Is Nanofat the Long-Awaited Treatment for Hypertensive Ischemic Leg Ulcers?

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Martorell ulcer, also known as HYTILU or Martorell hypertensive ischemic ulcer, is a type of chronic wound that typically occurs on the lower extremities. It was first described by the Spanish dermatologist Fernandes Martorell in 1945.¹ These ulcers are primarily associated with poorly controlled hypertension and arteriolosclerosis, leading to reduced blood flow and ischemia in the affected areas. The characteristics of Martorell ulcer include painful, necrotic ulcers with well-defined margins, which typically occur on the lateral aspects of the legs. The condition is more prevalent in middle-aged and older patients, with a higher incidence in women than men.¹ Some data show that in Europe, Martorell ulcer represents 10% to 15% of leg ulcers in patients hospitalized in a dermatological ward.²

The occurrence of HYTILU is linked to several risk factors, with hypertension being the primary one.^3–5 The main pathophysiological mechanism involves acquired microangiopathy of the dermal vessels due to arteriolosclerosis.^6,7 These changes in vessel walls lead to narrowing and subsequent arteriolar occlusion, resulting in decreased local perfusion, ischemia, and painful skin necrosis. This necrosis evolves into an ulceration that is characterized by a necrotic base bordered by a purpuric border, reflecting the progression of the inflammatory process. The ulcer may become deep, exposing the underlying structures (tendons, bones, and vascular and neural structures). The classic location, reported in 90% of cases, is the posterolateral aspect of the lower two-thirds of the leg.^3,8 

There is a discrepancy between the initial size of the ulcer (small) and the initial level of pain (high). The pain is intense, permanent, and chronic, impairing the quality of the patient’s sleep.⁴ Sixty percent of patients with HYTILU use opioids.⁹ Chronic pain and the need for daily local care lead to impaired quality of life and social isolation.^4,10 Treatment of these ulcers is a real challenge. Control of risk factors is the cornerstone of treatment but is not sufficient to stop the progression of HYTILU.^3,4,7 

Currently, medical-surgical treatment of HYTILU is associated with an average healing time of 4 to 11 months.^11–13 Necrosectomy followed by skin grafting is the most effective treatment.⁴ However, skin grafting is the only real surgical treatment, yielding poor results. In fact, studies indicate that there is a high incidence of skin graft necrosis, approximately 40%, along with a recurrence of the pathology.^4,9 This method, necrosectomy followed by skin grafting, is used primarily to alleviate severe pain.^1,10,12 

ADSCs represent a promising frontier in regenerative medicine because of their abundant presence in adipose tissue, ease of harvest, and potent regenerative capabilities. These multipotent cells have the capacity to differentiate into various cell lineages that are not limited to adipocytes, chondrocytes, osteocytes, or myocytes, making them versatile tools for tissue engineering and regenerative therapies.¹⁴ Moreover, ADSCs secrete a range of bioactive molecules that modulate the immune response, increase angiogenesis, and support tissue repair, making them particularly valuable for treating chronic wounds and ischemic conditions.¹⁵ 

Ebrahimian et al¹⁶ conducted an experimental study on mice and demonstrated the efficacy of ADSCs in promoting wound healing. These cells not only have the ability to morph into keratinocytes but also secrete pivotal growth factors, namely, KGF and VEGF. KGF, which belongs to the fibroblast growth factor family, plays crucial roles in both the development of epithelial tissues and the wound healing process, highlighting the versatile therapeutic potential of ADSCs in tissue regeneration.^16,17 

Given the complex pathophysiology of HYTILU, characterized by arteriolosclerosis-induced ischemia leading to painful ulcerations, the regenerative potential of ADSCs offers a novel therapeutic pathway. By promoting neovascularization and modulating the inflammatory milieu, ADSCs can address the underlying ischemia and support the healing of these recalcitrant ulcers.¹⁸ The autologous nature of ADSC therapy circumvents the risks associated with immune rejection, further highlighting its therapeutic appeal for conditions such as HYTILU, for which current treatment options are limited and often unsatisfactory.

The primary aim of the present preliminary study was to evaluate the therapeutic efficacy of adipose-derived mesenchymal stem cell autografting via the nanofat technique in patients with HYTILU. The secondary objectives were to determine the analgesic effect of this technique and its role in improving patients’ quality of life.

