Characterization of Normal Tissues and Metastatic Adenocarcinoma of the Lungs using Lectin Histochemistry
Ourlad Alzeus G. Tantengco
Institute of Biology, College of Science, University of the Philippines Diliman, Quezon City, Philippines, 1101
ABSTRACT Lectin Histochemistry shows that lectin can be utilized as specific markers for cell population. Several lectins were studied in this paper including Helix pomatia agglutinin (HPA), which is used as primary marker-based predictor of prognosis on various adenocarcinoma cells. It was observed that expression of HPA, PHA-L and UEA-I are strongly associated with metastatic spread in several cells of the primary tumor and to their lymph node metastases. Detection of lectin binding using the simple avidin-biotin method
is advantageous because of its availability, speed, suitability for Western blotting.
By the virtue of studying the pattern of glycosylation and glycoprofiles of cells and tissues, better understanding regarding disease mechanisms can be achieved. Disruption of cellular
glyscosylation mechanisms can result in changes in the extension of oligosaccharide chains; this is associated with tumor progression and metastasis. INTRODUCTION The respiratory system which macrophages, alveolar epithelial cells
(pneumocytes), mast cells and the capillary network. Observation of migratory cell population or changes in the normal number of resident cell population is vital in understanding the dynamics of various diseases inside the organ system.
includes the lungs is a branching system of tubes that link the site of gas exchange with the external environment. It provides a conduit to ensure uninterrupted supply of air but it also serves as passageway for toxins, chemical substances and pathogens.
(Barkhordari et. al., 2004) The study of glycan in cells and tissues is attracting greater attention because of its use for characterization of cell population. By the virtue of studying the
Understanding the normal histology of the lungs allow us to observe the structures and function of the resident cell populations of pulmonary tissues such as alveolar
This coat is called glycocalyx and is involved in cell to cell and cell to matrix interaction which is important in tumor metastasis. Lectin Histochemistry shows that lectin can be utilized as specific markers for cell
adenocarcinoma of the lung is rising. 2004). al.2
pattern of glycosylation and glycoprofiles of cells and tissues.. Sambucus nigra (SNA) and Maackia amurensis (MAA) also
prognostic markers. it was improved with
immunohistology of the cells (Barkhordari et.
glycosylation. The worldwide incidence of
sequences of certain cell population. Changes in the glycosylation of this carbohydrate rich coat are associated with metastatic potential of tumors and therefore have potential as markers for lung cancer prognosis. Lectins used in immunohistology of the normal lungs are as follows: Galanthus nivalis (GNA) and (NPA) for Narcissus alveolar esculentum
pseudonarcissus macrophages.. 2002). A carbohydrate rich coat covers the outer surface of mammalian cells. al. Phytolacca americana (PAA) and Triticum vulgaris-succinylated (sWGA) for type I pneumocytes. 2007) The aim of current researches is to analyze the glycoconjugate expression of normal lung and adenocarcinomas of the lung by using several lectins to define a prognostic marker for cancer.
(LEA). (Thom et. Lectins.. are used in analysis of glycans. The classification of primary lung carcinomas is probably one of the least precise among all the solid tumors (Laack et. but patchy staining by Arachis hypogaea (AHA). Since lectins can detect protein these changes in
comprehensive lectin-binding analysis of adenocarcinomas undertaken to of the lung possible was new
Phaseolus vulgaris leukoagglutinin (l-PHA) and Anguilla anguilla (AAA) for endothelial cells and Dolichos biflorus (DBA) for bronchial epithelium. al. a group of specific glycoproteins present in animal as well as plant cells. The universal staining of type I pneumocytes by LEA.
. better understanding regarding disease mechanisms can be Immunohistology of Normal Lungs Lectins can be used as reliable markers for identification of the cells and studying subtle differences in sugar
achieved. PAA and sWGA. Hippeastrum hybrid (HHA) and MPA for type II pneumocytes. Psophocarpus tetragonolobus (PTL-II). In order to achieve better characterization of cell populations using lectins.
The advantage of this kind of method is its high specificity in labeling therefore eliminating possible problems concerning sterical intereference of reporter molecule with carbohydrate combining site of lectins.3
indicated not only the presence of an abundant set of the related glycoforms in these cells in normal lung. but also the possibility of two distinct populations or a pattern of differentiation which is
squamous-cell carcinoma and its regional lymph node metastases. Lectin Immunohistology of normal lungs helps in decoding the patterns of glycosylation of cells which is helpful in understanding the mechanisms of diseases such as metastasis of adenocarcinoma.
complex multi-layered method to detect lectin binding specifically for HPA. thyroid and prostate. colorectum. esophagus.. 2003). ovary. Furthermore.
Labelling Methods for Detection of Lectin Binding Current studies have used the
Labeled Lectins for Histochemical Detection of Metastasis Helix pomatia agglutinin (HPA) binding was the primary marker-based predictor of prognosis cells such on as various breast. The disadvantage of this method is the
leucoagglutinin (PHA-L) has been reported by several studies as an effective marker that detects the tumor development in breast and colon cancer. the lectin Ulex europaeus agglutinin-I (UEA-I) was investigated in breast cancer where its binding related to disease-free interval and survival. Phaseolus vulgaris
incorporation of horseradish peroxidase labeled avidin or streptavidin (Brooks & Wilkinson. It has also been established as an excellent marker for human endothelial cells (Laack et. There is also a significant correlation between the expression of PHA-
morphologically unapparent (Barkhordari et. The tissue specimen is incubated with
unconjugated HPA which recognizes the GalNAc residues on cellular glycoproteins. However. then a biotin labeled secondary anti-rabbit antibody and the final layering is through
stomach. al. al.. 2004). HPA has been used to define the metastatic phenotype of these tumors. Multiple layering starts with a polyclonal rabbit antibody directed against HPA. 2002). HPA has not been the only lectin that has provided useful prognostic information.
