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Fulltext Latex C-serum from Hevea brasiliensis induces non-apoptotic cell death in hepatocellular carcinoma cell line (HepG2)

Lam KL, Yang KL, Sunderasan E, Ong MT

Cell Prolif, 2012 Dec;45(6):577-85.
PMID: 23046445 DOI: 10.1111/j.1365-2184.2012.00841.x

OBJECTIVES: Latex from Hevea brasiliensis (natural rubber tree primarily cultivated for its rubber particles) has no known primary metabolic function, although its biological role is as a plant defence system. The present study has evaluated specific anti-proliferative effects of latex whole C-serum and its subfractions, on human cancer cell lines.
MATERIALS AND METHODS: Cell viability assay using MTT, DNA fragmentation assay and real-time PCR were used to evaluate the cytotoxic effects of latex whole C-serum and its subfractions on the cell lines.
RESULTS: MTT assay revealed very low LC(50) values, 2.0 and 280 ng/ml, for DCS and DCP treatments, respectively. DCS was proven to be more potent compared to DCP, in conferring specific anti-proliferative effects on the cancer cell lines. The study also indicated that anti-proliferative activity of pre-heated C-serum fractions diminished significantly.
CONCLUSION: Although noteworthy cell death was reported, DNA fragmentation assay and real-time PCR confirmed that that induced by latex C-serum subfractions was not promoted via the classical apoptotic signalling pathway.
Allergenic proteins of natural rubber latex

Yeang HY, Arif SA, Yusof F, Sunderasan E

Methods, 2002 May;27(1):32-45.
PMID: 12079415 DOI: 10.1016/S1046-2023(02)00049-X

As the living cytoplasm of laticiferous cells, Hevea brasiliensis latex is a rich blend of organic substances that include a mélange of proteins. A small number of these proteins have given rise to the problem of latex allergy. The salient characteristics of H. brasiliensis latex allergens that are recognized by the International Union of Immunological Societies (IUIS) are reviewed. These are the proteins associated with the rubber particles, the cytosolic C-serum proteins and the B-serum proteins that originate mainly from the lutoids. Procedures for the isolation and purification of latex allergens are discussed, from latex collection in the field to various preparative approaches adopted in the laboratory. As interest in recombinant latex allergens increases, there is a need to validate recombinant proteins to ascertain equivalence with their native counterparts when used in immunological studies, diagnostics, and immunotherapy.
Anti-fungal effect of Hevea brasiliensis latex C-serum on Aspergillus niger

Daruliza KM, Lam KL, Yang KL, Priscilla JT, Sunderasan E, Ong MT

Eur Rev Med Pharmacol Sci, 2011 Sep;15(9):1027-33.
PMID: 22013725

Hevea brasiliensis extract could potentially be employed as a relatively low cost resource for various anti-fungal activities due to the simplicity of latex preparation and the abundance of latex that can be obtained in rubber producing regions. The present study was aimed at examining the species specific anti-fungal property of H. brasilensis latex C-serum against Aspergillus niger.
Anti-Candida albicans activity and brine shrimp lethality test of Hevea brasiliensis latex B-serum

Daruliza KM, Yang KL, Lam KL, Priscilla JT, Sunderasan E, Ong MT

Eur Rev Med Pharmacol Sci, 2011 Oct;15(10):1163-71.
PMID: 22165677

Hevea brasiliensis extracts could potentially be employed as a relatively low cost resource for various anti-fungal activities due to the simplicity of the extract preparation and its abundance especially in the tropical region. Latex B-serum was reported to have anti-cancer property and its specificity in anti-fungal property has not been elucidated. The present study was conducted to determine the anti-fungal activity of Hevea latex B-serum against Candida (C.) albicans (a rounded cell fungus) and Aspergillus (A.) niger (a filamentous fungus).
Morphological and Inter Simple Sequence Repeat (ISSR) markers analyses of Corynespora cassiicola isolates from rubber plantations in Malaysia

Nghia NA, Kadir J, Sunderasan E, Puad Abdullah M, Malik A, Napis S

Mycopathologia, 2008 Oct;166(4):189-201.
PMID: 18568417 DOI: 10.1007/s11046-008-9138-8

Morphological features and Inter Simple Sequence Repeat (ISSR) polymorphism were employed to analyse 21 Corynespora cassiicola isolates obtained from a number of Hevea clones grown in rubber plantations in Malaysia. The C. cassiicola isolates used in this study were collected from several states in Malaysia from 1998 to 2005. The morphology of the isolates was characteristic of that previously described for C. cassiicola. Variations in colony and conidial morphology were observed not only among isolates but also within a single isolate with no inclination to either clonal or geographical origin of the isolates. ISSR analysis delineated the isolates into two distinct clusters. The dendrogram created from UPGMA analysis based on Nei and Li's coefficient (calculated from the binary matrix data of 106 amplified DNA bands generated from 8 ISSR primers) showed that cluster 1 encompasses 12 isolates from the states of Johor and Selangor (this cluster was further split into 2 sub clusters (1A, 1B), sub cluster 1B consists of a unique isolate, CKT05D); while cluster 2 comprises of 9 isolates that were obtained from the other states. Detached leaf assay performed on selected Hevea clones showed that the pathogenicity of representative isolates from cluster 1 (with the exception of CKT05D) resembled that of race 1; and isolates in cluster 2 showed pathogenicity similar to race 2 of the fungus that was previously identified in Malaysia. The isolate CKT05D from sub cluster 1B showed pathogenicity dissimilar to either race 1 or race 2.
The 14.6 kd rubber elongation factor (Hev b 1) and 24 kd (Hev b 3) rubber particle proteins are recognized by IgE from patients with spina bifida and latex allergy