From September 9, 2019, to July 24, 2023, 23 patients with HYTILU were treated in the Department of Maxillofacial, Plastic, and Reconstructive Surgery at the University Hospital of Dijon in Dijon, France. All 23 patients were included in this retrospective, single-center pilot study after providing signed informed consent forms. Adult patients with at least 1 HYTILU lesion were included in the study. The diagnosis was clinical and included a necrotic leg ulcer, chronic high blood pressure, diabetes, and severe persistent pain. The exclusion criteria are listed in the decision flow diagram (Figure 1).

Surgical technique

The same surgical procedure was used for each patient. It was performed under local anesthesia and lasted approximately 45 minutes. The procedure consisted of 2 steps: liposuction of the abdomen or thigh with preparation of the nanofat according to the Tonnard technique,^16,19 followed by intradermal injection of the nanofat at the recipient site.

The first stage of the operation began with local anesthesia at the donor site. A small incision on the liposuction area allowed infiltration of the anesthetic mixture. Suction-assisted liposuction was performed under vacuum using a multiperforated cannula with 1-mm–diameter holes (Trivisonno Micro Harvester; Tulip Medical Products) connected to a 10-mL Luer Lock syringe. The collected adipose tissue was distributed into several 10-mL syringes and left to settle. The incision area was sutured and covered with a dry dressing. After a few minutes of settling, 3 phases were distinguished as follows, from top to bottom: an oily layer (damaged adipocytes), purified fat, and a lower layer consisting of blood and the infiltration product. In the last phase, the cup was emptied on a compress. The oil phase was removed by transferring only the fat into a new 10-mL syringe.

The fat was then emulsified. A filled and an empty 10-mL syringe were connected via a 2.4-mm female-female Luer Lock connector (Anaerobic Sizing Transfer; Tulip Medical Products), through which 30 passes were made. This process was repeated through a similar 1.4-mm and then a 1.2-mm connector. This resulted in a whitish solution.

The next step was a single pass of this solution through a proprietary reusable adipose tissue device that uniformly sizes harvested tissue for injection and incorporates a 500-µm disposable filter (NanoTransfer Set Generation I; Tulip Medical Products [hereinafter “transfer device”]). This filter retained fat cell debris and remaining connective tissue. On the other side of the filter, an empty 1-mL syringe screwed onto the transfer device was used to collect the adipocyte-free nanofat. Several 1-mL syringes of nanofat were then ready for injection. Local anesthesia was then applied at the recipient site. Once the detersion and trimming were completed, using a 1-mL syringe and a 30-gauge needle the nanofat was injected intradermally on the edges around the ulcer and under the wound bed at a rate of 0.1 mL every 2 cm. The first dressing, which had a silicone interface, was left in place for 3 days. Class II venous support, defined as moderate compression of 15 to 20 mm Hg according to the French classification for venous limb compression, was provided if indicated. The first dressing change was performed on day 3 in clinic and then daily at home. No immobilization or off-loading of the lower limb was necessary, and patients were able to walk immediately after the procedure.

Primary and secondary outcomes

The primary outcome was the percentage of ulcers healed 3 and 6 months postoperative. Photographs of the wounds were taken by a doctor using a digital single-lens reflex camera with a fixed 50-mm focal length lens. At each follow-up visit, the photographs were imported into the open-source image processing software ImageJ²⁰ and the residual surface area was assessed. Calibration was performed to obtain the equivalent of 1 cm in pixels to obtain a surface area result in square centimeters.

Assessment of pain via the VAS, the scar characteristics, and the effect of scarring on quality of life were the 3 secondary end points. The study of the scar after complete healing of the ulcer was performed via 2 questionnaires: the SCAR-Q patient-reported outcome measure, and the VSS,²¹ which was completed by the operator. In this study, the 3 SCAR-Q module scores (appearance, symptoms, and psychosocial impact) were summed for a total of 300, differing from the standard method in which each module is scored separately out of 100. Higher scores indicate better outcomes, with 300 being the maximum possible score. The VSS is a widely used tool for assessing scar severity on the basis of vascularity, pliability, height, and pigmentation. Scores range from 0 to 13, with higher scores indicating more severe scarring. The effect on quality of life was assessed via the Nottingham Health Profile²² before treatment, at 3 months after injection, and after complete healing.

Clinical follow-up with photographs and VAS measurements was performed preoperatively, on the day of surgery (day 0) and then at day 3, day 7, day 15, month 1, month 2, and month 3 postoperatively, and then every month until complete healing was achieved.

