UnC samples were not exposed to UV light and not pre-treated with NanoPCL-M (UnC = 1), T samples were pre-treated with NanoPCL-M before UV stress exposure

UnC samples were not exposed to UV light and not pre-treated with NanoPCL-M (UnC = 1), T samples were pre-treated with NanoPCL-M before UV stress exposure. proliferation of both fibroblasts and SSCs. Our results demonstrate that-M is able to preserve SSCs features and collagen depot after UV-induced senescence, suggesting their capability to retain a young phenotype. L.), an endemic herbaceous plant present in Sardinia and throughout the Mediterranean area [8], is now emerging for its peculiar properties. Myrtle berries, leaves, and brushwood are often noted for their antimicrobial, antioxidant, and anti-inflammatory properties, and for enhancing wound healing process, both in vitro and in vivo [8,47,48,49]. Moreover, the seeds of this plant are rich in hydrolyzable tannins and ellagitannins like galloylquinic acid, monogalloylhexose, ellagic acid hexoside and ellagic acid, Oenothein B, and eugeniflorin D2 [50,51]. We previously demonstrated the PEG6-(CH2CO2H)2 capability of myrtle extracts to counteract the senescence process in adipose derived stem cells [10,52]. We therefore developed a nanodevice combining nanofibers and myrtle extracts from seeds in an attempt to obtain a controlled topical release of specific KPNA3 phytochemicals (NanoPCL-M). We previously described the effect of the treatment with NanoPCL-M against UV-related damage on skin cell populations exposed to UV in a dynamic model within a bioreactor, highlighting the ability of NanoPCL-M to prevent aging in epidermal keratinocytes in a 3D structure, modulating the expression of stemness genes of SSCs [53]. In the present study, we aimed to evaluate the effect of-M, pre-treatment on SSCs and fibroblasts behavior after UV exposure, focusing on extracellular matrix changes. In particular, we investigated the effect of-M on PEG6-(CH2CO2H)2 the molecular events modulating stem cell senescence, including SIRT1 and SIRT2, and HAS2 gene expression. Moreover, we evaluated Collagen I production by SSCs and fibroblasts to gain insights into the consequences of our approach at the level of the extracellular environment. 2. Materials and Methods 2.1. Extract Preparations Myrtle extracts were prepared using the by-products of the industrial preparation of myrtle liqueur as raw material. Seeds were removed from berries, freeze-dried, and powdered to obtain a homogeneous sample. Seven grams of powder were extracted PEG6-(CH2CO2H)2 twice with 60 mL of an ethanol/water solution (70% EtOH). In both extractions, the mixtures were sonicated in an ultrasonic cleaner (VWR International, Leuven, Belgium) for 1 h at 25 C, then centrifuged at 3000 for 10 min. The organic extracts were combined and filtered with Whatman 4 filter paper, evaporated to dryness under nitrogen flow to remove ethanol, then freeze-dried to remove water. 2.2. Nanodevice Fabrication Nanofibers were prepared from polycaprolactone (PCL) polymeric solution through a needleless direct-current electrospinning method. Electrospinning was performed using 10% (= 6) and data are expressed as median standard deviation, assuming a statistically significant value of 0.05 (*). 3. Results 3.1. Nanodevice Characterization The PCL scaffold included polymeric fibers, wherein more than 83 % of all included fibers has diameters on a nanometric scale, called nanofibers, as illustrated in Figure 1. Open in a separate window Figure 1 (a) Fiber diameter distribution of PCL nanofibers. (b) SEM picture of PCL nanofibers. 3.2. NanoPCL-M Allows a Controlled Release of Myrtle Extracts Spectrophotometry revealed a gradual and constant release of the myrtle extracts from the PCL nanofiber device (Figure 2). The release was 0.16 0.05 mg/day for 7 days. Open in a separate window Figure 2 Release of myrtle extracts from NanoPCL-M for 7 days. The amount of extracts was evaluated as absorbance OD detected at 280 nm, and is expressed as mg of extracts/day. 3.3. NanoPCL-M Increases Cell Proliferation The BrdU assay revealed that NanoPCL-M pre-treatment PEG6-(CH2CO2H)2 significantly increased the proliferation of PEG6-(CH2CO2H)2 the analyzed cell populations (Figure 3). The same figure shows that T samples reached a proliferation rate similar to what was observed for UnC, after 3 days of pre-treatment. Moreover, a significantly higher proliferation rate compared with UnC could be observed when cells were pre-treated for 7 days. Open in a separate window Figure 3 Effect of pre-treatment with NanoPCL-M on cell proliferation. HFF1 (a) and SSCs (b) were pre-treated with NanoPCL-M for 3 and 7 days, and then exposed to UV light as described. Cell viability is expressed in OD units as compared with untreated cells UnC. Data are expressed as mean SD referring to the control. * 0.05. 3.4. NanoPCL-M Modulates Sirtuins and.