Yeang HY, Cheong KF, Sunderasan E, Hamzah S, Chew NP, Hamid S, et al.

J Allergy Clin Immunol, 1996 Sep;98(3):628-39.
PMID: 8828541 DOI: 10.1016/s0091-6749(96)70097-0

Two major water-insoluble proteins are located on the surface of rubber particles in Hevea brasiliensis latex. A 14.6 kd protein (Hev b 1), found mainly on large rubber particles (> 350 mm in diameter), and a 24 kd protein (Hev b 3), found mainly on small rubber particles (average diameter, 70 nm), are recognized by IgE from patients with spina bifida and latex allergy. Although Hev b 1 (also called the rubber elongation factor [REF]) has previously been reported as a major latex allergen, this conclusion has been disputed on the basis of results from other studies. The allergenicity of Hev b 1 is verified in this study by testing the recombinant protein generated from its gene. Because allergenicity is confined to patients with spina bifida and not observed in adults sensitive to latex, it is not a major latex allergen. The identification of Hev b 3 as another allergen originating from rubber particles is confirmed by immunogold labeling and electron microscopy. Observations with the monoclonal antibody USM/RC2 developed against Hev b 3 show that the protein has a tendency to fragment into several polypeptides of lower molecular weight (from 24 kd to about 5 kd) when stored at -20 degrees C. There is also indication of protein aggregation from the appearance of proteins with molecular weights greater than 24 kd. Fragmentation of Hev b 3 is induced immediately on he addition of latex B-serum, which is normally compartmentalized in the lutoids in fresh latex. In the preparation of ammoniated latex (used for the manufacture of latex products), the lutoids are ruptured, and the released B-serum reacts with Hev b 3 on the rubber particles to give rise to an array of low molecular weight polypeptides that are allergenic to patients with spina bifida.
Molecular cloning and immunoglobulin E reactivity of a natural rubber latex lecithinase homologue, the major allergenic component of Hev b 4

Sunderasan E, Bahari A, Arif SA, Zainal Z, Hamilton RG, Yeang HY

Clin Exp Allergy, 2005 Nov;35(11):1490-5.
PMID: 16297147 DOI: 10.1111/j.1365-2222.2005.02371.x

BACKGROUND:
Hev b 4 is an allergenic natural rubber latex (NRL) protein complex that is reactive in skin prick tests and in vitro immunoassays. On SDS-polyacrylamide gel electrophoresis (SDS-PAGE), Hev b 4 is discerned predominantly at 53-55 kDa together with a 57 kDa minor component previously identified as a cyanogenic glucosidase. Of the 13 NRL allergens recognized by the International Union of Immunological Societies, the 53-55 kDa Hev b 4 major protein is the only candidate that lacks complete cDNA and protein sequence information.

OBJECTIVE:
We sought to clone the transcript encoding the Hev b 4 major protein, and characterize the native protein and its recombinant form in relation to IgE binding.

METHODS:
The 5'/3' rapid amplification of cDNA ends method was employed to obtain the complete cDNA of the Hev b 4 major protein. A recombinant form of the protein was over-expressed in Escherichia coli. The native Hev b 4 major protein was deglycosylated by trifluoromethane sulphonic acid. Western immunoblots of the native, deglycosylated and recombinant proteins were performed using both polyclonal antibodies and sera from latex-allergic patients.

RESULTS:
The cDNA encoding the Hev b 4 major protein was cloned. Its open reading frame matched lecithinases in the conserved domain database and contained 10 predicted glycosylation sites. Detection of glycans on the Hev b 4 lecithinase homologue confirmed it to be a glycoprotein. The deglycosylated lecithinase homologue was discerned at 40 kDa on SDS-PAGE, this being comparable to the 38.53 kDa mass predicted by its cDNA. Deglycosylation of the lecithinase homologue resulted in the loss of IgE recognition, although reactivity to polyclonal rabbit anti-Hev b 4 was retained. IgE from latex-allergic patients also failed to recognize the non-glycosylated E. coli recombinant lecithinase homologue.

CONCLUSION:
The IgE epitopes of the Hev b 4 lecithinase homologue reside mainly in its carbohydrate moiety, which also account for the discrepancy between the observed molecular weight of the protein and the value calculated from its cDNA.