Statistical analysis

The results are expressed as mean (SD) for populations with a normal distribution, and as median (IQR) for populations with an asymmetric distribution. The primary end point (ie, the ulcer area) was compared via the nonparametric Wilcoxon matched-pairs signed rank test, whereas the other quantitative variables (VAS, quality of life improvement) are expressed as differences in means and were analyzed via analysis of variance. Alpha was set at .05, and P < .05 was considered significant. Calculations were performed via GraphPad Prism software (version 10; GraphPad Software).

The study included 23 patients, with 6 females and 17 males. Patient age ranged from 51 years to 88 years (mean, 74 years). All patients had been living with hypertension for many years (average, 24.5 years). Furthermore, 82.6% of the patients had diabetes, with a mean (SD) hemoglobin A1c of 7.1% (9.0%). The mean duration of diabetes was 19 years. Fourteen ulcers were located on the anterolateral aspect of the leg, 6 were located posterolateral, and 3 were circumferential. The mean ulcer duration pretreatment was 10 months (Table 1).

Initial ulcer size varied, ranging from 4.8 cm² to 158.1 cm² (mean, 39.69 cm²; median, 30.9 cm² [IQR, 21.9 cm²–38.9 cm²]) (Figure 2). The average wound healing rate was 89.29% at 3 months, improving to 99.29% at 6 months (P < .0001). The healing time ranged from 3 to 12 months, with a mean of 4.65 months. All patients achieved complete healing (Figure 3).

All 23 patients experienced pain before treatment, with a mean (SD) VAS pain score before nanofat injection of 5.87 (2.12), which significantly decreased to 2.87 (2.38) 1 week after treatment and to 0.39 (1.08) at 3 months after the injection (P < .0001 for both time points). Before treatment, 15 patients were using opioids. All patients stopped using opioids; the mean (SD) time to stopping opioids was 2.4 (1.73) weeks.

Once complete healing was achieved, patients were assessed using the SCAR-Q and the VSS. The SCAR-Q scores ranged from 278 to 300 (mean [SD], 286 [4]). Scar quality assessment scores per the VSS ranged from 2 to 7 (mean [SD] 4.00 [1.21]) (Table 1).

Significant changes in the effect on the quality-of-life criteria measured by the Nottingham Health Profile were shown, with 100% of patients experiencing pain before surgery (n = 23) and 0% (n = 0) experiencing pain after complete healing. Physical mobility issues were reported by 100% of patients before surgery (n = 23) vs 4.3% after healing (n = 1). Social isolation was reported by 78.3% of patients before surgery (n = 18) and by 26.1% (n = 6) after healing. Energy level was affected in 82.6% of patients before surgery (n = 19) and in 21.7% (n = 5) after surgery. Emotional reactions were noted by 100% of patients before surgery (n = 23) and by 30.4% (n = 7) after surgery. Sleep disturbances were reported by 95.7% of patients before surgery (n = 22) and 17.4% (n = 4) after healing (Table 2).

The present study evaluated the effectiveness of nanofat injections in treating HYTILU. The study authors did not anticipate these unrealistic results. The treatment not only resulted in complete healing of the wounds but also significantly decreased pain. 

The Dermatology Department at Dijon University Hospital, a major health care institution with over 1800 beds, plays a pivotal role in patient care. This department includes a specialized unit with 36 beds for hospitalized patients and a consultation area managing approximately 18000 visits each year. The department serves as a regional reference center for dermatological conditions, and the staff have expertise in areas such as wound care, with a particular focus on patients with HYTILU in the eastern region of France.

The findings of the present study, which included a wide range of initial ulcer sizes and in which complete healing was achieved in all patients within an average 4.65 months, underscore the robust healing capabilities of nanofat. Particularly notable was the substantial improvement in mean wound healing rate, which significantly increased from 89.29% at 3 months to 99.29% by 6 months (P < .0001). The maximum healing time was observed in patient 3, who had a large circumferential ulcer (Table 1). Only this patient required a second operation at month 3 because of insufficient and slow healing. This second procedure resulted in complete healing 6 months after the second injection (12 months after the first injection). These findings resonate with the regenerative properties of nanofat reported by Tonnard et al¹⁹ in 2013, who highlighted the potential of nanofat for skin rejuvenation and its applicability in improving the management of chronic wounds.

A prospective study by Chopinaud et al,²³ published in 2017, investigated autologous fat grafting for the treatment of hypertensive leg ulcers in 10 patients of whom one smoker was lost to follow-up at 5 months, leaving 9 patients for analysis. Liposuction and fat grafting were performed under general anesthesia. After centrifugation, the purified fat was injected around and under the wound without prior emulsification or filtration. Those authors reported a significant decrease in ulcer area, with complete healing in 1 patient at 3 months and in 3 other patients at 4 months. Among the 9 nonsmokers, the median healing percentage was 73.2% at 3 months and 93.1% at 6 months. Additionally, a significant reduction in pain was reported, with a mean VAS score of less than 1 out of 10 from the first month onward (P < .001).

