and Keratinocyte Grafts in the Treatment of Chronic Foot Ulcers in Aging Type 2 Diabetic Patients

Abstract

Background: We assessed the tolerability and efficacy of autologous skin cell grafts in older type 2 diabetic patients with chronic foot ulcers. Methods: Treatment with Hyalograft 3D and Laserskin Autograft was proposed to a consecutive series of type 2 diabetic patients older than 65 years affected by long-standing (>6 months) foot ulcers with an area greater than 15 cm². Ulcer healing rates and measurements of ulcer area were determined monthly for 12 months. Results: Seven patients with 12 ulcers, nine of which received the described treatment, were enrolled. During 12-month follow-up, all of the ulcers healed except one. In the remaining eight ulcers, the median healing time was 21 weeks (interquartile range, 4–29 weeks). Conclusions: Autologous skin cell grafts are feasible, well tolerated, and apparently effective in the treatment of diabetic ulcers of the lower limbs in advanced age. Age did not seem to moderate healing times.

Grafts formed from autologous fibroblasts and keratinocytes grown on a scaffold made from the benzyl ester of hyaluronic acid have been developed for the treatment of skin ulcers, including those of diabetic feet.[1] Case reports[2] and observational studies[3,4,5] showed that autologous fibroblast and keratinocyte grafts cultivated on a hyaluronic acid–derived scaffold (Hyalograft 3D [H3D] and Laserskin Autograft [LSK]; Fidia Advanced Biopolymers, Abano Terme, Italy) were effective and well tolerated in patients affected by diabetic foot ulcers. This result was confirmed by a multicenter, openlabel, randomized clinical trial.[3]

Most type 2 diabetic patients with foot ulcers are older than 65 years,[6,7] whereas many interventional studies have been performed in mostly middle-aged participants.[3,4,5] Advanced age is a well-known predictor of unfavorable outcome in diabetic foot ulcers,[4,8] even if some authors disagree[9]; in fact, frailty, memory loss, immobility, and lack of independence, which are all associated with advanced age, could have a marked effect on wound healing. It is, therefore, reasonable to expect lower efficacy of treatment in older patients. Despite this fact, previous studies of autologous fibroblast and keratinocyte grafts (H3D/LSK) did not specifically explore the efficacy of such therapy in advanced age.

Research Design and Methods

Treatment with H3D/LSK was proposed to a consecutive series of type 2 diabetic patients older than 65 years referred to the Outpatient Diabetes Clinic of the Geriatric Unit of Careggi Hospital in Florence, Italy, for the treatment of long-standing (>6 months) foot ulcers with an area greater than 15 cm²; those with an ankle-brachial index less than 0.5 and those with exposed bone or osteomyelitis diagnosed by radiography were excluded. Antibiotic treatment continued until eradication of infection, demonstrated through a swab for bacteriologic analysis, before treatment with H3D/LSK; pressure relief for plantar ulcers was obtained with removable casts.

Ulcers were classified according to the University of Texas score.[10] The ulcer area in each patient was traced using a transparent plastic grid (OpSite; Smith & Nephew, London, England). The wound area was later calculated by computerized morphometric measurement.

The technique for H3D/LSK treatment has been described elsewhere in greater detail.[3] Briefly, an abdominal skin biopsy sample (1–2 cm², 0.8 mm deep) was sent to the TissueTech Autograft laboratory (Abano Terme, Padua, Italy), where fibroblasts and keratinocytes were isolated, propagated, and then seeded on two distinct biodegradable scaffolds. Fibroblasts (H3D) were grafted onto the debrided and cleansed wound. Further grafts, when required, were applied on the wound cleansed with physio-logic solution. Approximately 7 to 10 days after the last H3D grafting, the ulcer received keratinocytes (LSK). Subsequent ulcer care was provided by the clinic staff until healing (complete reepithelialization) or up to 1 year after the first graft.

Costs were calculated on the basis of direct expenses (for H3D/LSK and accessory materials), adding the cost of personnel with current salaries (€26.70 and €18.10 per hour for physicians and nurses, respectively), computed on the basis of actual time spent with each patient.