PHA-L and UEA-I are strongly associated with metastatic spread in several cells of the primary tumor and to their lymph node metastases. Disruption of cellular glyscosylation mechanisms can result in changes in the extension of oligosaccharide chains. Due to these disadvantages.
Adenocarcinoma Lung The classical way of assessing the tumor spread is through anatomical
application of multivariate analysis only HPA binding remained as a significant independent prognostic indicator. 2003) It was observed that expression of HPA. Several researches about adenocarcinoma other than lung. Using univariate analysis. VAA showed approximately
uncommon terminal residue in the glycans
. Recent studies show 159 cases of lung cancer that were investigated for their capacity to bind Viscum album L.
However. (Brooks & Wilkinson. This classical TNM method is disadvantageous because this will not give any understanding of the metastatic capability of the tumor. an Lung Cancer-Specific Lectin According to current researches
reagarding lectin Histochemistry of the lungs. specific lectins for lung cancer cases are not yet worked out widely.
removed. (VAA) and galectin-1 and for Lewis antigen reactivity.
researchers resorted to a simpler detection method using commercially widely-
available reagents such as simple avidinbiotin method. this is associated with tumor progression and metastasis. VVA has a high affinity for the disaccharide GalNAcα1 and 3Galβ1-. the lectin from the Roman snail. HPA recognizes oligosaccharides with a terminal α-glycosidically-linked Nacetylgalactosamine (α-GalNAc). Biotin-labeled HPA detected with an advitin-biotin complex (ABC) method has been used as prognostic and biological marker in breast cancer and animal models of human melanoma. Helix pomatia agglutinin (HPA). This method is also unsuitable for Western blotting because the reagents are cross reactive with
of normal cells.4
availability of the polyclonal antibody and the length of time consumed. has been used to define the metastatic phenotype of these tumors. it was found out that these lectins bind to the primary tumors and therefore can be used as prognostic markers for overall and relapse-free survival after the tumor was
antibodies used in multiple layering.
description of tumor.
(VAA) and galectin-1 also provided useful
adenocarcinomas has not been elucidated completely. There is no direct correlation between the presence on VAA binding capacity and survival whereas better
prognosis has been observed due to the presence of the galectin-1 binding sites (Sherwani et. as it is associated with the presence of metastases in lymph nodes.
CONCLUSION Lectins are used in characterization of the glycoconjugate content of normal and malignant tissues of the lungs. and.. the molecular basis why HPA binds preferentially to metastasizing
development. Galectins are a family of proteins that includes calcium independent galactoside binding lectins. Viscum album
L. Several methods of labeling lectin binding were also presented here including the multilayering method and the use of simple avidin-biotin method for detection of HPA binding..
. Avidin-biotin method is simple and advantageous because it does not require access to polyclonal antibodies against the lectin. Expression of several lectins
carcinomas. whose functional role has to be elucidated in future studies. The precise structure of the
HPA binding oligosaccharide associated with metastasis has not been identified yet. 2003). 84% of the lung tumours showed HPA binding (Laack et. However. al. PHA-L and UEA-I are strongly associated with metastatic spread in several cells of the primary tumor and to their lymph node metastases. Adenocarcinomas and
detectable by the lectin HPA and other lectins. is one of the most frequently reported glycosylation changes in
branchioalveolar carcinomas were more frequently labelled than squamous
70% moderate to strong binding to lung tumour. Deviation from the lectin labelling and expression present in the normal tissues is also one of the changes in malignancy. 2002). as
The use of lectin Histochemistry is of prognostic significance in many human cancer types. a monosialylated oligosaccharide of 4. upon application of multivariate analysis only HPA binding remained as a significant independent prognostic indicator for adenocarcinoma of the lungs.
including HPA. The aberrant expression of GalNAc oligosaccharides. al.58 glucose units termed HPA gly-1 has been identified in HPA-positive breast cancers.
160. Eckart et. 3. Validation of a simple avidin-biotin detection method for Helix pomatia lectin (HPA) binding as a prognostic marker in cancer.
Journal of Molecular Histology 35: 147–156. al. American Journal of Pathology. Asma Farhat et. Susan A & Diane Wilkinson. Netherlands. Acta Histochemica. Lectin Lung. Pages 391–397 Sherwani. al. Lectin Histochemistry of Resected Adenocarcinoma of the Lung. Ina et. 2003. al. 18 (2) 169-180
. Vol. Kluwer Academic Publishers. March 2002
Thom. Abolfazl of et. Issue 3. Lectin histochemistry of metastatic adenocarcinomas of the lung. 2004
Brooks. Lung Cancer Volume 56.
Characterization of Lectins and Their Specificity in Carcinomas An
REFERENCES Barkhordari. Indian Journal of Clinical Biochemistry. No. June 2007. 105(3) 205–212 (2003)