Necrosectomy followed by skin grafting is currently the standard treatment for HYTILU. The graft promotes epithelialization and reduces healing time by releasing growth factors and vasodilator peptides.²⁴ In 30% to 40% of cases, however, multiple successive grafts are necessary. In a retrospective series involving 31 patients with HYTILU, 26 of 29 patients received a skin graft immediately following necrosectomy; healing was achieved with a single skin graft in 14 patients, while the remaining 12 experienced recurrence despite the initial graft.¹ In the present study, only patient number 3 required a second injection of nanofat.

Postoperative pain reduction was another significant outcome in the present study, with the average VAS pain score significantly decreasing from 5.87 before treatment to 0.39 at 3 months after nanofat injection (P < .0001). This substantial decrease in pain, coupled with the cessation of opioid use in all patients, highlights the analgesic properties of nanofat grafting. This finding is in line with those of Jan et al,²⁵ who in 2019 demonstrated that nanofat is an effective solution for alleviating pain in postburn scars; together, these findings suggest the broad analgesic application of nanofat in various types of chronic wounds.

Skin graft harvest can cause pain for several weeks and can leave a scar at the donor site. A scar from a grafted ulcer often differs in color and elasticity from the surrounding skin. When skin grafts necrose, direct healing results in sclerotic, fibrous, hypo- and/or hyperpigmented scars, along with vascular abnormalities. ADSCs are typically collected under local anesthesia on an outpatient basis. Nanofat, which does not utilize healthy skin, leaves only a discreet scar in the liposuction area. Injected ADSCs around the ulcer facilitate epidermal restoration without atrophy. The healed skin is of high quality, supple, and nearly identical in color to the surrounding skin, as confirmed by the VSS and SCAR-Q results.^26,27 Stem cells begin to proliferate between the first and fourth week after injection,²⁸ stimulating various wound healing agents through their paracrine, anti-inflammatory, and proangiogenic functions.²⁹ Tissue remodeling occurs within the first 3 months after injection, with tissue action persisting beyond 3 months and peaking between 4 and 6 months postoperative.^19,30

Adverse effects were minimal in the present study, with some patients experiencing infection, hematoma, or pain at the liposuction site. At the injection site, reactions included temporary discoloration of the yellowish skin, inflammation, and short-term swelling. Notably, patient number 4 in the present study died. This patient had numerous comorbidities and developed septic shock of pulmonary origin 7 months after inclusion.

Moreover, the results of the posttreatment assessment of scar quality and quality of life in the present study further corroborate the comprehensive benefits of nanofat grafting. The SCAR-Q scores, ranging from 278 to 300 (mean, 286), and the VSS scores, ranging from 2 to 7 (mean, 4), indicated substantial improvements in scar appearance and quality (Table 1). These improvements, combined with dramatic enhancements in Nottingham Health Profile quality-of-life criteria (Table 2), from pain and physical mobility issues to emotional reactions and sleep disturbances, highlight the multifaceted effect of nanofat treatment. These outcomes underscore not only the regenerative capacity of nanofat but also its role in improving patients’ overall quality of life, which is consistent with the findings of Uyulmaz et al³¹ on the beneficial effects of nanofat on skin and scar quality. In the present study, all procedures were performed under local anesthesia, and no cases of recurrence were identified at nearly 1 year in the 22 surviving patients. 

The present study has limitations. The small sample size of 23 patients may limit the generalizability of these findings to a broader population. Additionally, the retrospective single-center design could introduce selection and reporting biases. Furthermore, the absence of a control group in the study design prevents definitive conclusions about the clinical improvements being solely attributable to nanofat grafting because other confounding factors could also play a role.

The present study showed promising results, with complete healing at an average of 4.65 months, a decrease in mean VAS pain score to 0.39 at 3 months, and cessation of opioid use at a mean of 2.4 weeks after the start of nanofat treatment, as well as good-quality scarring per SCAR-Q and VSS scores. Nottingham Health Profile scores indicated greatly improved quality of life after nanofat treatment. The authors of the present study plan to design a comparative study to assess the efficacy of nanofat grafting vs skin grafting, the current standard procedure, in the management of HYTILUs.