LED treatment of ischemia challenged tissue improved early wound healing by enhancing angiogenesis irrespective of the wavelength thus delineating this noninvasive means as a potential, cost effective tool in complicated wounds.

Low level light therapy receives increasing interest in the fields of tissue regeneration and wound healing. Several in vivo studies demonstrated the positive effects of LLLT on angiogenesis. This study aimed to investigate the underlying properties in vitro by comparing the effects of light therapy by light emitting diodes of different wavelengths on endothelial cells in vitro. Human umbilical vein endothelial cells were treated with either 475 nm, 516 nm or 635 nm light. Control cells were not illuminated. 2D proliferation was quantified by manual counting. HUVEC migration was analyzed by performing a 2D wound scratch assay and a 3D bead assay. The influence of LLLT on early vasculogenic events was determined in a 3D fibrin co-culture model with adipose-derived stem cells. Stimulation with both red and green pulsed LED light significantly increased HUVEC proliferation and 3D migration. Moreover, HUVEC showed increased 2D migration potential with green light stimulation. The treatment with blue light was ineffective. Several parameters showed that green light was even more potent to stimulate proliferation and migration of endothelial cells than clinically well-established red light therapy. Further studies have to focus on intracellular mechanisms induced by different wavelengths in order to optimize this promising therapy in tissue regeneration.

Extracorporeal shock waves (ESWTs) are “mechanical” waves, widely used in regenerative medicine, including soft tissue wound repair. Although already being used in the clinical practice, the mechanism of action underlying their biological activities is still not fully understood. In the present paper we tried to elucidate whether a proinflammatory effect may contribute to the regenerative potential of shock waves treatment. For this purpose, we exposed human foreskin fibroblasts (HFF1 cells) to an ESWT treatment (100 pulses using energy flux densities of 0.19 mJ/mm2 at 3 Hz), followed by cell analyses after 5 min, up to 48 h. We then evaluated cell proliferation, reactive oxygen species generation, ATP release, and cytokine production. Cells cultured in the presence of lipopolysaccharide (LPS), to induce inflammation, were used as a positive control, indicating that LPS-mediated induction of a proinflammatory pattern in HFF1 increased their proliferation. Here, we provide evidence that ESWTs affected fibroblast proliferation through the overexpression of selected cytokines involved in the establishment of a proinflammatory program, superimposable to what was observed in LPS-treated cells. The possibility that inflammatory circuits can be modulated by ESWT mechanotransduction may disclose novel hypothesis on their biological underpinning and expand the fields of their biomedical application.

A highly interesting source for adult stem cells is adipose tissue, from which the stromal vascular fraction (SVF)-a heterogeneous cell population including the adipose-derived stromal/stem cells-can be obtained. To enhance the regenerative potential of freshly isolated SVF cells, low-level light therapy (LLLT) was used. The effects of pulsed blue (475 nm), green (516 nm), and red (635 nm) light from light-emitting diodes applied on freshly isolated SVF were analysed regarding cell phenotype, cell number, viability, adenosine triphosphate content, cytotoxicity, and proliferation but also osteogenic, adipogenic, and proangiogenic differentiation potential. The colony-forming unit fibroblast assay revealed a significantly increased colony size after LLLT with red light compared with untreated cells, whereas the frequency of colony-forming cells was not affected. LLLT with green and red light resulted in a stronger capacity to form vascular tubes by SVF when cultured within 3D fibrin matrices compared with untreated cells, which was corroborated by increased number and length of the single tubes and a significantly higher concentration of vascular endothelial growth factor. Our study showed beneficial effects after LLLT on the vascularization potential and proliferation capacity of SVF cells. Therefore, LLLT using pulsed light-emitting diode light might represent a new approach for activation of freshly isolated SVF cells for direct clinical application.

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Melting points were determined on a Mel-Temp capillary melting points apparatus, model 1001, and are uncorrected. Elemental (C, H, N, S) analysis of the samples was carried out in the Center for Instrumental Analysis, Faculty of Chemistry, Belgrade. IR spectra were obtained on a Perkin Elmer Spectrum One FT-IR spectrometer with a KBr disc. 1 H and 13 C-NMR spectra were recorded on a Varian Gemini 200 MHz spectrometer. UV-Vis spectra were recorded on a double beam UV-Vis spectrophotometer model Cary 300 (Agilent Technologies, Santa Clara, USA) with 1.0 cm quartz cells. General procedure for the synthesis of 4a-tTo the aqueous solution of NaOH (21.887 mmol, 3.8 mL of distilled H2O) monochloroacetic acid (10.94 mmol, 1033.83mg) and vanillin (9.48 mmol, 1442.38 mg) were added, and the reaction mixture was refluxed for 100 minutes. Afterwards, the mixture in the flask was cooled to room temperature, 15 mL of distilled H2O was added and the solution was acidified with 2M HCl to pH 2, whereby a beige precipitate was formed. The suspension was cooled in the refrigerator for 2 h, the precipitate was filtrated, washed with a small amount of cold water and dried over anhydrous CaCl2. The crude product was purified by dissolving in hot distilled water. After cooling in a refrigerator for 2 h, brown-yellow crystals were formed. The compound was filtered off, dried over anhydrous CaCl2 and used for further synthesis. In the next step, the compound 2 (2.857 mmol, 600.49 mg) was measured into a 50 mL flask, followed by the addition of dry, freshly distilled CH2Cl2 (14.28 mL), SOCl2 (28.57 mmol, 2.07 mL) and 4 drops of DMF. After 12 h of stirring at room temperature, the solvent was evaporated under reduced pressure, and the excess of SOCl2 was removed by azeotropic distillation with toluene.Then, a corresponding amine (2.857 mmol), NaHCO3 (5.714 mmol, 479.85 mg) and dry CH2Cl2 (14.28 mL) were added to the formed acid chloride, without its previous isolation, and the reaction mixture was further stirred at room temperature for additional 4.5 h. After completion, CH2Cl2 was evaporated under reduced pressure, 28.60 mL of distilled water was added to the residue, and the pH was adjusted to 10 by addition of solid Na2CO3. In the case of compounds S4 4b, 4i and 4t this suspension was left overnight at room temperature, vigorously stirred for 20 minutes the next day and then filtered. In all other cases after pH adjustment, the suspension was stirred for 20 minutes and filtered. With exception of 4o, which was obtained with satisfactory purity and used for further synthesis without purification, all other 4a-t derivatives were purified by dissolving in an appropriate solution of hot aqueous ethanol, and left overnight in the refrigerator. Compounds 4a-g, 4j, 4l, 4n, 4r and 4s-t were recrystallized from 50% aqueous solution of EtOH, 4i from 55%, 4m and 4p from 60% EtOH, while 4h and 4k were purified by recrystallization from 65% EtOH. Derivatives 4a-t were obtained in satisfactory yields ranging from 42 to 78%. General procedure for the...

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