RESUMEN
BACKGROUND: Wound healing is an active, complex, integrated series of cellular, physiological, and biochemical changes initiated by the stimulus of injury in a tissue. The present study was performed to investigate the potential wound healing abilities of Sargassum ilicifolium crude extracts (CE) that were characterized by 1H NMR and FTIR Spectrometric measurements. MATERIALS AND METHODS: Seaweed samples were collected from southern coastal sites of Sri Lanka. To determine the cytotoxicity and proliferation of S. ilicifolium CE were used for the MTT and alamarBlue assays respectively. The scratch and exclusion wound models were used to HaCaT and HDF cells to assess the cell proliferation and migration. RAW 264.7 cells (macrophages) were used to evaluate Nitric Oxide (NO) production and phagocytosis activities. Moreover, Fifteen, 8-week-old, female, New Zealand rabbits were selected and divided into five groups: excision skin wounds (10.40 ± 0.60 mm) were induced in groups I, II, and III. Rabbits in groups I and IV were given S. ilicifolium CE (orally, 100 mg/kg day, two weeks), whereas groups II and V were given equal amounts of distilled water. Wound healing properties were measured and wound tissue samples were collated, formalin-fixed, wax-embedded, stained (Hematoxylin and Eosin; Van Gieson) and examined for the healing process. RESULTS: Anti-inflammatory and wound healing activities were observed in RAW 264.7, HDF and HaCaT cells treated with S. ilicifolium aqueous extracts when compared to the control groups. S. ilicifolium extracts concentration 8 - 4 µg/µL, (P<0.05) had remarkable the highest proliferative and migratory effects on RAW 264.7, HDF and HaCaT cells when compared with the control. RAW 264.7 cell proliferation and/or migration were higher in S. ilicifolium extracts (4 µg/µL, 232.8 ± 10.07%) compared with the control (100 %). Scratch wound healing were remarkably enhanced in 24 h, 48 h (P<0.05) when treated with S. ilicifolium on HaCaT cells. Rabbits treated with the CE of S. ilicifolium showed a significantly increased wound healing activities (P<0.05) within three days with a close wound area of 57.21 ± 0.77 % compared with control group (26.63 ± 1.09 %). Histopathology, aspartate aminotransferase and alanine aminotransferase levels evidenced no toxic effects on seaweed treated groups. Histopathological results also revealed that the healing process was significantly faster in the rabbit groups which were as treated with CE of S. ilicifolium orally with the evidence of enhanced early granulation tissue (connective tissue and angiogenesis) and significant epithelization compared to the control. CONCLUSIONS: Cell proliferation and migration are significantly faster when treated with S. ilicifolium aqueous extracts. Moreover, there are no toxic effect of S. ilicifolium aqueous extracts on RAW 264.7, HDF and HaCaT cell lines. In this study, it is revealed that S. ilicifolium has potential remedial agent; D-Mannitol for skin wound healing properties that by promote keratinocyte and fibroblast proliferation and migration. These findings show that S. ilicifolium have promising wound healing properties.
Asunto(s)
Sargassum , Femenino , Conejos , Animales , Cicatrización de Heridas , Aspartato Aminotransferasas , Bioensayo , Proliferación CelularRESUMEN
Seaweeds have been regarded as a reservoir of biologically active molecules that are important in the pharmaceutical industry. The aim of the present study was to explore the wound healing properties and to assess the safety of the seaweed Sargassum ilicifolium and Ulva lactuca. Enhanced cell proliferation and cell migration activities were observed in L929 cells treated with S. ilicifolium extract compared to U. lactuca extract treated cells and the control group. In-vivo experiments were conducted using five groups (10 in each) of Albino mice (BALB/c). Mice in group I and group II were treated (Orally, 100 mg/kg BW/day) with aqueous extracts of S. ilicifolium and U. lactuca, respectively for 14 days. Treatment group III received a topical application of the aqueous extract of S. ilicifolium (25% w/w) and ointment base (75% w/w) (2 g/kg BW/day, for 14 days). Group IV (Control) received an equal amount of distilled water, orally and mice in group V kept without wounds. The extract from S. ilicifolium showed stronger wound healing properties than the one from Ulva lactuca. Histopathological findings also revealed that the healing process was significantly enhanced in the mice group treated orally with S. ilicifolium aqueous extract. These findings show that S. ilicifolium species possess promising wound healing properties in-vitro and in-vivo.
