Elsevier

Journal of Ethnopharmacology

Volume 137, Issue 1, 1 September 2011, Pages 352-358
Journal of Ethnopharmacology

Typha latifolia L. fruit polysaccharides induce the differentiation and stimulate the proliferation of human keratinocytes in vitro

https://doi.org/10.1016/j.jep.2011.05.042Get rights and content

Abstract

Ethnopharmacological relevance

In Northern America Typha latifolia L. (Typhaceae) fruits are used for more than 4000 years for treatment of skin disorders, burns and as wound dressing to absorb the ichors.

Aim of the study

The following studies attempted to characterize water-soluble polysaccharides from aqueous Typha latifolia extracts and to investigate the influence of the polymers on cell physiology of human dermal fibroblasts (NHDF) and epidermal keratinocytes (NHEK).

Materials and methods

Water-soluble raw polysaccharides (RPS) were isolated from Typha latifolia fruits and fractionated by anion exchange chromatography (AEC) and size exclusion chromatography (GPC). Fractions obtained were characterized concerning monosaccharide composition by HPAEC-PAD. The bioactivity of the polysaccharides was investigated on cell viability, proliferation, differentiation and gene expression NHDF of NHEK.

Results

RPS was fractionated into 5 heterodisperse fractions (TL1–TL5). The polysaccharides were composed mainly of glucose (more than 50% in RPS and TL4), galactose, xylose, mannose, glucuronic acid, galacturonic acid, arabinose, ribose, fucose, rhamnose, and fructose with differing amounts concerning to RPS and AEC-fractions. Proteins were detected in the RPS (10%) and to a less extend in TL1–TL3 (1–3%). TL1–TL3 significantly increased the proliferation of keratinocytes, whereas TL4 was shown to be a potent inductor of the early differentiation process of keratinocytes. Gene expression analysis supported these results since Smad3 and PKC-α, known to be part of signal pathways leading to cell differentiation, were significantly up regulated.

Effects on fibroblasts were not observed, indicating cell specific activity of the polysaccharides.

Conclusion

The results clearly indicate a rationale for the traditional use of Typha latifolia fruits extracts for wound healing to the strong stimulatory activity of the polysaccharides on keratinocytes proliferation and early differentiation, major activities necessary for potent wound-healing agents.

Highlights

► Typha latifolia fruits were used for skin treatment. ► We isolated and characterized the water-soluble polysaccharides and tested their bioactivity on human skin cells. ► Five heterodisperse polysaccharide fractions were obtained differing in their monosaccharide composition. ► Effects on fibroblasts were not observed but keratinocyte proliferation was increased by three fractions while the fraction with high beta-D-glucose content induced the differentiation.

Introduction

Typha latifolia L., Typhaceae (bulrush, cattail or reed mace) is a commelinid monocotyledon. In Northern America the fruits (achene and perigone hairs) were used as wound dressing (Moermann, 1998) to absorb the ichors. In the Dr. Dukes Phytochemical and Ethnobotanical Database Typha latifolia is one plant pronounced to be traditionally used for wound healing (Duke, 2006). Medicinal use is also reported for roots and the sticky juice between young leaves (Carman, 2006). The plant is herbaceous, rhizomatous and perennial with long sword-like leaves from the bare stalk which terminates in a cylindrical inflorescence of female flowers immediately below the male flowers, erupting as fluff when matured. The habitat includes shallow water of lake and pond edges, marshes, and ditches at elevations lower than 2000 m. Phytochemical analysis of Typha latifolia was done so far on leaves, roots, rhizomes and pollen. Alkanolamides (Peru et al., 2004) and flavonol glucosides (Sick Woo et al., 1983) were found in leaves. Contents of carbohydrates and sugars were investigated in overwintering parts (Kausch et al., 1981, Biesboer, 1984) and the stem (Pollard, 1982). Carotinoid-like compounds and allelopathic sterols of the whole plants were also characterized (Dell Greca et al., 1990a, Dell Greca et al., 1990b) as the lipid fraction of Typha latifolia pollen (Caffrey et al., 1987).

Polysaccharides, tannins and glycosides are the most plausible phytochemicals that would be extracted in a wet wound atmosphere from plant derived wound dressings. A direct influence on the cell physiology of epidermal and connective tissue cell is possible since such effects have been described for other carbohydrates like alginates (Edwards et al., 2003), glycosaminoglycans (Kirker et al., 2002) and β-glucan (Delatte et al., 2001). Recent reports also indicate that such polysaccharide-mediated effects are strongly dependent on the structural features of the polymers. (Kirker et al., 2002, Ni et al., 2004).

