Iontophoresis of treprostinil promotes wound healing in a murine model of scleroderma-related ulcers.

OBJECTIVE: Systemic sclerosis (SSc) is a rare, chronic disease characterized by fibrosis, vascular alterations and digital ulcerations. Few drugs have shown efficacy to enhance wound healing of existing SSc-related ulcers. Local delivery of treprostinil, a prostacyclin analogue, may improve wound healing. The present work aimed first at developing a mouse model of SSc-related ulcerations and second at assessing the effect of iontophoresis of treprostinil on wound healing. METHODS: We used two murine models of SSc: chemically induced with HOCl, and urokinase-type plasminogen activator receptor (uPAR)-deficient. Excisional wounding was performed on the dorsal midline with a biopsy punch. Animals were randomized into three groups: treated with electrostimulation alone, with treprostinil iontophoresis or untreated. We assessed wound healing over time, as well as skin microvascular reactivity, inflammation, microvessel density and collagen distribution, before wounding and after re-epithelialization. RESULTS: uPAR-/- mice, but not HOCl-treated mice, showed impaired wound healing and decreased microvascular reactivity compared with their controls. Treprostinil iontophoresis improved wound healing and microvascular density and decreased inflammation in uPAR-/- mice, while electro-stimulation did not. However, treprostinil had no effect on microvascular reactivity and collagen distribution. CONCLUSION: This study suggests that excisional wounds in uPAR-/- mice are a relevant model of SSc-related ulcers. In addition, treprostinil iontophoresis enhances wound healing in this model. Further work in now needed to show whether this effect translates in humans.

Alcohol-related hepatocellular carcinoma is a heterogenous condition: Lessons from a latent class analysis.

BACKGROUND: Alcohol-associated hepatocellular carcinoma (AL-HCC) poor prognosis has been attributed to diagnosis at a later stage. However, host factors and specific health trajectories have been associated with severe outcomes in alcohol-related liver disease. We hypothesize AL-HCC is not a homogeneous condition but encompasses subgroups yielding different outcomes. AIMS: Our aim was to provide a first attempt at a clinical phenotyping of AL-HCC. METHODS: We analysed data for the calendar years 2007-2013 from the French nationwide administrative hospital database. We selected patients with AL-HCC only. Clustering of AL-HCC phenotypes was performed by latent class analysis (LCA). RESULTS: The study included 11 363 patients with AL-HCC, mainly male (89.6%), median age 67 years [IQR: 61; 74] of which 71.2% had at least one metabolic comorbidity. Five phenotypes were identified. Phenotype 1 (41.4%) displayed high rates of unrecognized cirrhosis prior to HCC diagnosis (81%), low rates of metabolic comorbidities (diabetes 13%), and mostly compensated liver disease at HCC diagnosis while the four other phenotypes displayed high rates of metabolic comorbidities (diabetes up to 100%), various patterns of liver disease trajectories and overall 42% unrecognized cirrhosis. In adjusted survival analysis, compared to phenotype 1, risk of death after HCC diagnosis was significantly different for all phenotypes. CONCLUSION: LCA uncovers AL-HCC is a heterogeneous condition with distinct phenotypes yielding specific survival outcomes. Frequent unrecognized cirrhosis prior to HCC underlines the urgent need for implementing strategies to identify the underlying liver disease prior to HCC onset in patients with documented alcohol use disorders and metabolic comorbidities.

Hierarchical evaluation of electrical stimulation protocols for chronic wound healing: An effect size meta-analysis.

Electrical stimulation (ES) has been tested for decades to improve chronic wound healing. However, uncertainty remains on the magnitude of the efficacy and on the best applicable protocol. We conducted an effect size meta-analysis to assess the overall efficacy of ES on wound healing, to compare the efficacy of the different modalities of electrical stimulation, and to determine whether efficacy differs depending on the wound etiology, size, and age of the chronic wound. Twenty-nine randomized clinical trials with 1,510 patients and 1,753 ulcers were selected. Overall efficacy of ES on would healing was a 0.72 SMD (95% CI: 0.48, 1) corresponding to a moderate to large effect size. We found that unidirectional high voltage pulsed current (HVPC) with the active electrode over the wound was the best evidence-based protocol to improve wound healing with a 0.8 SMD (95% CI: 0.38, 1.21), while evaluation of the efficacy of direct current was limited by the small number of studies. ES was more effective on pressure ulcers compared to venous and diabetic ulcers, and efficacy trended to be inversely associated with the wound size and duration. This study confirms the overall efficacy of ES to enhance healing of chronic wounds and highlights the superiority of HVPC over other type of currents, which is more effective on pressure ulcers, and inversely associated with the wound size and duration. This will enable to standardize future ES practices.