RESUMEN
BACKGROUND: Seaweeds are an important source of bioactive compounds which are applied in various aspects of medicinal investigations. The present study was conducted to investigate cytoxicity (in-vitro and in-vivo) and wound healing activity of different seaweed species in Sri Lanka. METHODS: Twenty-three seaweed samples, belonging to Phaeophyta (Brown), Chlorophyta (Green) and Rhodophyta (Red) were used for the experiments. Samples were collected from the inter-tidal and the sub-tidal habitats around Sri Lankan coast (Southern, Northern and North-western). Aqueous seaweed extracts were tested for cytotoxic and wound healing activity; in-vitro and in-vivo. To determine toxicity of aqueous seaweed extracts, brine shrimp lethality assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) colorimetric assay on mouse fibroblasts (L929) cell line were performed. Cell migration induction of seaweed extracts was assessed by scratch wound healing assay using L929 cell line. Based on the our previous experiments S.ilicifolium (SW23) was selected for the in vivo study to confirm our hypothesis. Albino mice (BALB/c) were divided into three groups (12 in each) and a circular area (44.07 ± 02.51 mm2) of full skin was excised to create a wound in mice group II and III. Group III received aqueous extract of Sargasum illicifolium (400 mg/kg BW/day for 12 days, orally), Group II received distilled water for 12 days whereas Group I was used as the control group and it was tested without forming wounds and without providing any treatment. Further, the expression level of Tumor Necrosis Factor (TNF-α) and Transforming Growth Factor-ß (TGF-ß) via RT-PCR were measured every three days until the end of the experiment. RESULTS: Phytochemical tests showed positive results to flavonoids in all the selected green seaweeds and alkaloids were observed in red seaweeds. In the toxicity assay, red seaweed, Acanthophora spicifera (SW17) was found to be highly effective on nauplii of brine shrimp (LC50 = 0.072 µg/µl). LC50 value of green seaweed species, Caulerpa racemosa (SW02 and SW08) and Caulerpa sertularioides (SW10) was not found within the tested concentration series. The highest cytotoxic effect on L929 cell line was exhibited by aqueous extracts of red seaweed; Jania adhaereus with 50.70 ± 7.304% cell viability compared with control group. The highest cell migration activity was observed in L929 cell line group treated with extracts of green seaweed namely; Halimeda opuntin (SW07) and extracts of brown seaweed namely; Stoechospermum polypodioides (SW11). Extracts of S. illicifolium (SW23) exhibited a significantly enhanced wound healing activity in mice group III within three days (P < 0.05) with an open wound area of 17.35 ± 1.94 mm2 compared with control group (26.29 ± 2.42 mm2). TGF-ß gene expression peaked on 6th day of post-wound and subsequently decreased on 9th day of post-wound in mice group III. TNF-α expression was suppressed in mice group III whereas it was elevated in group II. TGF-ß expression is enhanced in the treatment group compared to the control group. CONCLUSIONS: Aqueous extracts of selected seaweeds are a significant source of potential compounds with wound healing properties, which might be helpful in the healing of various wounds. This also infers that many species of brown and red seaweeds have the potential of wound healing, specifically, Sargasum illicifolium and Jania adhaereus could be a potential candidate for in-vivo studies related to wound healing and cancer therapy in the near future.
RESUMEN
We have previously demonstrated a correlation between clinical paroxysms in Plasmodium vivax malarial infections and the appearance in patients' plasma of factors that kill blood stage parasites (gametocytes). This activity was, as previously shown, dependent on the presence in paroxysm plasma of tumour necrosis factor-alpha (TNF-alpha), which acts in conjunction with other 'complementary' factors. Here we have identified a parasite component which is essential for this activity and functions as a 'complementary' factor together with TNF, and a third component of unknown origin. The P. vivax parasite component present in paroxysm plasma can be substituted with a blood-stage schizont extract of either P. vivax or P. falciparum. This was demonstrated by restoring the parasite-killing activity to post-paroxysm plasma (from which it was absent) with the addition of the extracts together with TNF. The active materials in these extracts, however, are different from the natural components in P. vivax paroxysm plasma, i.e. while the schizont extracts are immunologically cross-reactive between species, the activity of the natural P. vivax toxin(s) in patients' plasma is neutralized only by the homologous antisera. Plasmodium falciparum infections have neither distinct paroxysms nor parasite-killing activity in plasma. The pronounced paroxysms of P. vivax infections may thus be due in part of a species-specific toxin(s).