Wound healing is a dynamic and complex regenerative process in which different cell species of the skin and the immune system working together in three distinct phases: the inflammatory phase, the reepitheliazation phase and the remodeling phase. During reepitheliazation keratinocyte proliferate at the wound edge, consequently differentiate to corneocytes and form a thin new epidermis. This process is followed by migration of fibroblasts to the wound edge subsequently proliferating and producing extracellular matrix. During the remodeling phase fibroblasts transform into myofibroblasts and deposit collagen (Werner and Grose, 2003). While each cell type has its own function their cross talk by growth factors and cytokines is essential for wound closure (Werner et al., 2007). The wound healing process is mostly investigated by in vivo models but for ethical and financial reasons these models are for screening usually changed to in vitro bioassays. The established in vitro cell culture based models cover the different wound healing processes like proliferation, metabolic activity and differentiation of human skin fibroblasts and keratinocytes.

The aim of this study was to isolate the polysaccharides of an aqueous extract of Typha latifolia fruit and to investigate their effect on human skin cells.

Section snippets

General

All chemicals used were of analytical quality and were purchased from Diagonal (Münster, Germany). Cell culture media supplements are from PAA (Coelbe, Germany) as not mentioned otherwise.

Isolation, fractionation and characterization of polysaccharide

Typha latifolia L. fruits were collected from a private pond in the north western part of Germany (Münster, Westfalen) in late autumn 2006. A reference specimen (TL561/D) is placed at the Institute of Pharmaceutical Biology and Phytochemistry in Münster. The fruits (215 g), including the trichomes were grinded

Polysaccharides from Typha latifolia fruits

Water-soluble polysaccharides were extracted from the fruits by aqueous extraction at room temperature, followed by ethanol precipitation of the raw polysaccharides (RPS) at a 1% yield and containing about 10% of carbohydrates, beside residual polyphenolic compounds and proteins. RPS was separated into five subfractions by anion exchange chromatography using a step gradient with increasing ionic strength of buffer. The main fraction TL3 was obtained at a buffer concentration of 0.25 mol/L, while

Discussion

The activity of the tested polysaccharides on human skin cells showed that the impure RPS reduced significantly cell proliferation and enzymatic activity pointing to a pro-apoptotic potential since the test for necrotic activity was negative. This effect is maybe reduced to the impurities, since it is known that some phenols are cytotoxic or pro-apoptotic and because of this effect was not shown in case of the AEC fractions (Ugartondo et al., 2007, Koppikar et al., 2010). Contrary to TL4, which

Conclusions

Regarding these results Typha latifolia fruit polysaccharides have even though less effects on reepitheliazation and remodeling, two phases of the wound healing process. In particular more information must be collected to find out the signal pathway responsible for the cells reactivity on these different polysaccharides and their structure.

Acknowledgements

The authors thank Cathleen Possemeyer for technical assistance and Dr. Lohse, Department of Paediatric Surgery, University of Münster for support with skin grafts. Financial support for the study was given by A. Hensel.

References (43)

  • Y. Ni et al.

    Isolation and characterization of structural components of Aloe vera L. leaf pulp

    International Immunopharmacology

    (2004)
  • P. Owens et al.

    The role of Smads in skin development

    Journal of Investigative Dermatology

    (2008)
  • S. Pastore et al.

    The epidermal growth factor receptor system in skin repair and inflammation

    Journal of Investigative Dermatology

    (2008)
  • C.J. Pollard

    Fructose oligosaccharides in monocotyledons: a possible delimitation of the order Uliales

    Biochemical Systematics and Ecology

    (1982)
  • T. Porstmann et al.

    Quantification of 5-bromo-2′-deoxyuridine into DNA: an enzyme immunoassay for the assessment of the lymphoid cell proliferative response

    Journal of Immunological Methods

    (1985)
  • W. Sick Woo et al.

    A flavonol glucoside from Typha latifolia

    Phytochemistry

    (1983)
  • B.G. Smith et al.

    The polysaccharide composition of Poales cell walls: Poaceae cell walls are not unique

    Biochemical Systematics and Ecology

    (1999)
  • B.K. Steele et al.

    Variable expression of some “housekeeping” genes during human keratinocyte differentiation

    Analytical Biochemistry

    (2002)
  • S. Werner et al.

    Keratinocyte–fibroblast interactions in wound healing

    Journal of Investigative Dermatology

    (2007)
  • M.B. Yaffe et al.

    Biophysical characterization of involucrin reveals a molecule ideally suited to function as an intermolecular cross-bridge of the keratinocyte cornified envelope

    Journal of Biological Chemistry

    (1992)
  • J. Zippel et al.

    A high molecular arabinogalactan from Ribes nigrum L.: influence on cell physiology of human skin fibroblasts and keratinocytes and internalization into cells via endosomal transport

    Carbohydrate Research

    (2009)
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