Anodal iontophoresis of a soluble guanylate cyclase stimulator induces a sustained increase in skin blood flow in rats.

The treatment of systemic sclerosis-related digital ulcers is challenging. Although the only effective drugs are prostacyclin analogs, their use is limited by vasodilation-related adverse reactions. In this study, we assessed the local iontophoresis administration of three soluble guanylate cyclase (A-350619 [3-[2-[(4-chlorophenyl)thiophenyl]-N-[4-(dimethylamino)butyl]-2-propenamide hydrochloride], SIN-1 [amino-3-morpholinyl-1,2,3-oxadiazolium chloride], and CFM 1571 [3-[3-(dimethylamino)propoxy]-N-(4-methoxyphenyl)-1-(phenylmethyl)-1H-pyrazole-5-carboxamide hydrochloride]) and two nonprostanoid prostaglandin I2 (prostacyclin) receptor agonists (MRE-269 [[4-[(5,6-diphenylpyrazinyl)(1-methylethyl)amino]butoxy]-acetic acid] and BMY 45778 [[3-(4,5-diphenyl[2,4'-bioxazol]-5'-yl)phenoxy]acetic acid]) to induce vasodilation onto the hindquarters of anesthetized rats. Skin blood flow was quantified using laser Doppler imaging during the whole experience, and safety was assessed by continuous recording of blood pressure and histopathological examination. Anodal iontophoresis of A-350619 (7.54 mM) induced a sustained increase in cutaneous blood flow (P = 0.008 vs. control). All other drugs exhibited poor or no effect on skin blood flow. Vasodilation with A-350619 iontophoresis was concentration-dependent (7.5, 0.75, and 0.075 mM; P < 0.001, Jonckheere-Terpstra trend test), and repeated administrations do not suggest any risk of tolerance. This study also compared continuous versus intermittent iontophoresis protocols. Continuous anodal iontophoresis of A-350619 at 7.5 mM increases cutaneous blood flow with good local tolerance. Iontophoresis of soluble guanylate cyclase stimulators should be investigated as potential local therapy for digital ulceration in patients with scleroderma.

Chronic intermittent hypoxia, a hallmark of obstructive sleep apnea, promotes 4T1 breast cancer development through endothelin-1 receptors.

The association between obstructive sleep apnea (OSA) and cancer is still debated and data are scarce regarding the link between OSA and breast cancer progression. Since conclusive epidemiological studies require large sample sizes and sufficient duration of exposure before incident cancer occurrence, basic science studies represent the most promising approach to appropriately address the topic. Here we assessed the impact of intermittent hypoxia (IH), the major hallmark of OSA, on the development of breast cancer and explored the specific involvement of the endothelin signaling pathway. Original in vitro and in vivo models were used where 3D-spheroids or cultures of murine 4T1 breast cancer cells were submitted to IH cycles, and nude NMRI mice, orthotopically implanted with 4T1 cells, were submitted to chronic IH exposure before and after implantation. The role of the endothelin-1 in promoting cancer cell development was investigated using the dual endothelin receptor antagonist, macitentan. In vitro exposure to IH significantly increased 4T1 cell proliferation and migration. Meta-analysis of 4 independent in vivo experiments showed that chronic IH exposure promoted tumor growth, assessed by caliper measurement (overall standardized mean difference: 1.00 [0.45-1.55], p < 0.001), bioluminescence imaging (1.65 [0.59-2.71]; p < 0.01) and tumor weight (0.86 [0.31-1.41], p < 0.01), and enhanced metastatic pulmonary expansion (0.77 [0.12-1.42]; p = 0.01). Both in vitro and in vivo tumor-promoting effects of IH were reversed by macitentan. Overall, these findings demonstrate that chronic intermittent hypoxia exposure promotes breast cancer growth and malignancy and that dual endothelin receptor blockade prevents intermittent hypoxia-induced tumor development.

Treprostinil Hydrogel Iontophoresis in Systemic Sclerosis-Related Digital Skin Ulcers: A Safety Study.