Cognitive function was explored with the Mini-Mental State Examination,[11] and functional status was assessed through basic activities of daily living[12] and instrumental activities of daily living.[13] Depressive symptoms were explored with the Geriatric Depression Scale.[14] The 36-Item Short Form Health Survey[15] and the Well-being Enquiry for Diabetics16 were used to assess generic and disease-specific health-related quality of life. Data are reported as mean ± SD or median (interquartile range) and are compared with t or Mann-Whitney tests whenever appropriate.

Results

Four of the 11 patients to whom this treatment had been proposed were excluded: three denied their written informed consent and one had inadequate control of local infection with antibiotic treatment before H3D/LSK therapy. The final sample comprised seven patients with 12 ulcers, nine of which received the described treatment. Of those ulcers, five were plantar, two were dorsal, and two were in the leg. The mean ± SD number of H3D grafts per ulcer was 2.3 ± 1.3, and one LSK graft was sufficient in all cases.

One patient had undergone surgical revascularization of the affected limb approximately 15 years before enrollment; none of the patients had undergone any revascularization procedure in the 5 years preceding inclusion in the present study.

During 12-month follow-up, all of the ulcers healed except one (in a patient with two treated ulcers), which showed a surface reduction of 44%. In the remaining eight ulcers, the median healing time was 21 weeks (interquartile range, 4–29 weeks). No adverse reactions to grafts were observed.

Ulcers that healed within 21 weeks (5 weeks in four patients) showed a trend toward a greater surface at baseline, and a lower ankle-brachial index, although differences did not reach statistical significance. The other clinical characteristics, including age, functional status, depressive symptoms, and health-related quality of life at baseline, did not seem to discriminate between the two groups (

Table 1. Baseline Demographic and Clinical Characteristics^a.

Abbreviations: ABI, ankle-brachial index (assessed at the side affected by the ulcer); ACE-I, angiotensin-converting enzyme inhibitor; ARB, angiotensin receptor blocker; BADL, basic activities of daily living; BMI, body mass index (calculated as weight in kilograms divided by the square of the height in meters); GDS, Geriatric Depression Scale; HbA_1c, hemoglobin A_1c; IADL, instrumental activities of daily living; MMSE, Mini-Mental State Examination; SF-36, 36-Item Short Form Health Survey; ESR, Erythrocyte sedimentation rate; VPT, vibration perception threshold (assessed at the great toe in triplicate); WED, Wellbeing Enquiry for Diabetics. ^aThere were no statistically significant differences between the groups. ^bP < .10.

).

The mean ± SD total costs for each patient were €3,478 ± €1,491 (range, €1,755–€5,740); of these, €2,960 ± €1,257, €106 ± €96, and €413 ± €313 were attributable to H3D/LSK, other material, and personnel, respectively.

Conclusions

Healing of long-standing foot ulcers with a wide surface in diabetic patients represents a challenging threat; advanced patient age can further aggravate the prognosis. Therapeutic options capable of accelerating healing, therefore, have major relevance for clinicians. Autologous skin cell grafts have been shown to reduce healing times in diabetic patients provided that sufficient blood flow and adequate pressure relief is warranted.[3]

The present study suggests that such a technique is effective in older individuals. Interestingly, age did not seem to moderate healing times in this series of patients. In fact, the only factors apparently associated with a delay in healing were a wider ulcer surface and the impairment of blood flow. Factors potentially affecting would healing that are associated with advanced age, such as mood depression and impairment of functional status and cognitive function, did not seem to moderate treatment outcome; however, the limited size of the sample suggests that caution be used to interpret these results. Also note that other factors associated with aging, such as reduced mobility, which were not assessed in the present study, could interfere with wound healing.

Any consideration of costs should be based on the availability of financial resources. However, the costs registered seem to be compatible with those of most health-care systems. Note that delay in healing is associated with additional costs for ulcer care and with increased risk of amputations.[8] Furthermore, it should be considered that improved healing could have a major impact on patients’ health-related quality of life. Future studies should include prospective evaluations of this parameter, which, in the present survey, was measured only at baseline.

An uncontrolled study, such as this pilot investigation, cannot provide any conclusion on efficacy and cost-effectiveness of treatments. A randomized controlled trial is needed to verify the effects of this treatment in older individuals. However, these data suggest that autologous skin fibroblast and keratinocyte grafts are feasible, well tolerated, and apparently effective in the treatment of diabetic ulcers of the lower limbs in advanced age.