In vitro studies on the biosynthesis of allergens of parhenium hysterophorus and cytotoxicity of parthenin

Abstract

Parthenium hysterophorus, like several other members of the Compositae, causes allergic contact dermatitis and hay-fever in humans. Besides this, it is also ecologically dominant and is toxic to livestock. The allergic contact dermatitis and toxicity have been attributed to the presence of sesquiterpene lactones, and the hay-fever to allergenic proteins present in the pollen. The thesis entitled “In vitro studies on the biosynthesis of allergens of Parthenium hysterophorus and cytotoxicity of parthenin” is divided into six chapters.

Chapter 1 provides an introduction to the biologically active principles of the Compositae, and the use of tissue cultures as model systems to study morphogenesis, biosynthesis, and production of natural compounds.

In Chapter 2, the results on the establishment of tissue cultures from P. hysterophorus and its morphogenesis are presented. Callus cultures were obtained from leaf, stem, flower, and roots of P. hysterophorus. The leaf formed a friable and rapidly growing callus. Both root and shoot differentiation were obtained with the stem callus, while the leaf callus failed to regenerate any shoots. The leaf callus, when cultured in liquid medium, formed rapidly growing cell suspension cultures. Roots obtained from the leaf callus were cultured in liquid medium.

Chapter 3 describes the studies on the biosynthesis of sesquiterpene lactones in the tissue cultures. The culture extracts elicited a positive patch test response in patients showing allergic contact dermatitis to P. hysterophorus. A new High Performance Liquid Chromatography (HPLC) method was standardized to study variations in the levels of the sesquiterpene lactones during the various stages of differentiation. The rapid separation of the sesquiterpene lactones could be carried out by HPLC, and the major sesquiterpene lactone, parthenin, was quantitated. In the stem callus, shoot differentiation enhanced the levels of parthenin, though among the cultures, the unorganized leaf cell suspension cultures had the highest amount of parthenin. The sesquiterpene lactone pattern in the various plant parts and the cultures was studied.

In Chapter 4, studies on the protein allergens produced by the cell suspension cultures of P. hysterophorus are described. The suspension cultures produced protein antigens that were similar to the protein antigens in the pollen. These antigens were able to induce IgG and IgE in experimental animals. The major allergenic protein present in pollen was detectable in cultures, was isolated, and found to have a molecular weight of around 42,000 daltons. The culture extracts gave a positive prick-test response in patients suffering from allergic rhinitis who also gave a positive response to pollen extracts.

Several of the sesquiterpene lactones are not only allergenic but are also cytotoxic. The effects of parthenin, the major sesquiterpene lactone of P. hysterophorus, on cellular metabolism are presented in Chapter 5. Parthenin was cytotoxic to and inhibited macromolecular synthesis in mammalian cell cultures. Key cellular enzymes were also inhibited, and cAMP levels were enhanced on drug treatment. Cultured lymphocytes, on treatment with parthenin, showed a rise in the number of cells arrested at the metaphase stage of mitosis. Tetrahydroparthenin, in which the reactive groups—the -methylene and the unsaturated cyclopentenone system of parthenin—were reduced, showed no activity.

Chapter 6 summarizes the main results and conclusions of the investigation.