Digital skin ulcers are a severe complication of systemic sclerosis. The first-line treatment is intravenous iloprost, but it induces dose-limiting adverse effects. Local administration of treprostinil through skin iontophoresis may be a safe alternative. We conducted a 2-stage, randomized, placebo-controlled single-ascending-dose study in healthy volunteers and patients with systemic sclerosis-related digital ulcer. We further explored the effect of the procedure on skin blood flux. In a first group of healthy subjects, treprostinil and placebo iontophoresis were performed at 3 locations (ie, 6 skin sites): the sole of the foot, the leg, and the fingers. We used a 1-mg/mL hydrogel of treprostinil. We then randomly treated systemic sclerosis-related digital ulcers in a 3:1 ratio of treprostinil or placebo. We used concentrations from 0.1 to 1 mg/mL. All adverse events were recorded and rated according to the Common Terminology Criteria for Adverse Events (CTCAE), whereas skin microvascular blood flux was recorded with laser speckle contrast imaging. Among the 12 healthy volunteers, we observed 60 local adverse effects: burns, skin pain, erythema, and pruritus, graded 1 or 2 on the 5-point CTCAE scale. Treprostinil iontophoresis significantly increased skin blood flux on the leg (AUC0-4 h at 88 460% ± 6436% versus 12 730% ± 3397% baseline flux.min respectively; P < .001) and on the sole of the foot (AUC0-3 h at 20 124% ± 6119% versus 3142% ± 3036% baseline flux.min, respectively; P = .018) with a trend on the finger. Among 5 patients with systemic sclerosis-related digital ulcer, 2 resolutive local adverse effects were reported. Iontophoresis of treprostinil hydrogel was safe in systemic sclerosis patients with digital ulcer.

Impact of the acute local inhibition of soluble epoxide hydrolase on diabetic skin microcirculatory dysfunction.

The impact of the local inhibition of soluble epoxide hydrolase, which metabolizes vasodilator and anti-inflammatory epoxyeicosanoids, on diabetic skin microvascular dysfunction was assessed. In diabetic db/db mice, basal skin blood flow assessed using laser Doppler imaging was similar to that of control mice, but thermal hyperemia was markedly reduced. At 2 h after the topical administration of an aqueous gel containing the soluble epoxide hydrolase inhibitor trans-4-[4-(3-adamantan-1-yl-ureido)-cyclohexyloxy]-benzoic acid (t-AUCB: 400 mg/L), the peak concentration of t-AUCB was detected in the skin of diabetic mice, which quickly decreased thereafter. In parallel, 2 h after application of t-AUCB treatment, thermal hyperemia was increased compared to the control gel. Quantification of t-AUCB in plasma of treated animals showed no or low systemic diffusion. Furthermore, haematoxylin and eosin histological staining of skin biopsies showed that skin integrity was preserved in t-AUCB-treated mice. Finally, for pig ear skin, a surrogate for human skin, using Franz diffusion cells, we observed a continuous diffusion of t-AUCB from 2 h after application to beyond 24 h. A single topical administration of a soluble epoxide hydrolase inhibitor improves microcirculatory function in the skin of db/db mice and might represent a new therapeutic approach for preventing the development of skin complications in diabetic patients.

Effect of continuous vs pulsed iontophoresis of treprostinil on skin blood flow.

INTRODUCTION: Systemic sclerosis (SSc) is a rare disease affecting digital microcirculation, leading to finger ulcers and in some cases to amputation. Prostacyclin analogues can be used intravenously but their therapeutic effect is counterbalanced by potentially serious vasodilatation-induced side effects. Iontophoresis of treprostinil could be a promising local therapeutic alternative for SSc-related digital ulcers. Iontophoretic drug delivery is complex, and whether continuous or periodic current should be used remains debated. The objective of the present work is to compare the effect of continuous vs pulsed iontophoresis of treprostinil in rats. MATERIALS AND METHODS: Treprostinil (0.64 mM and 0.064 mM) and NaCl were delivered by cathodal iontophoresis onto the hindquarters of anaesthetized rats. Three protocols delivering the same quantity of current were compared: one was continuous (100 µA during 20 min) and two were periodic (B: twenty 1-min cycles with 200 µA during 30 s followed by 30 s Off; and C: twenty 1-min cycles with 600 µA during 10s followed by 50s Off) (n=8 for each protocol with each concentration). Skin blood flow was quantified using laser Doppler imaging and skin resistance was calculated with Ohm's law. RESULTS: All protocols induced a significant increase in skin blood flow. At the lower concentration (0.064 mM treprostinil) the pulsed 10/50 sequence significantly enhanced cutaneous blood flow (Table 1; Fig. 1B) compared to continuous iontophoresis or the 30/30 sequence. We noted that the pulsed iontophoresis of NaCl (10/50 sequence) induced a significant early increase in cutaneous blood flow in comparison with continuous iontophoresis. Skin resistance measures were negatively correlated with current intensity delivered. CONCLUSION: In conclusion, pulsed iontophoresis of treprostinil with a 10 s/50 s (On/Off) protocol at 600 µA increases the efficacy of iontophoresis at 0.064 mM but not at a tenfold higher concentration. Pulsed iontophoresis could be used to optimize treprostinil iontophoresis, to provide similar efficacy with decreased costs, and should now be tested on humans.