Effect of Induced Mechanical Leaf Damage on the Yield and Content of Bioactive Molecules in Leaves and Seeds of Tepary Beans (Phaseolus acutifolius).

Growing interest has recently been shown in Tepary beans (Phaseolus acutifolius) because they contain lectins and protease inhibitors that have been shown to have a specific cytotoxic effect on human cancer cells. Bean lectins offer protection against biotic and abiotic stress factors, so it is possible that mechanical foliar damage may increase lectin production. This study evaluates the effect of mechanical stress (foliar damage) on lectin and protease inhibitor content in Tepary beans. Seed yield was also analyzed, and phenolic content and antioxidant capacity (DPPH and TEAC) were determined in the leaves. An experimental design with random blocks of three treatments (T1: control group, T2: 50% mechanical foliar damage and T3: 80% mechanical foliar damage) was carried out. Mechanical foliar damage increased the amount of lectin binding units (LBUs) fivefold (from 1280 to 6542 LBUs in T3) but did not affect units of enzymatic activity (UEA) against trypsin (from 60.8 to 51 UEA in T3). Results show that controlled mechanical foliar damage could be used to induce overexpression of lectins in the seeds of Tepary beans. Mechanical foliar damage reduced seed production (-14.6%: from 1890 g to 1615 g in T3) and did not significantly increase phenolic compound levels in leaves.

Quantum Dot Labelling of Tepary Bean (Phaseolus acutifolius) Lectins by Microfluidics.

Lectins are bioactive proteins with the ability to recognize cell membrane carbohydrates in a specific way. Diverse plant lectins have shown diagnostic and therapeutic potential against cancer, and their cytotoxicity against transformed cells is mediated through the induction of apoptosis. Previous works have determined the cytotoxic activity of a Tepary bean (Phaseolus acutifolius) lectin fraction (TBLF) and its anti-tumorigenic effect on colon cancer. In this work, lectins from the TBLF were additionally purified by ionic-exchange chromatography. Two peaks with agglutination activity were obtained: one of them was named TBL-IE2 and showed a single protein band in two-dimensional electrophoresis; this one was thus selected for coupling to quantum dot (QD) nanoparticles by microfluidics (TBL-IE2-QD). The microfluidic method led to low sample usage, and resulted in homogeneous complexes, whose visualization was achieved using multiphoton and transmission electron microscopy. The average particle size (380 nm) and the average zeta potential (-18.51 mV) were determined. The cytotoxicity of the TBL-IE2 and TBL-IE2-QD was assayed on HT-29 colon cancer cells, showing no differences between them (p ≤ 0.05), where the LC50 values were 1.0 × 10-3 and 1.7 × 10-3 mg/mL, respectively. The microfluidic technique allowed control of the coupling between the QD and the protein, substantially improving the labelling process, providing a rapid and efficient method that enabled the traceability of lectins. Future studies will focus on the potential use of the QD-labelled lectin to recognize tumor tissues.

Tepary Bean (Phaseolus acutifolius) Lectins Induce Apoptosis and Cell Arrest in G0/G1 by P53(Ser46) Phosphorylation in Colon Cancer Cells.

A Tepary bean lectin fraction (TBLF) has been studied because it exhibits differential cytotoxic and anticancer effects on colon cancer. The present work focuses on the evaluation of the apoptotic mechanism of action on colon cancer cells. Initially, lethal concentrations (LC50) were obtained for the three studied cell lines (HT-29, RKO and SW-480). HT-29 showed the highest LC50, 10 and 100 times higher than that of RKO and SW-480 cells, respectively. Apoptosis was evaluated by flow cytometry, where HT-29 cells showed the highest levels of early and total apoptosis, caspases activity was confirmed and necrosis was discarded. The effect on cell cycle arrest was shown in the G0/G1 phase. Specific apoptosis-related gene expression was determined, where an increase in p53 and a decrease in Bcl-2 were observed. Expression of p53 gene showed the maximum level at 8 h with an important decrease at 12 and 24 h, also the phosphorylated p53(ser46) increased at 8 h. Our results show that TBLF induces apoptosis in colon cancer cells by p-p53(ser46) involvement. Further studies will focus on studying the specific signal transduction pathway.

Effects of intragastrically-administered Tepary bean lectins on digestive and immune organs: Preclinical evaluation.

Previous work showed that Tepary bean (Phaseolus acutifolius) lectins exhibit differential cytotoxic effects on cancer cell lines by apoptosis induction. In vivo studies using a Tepary bean lectin fraction (TBLF, 50 mg/kg of body weight) after colon cancer induction in rats showed that TBLF inhibited early precancerous lesions without systemic toxicity however, loss of body weight gain and activation of immune cells were observed. In order to know more about the possible adverse effects, we evaluated the administration of TBLF on digestive and immune organs. Sprague Dawley rats were administered TBLF for six weeks and allowed to recover for two weeks. Immune activation was observed through an increased lymphocyte-granulocyte ratio, an increased number of lymphoid follicles in intestinal Peyer's patches and a slight expansion of the splenic white pulp. Atrophy was observed in small intestine villi and crypt foci of the colon without normalization after the recovery period. Pancreas histopathology showed hypertrophy after the six-week administration period, particularly vacuolation and trabecular widening; but after the two-week recovery period atrophy was observed, suggesting a partial compensatory type process. Our results show that TBLF activates the immune system and affects digestive organs through direct interaction with intestinal epithelium, and indirectly by producing pancreatic hyperfunction. Further work will focus in longer recuperation periods after TBLF treatment.

Phaseolus acutifolius Lectin Fractions Exhibit Apoptotic Effects on Colon Cancer: Preclinical Studies Using Dimethilhydrazine or Azoxi-Methane as Cancer Induction Agents.

Phaseolus acutifolius (Tepary bean) lectins have been studied as cytotoxic molecules on colon cancer cells. The toxicological profile of a Tepary bean lectin fraction (TBLF) has shown low toxicity in experimental animals; exhibiting anti-nutritional effects such as a reduction in body weight gain and a decrease in food intake when using a dose of 50 mg/kg on alternate days for six weeks. Taking this information into account, the focus of this work was to evaluate the effect of the TBLF on colon cancer using 1,2-dimethylhydrazine (DMH) or azoxy-methane/dextran sodium sulfate (AOM/DSS) as colon cancer inductors. Rats were treated with DMH or AOM/DSS and then administered with TBFL (50 mg/kg) for six weeks. TBLF significantly decreased early tumorigenesis triggered by DMH by 70%, but without any evidence of an apoptotic effect. In an independent experiment, AOM/DSS was used to generate aberrant cryptic foci, which decreased by 50% after TBLF treatment. TBLF exhibited antiproliferative and proapoptotic effects related to a decrease of the signal transduction pathway protein Akt in its activated form and an increase of caspase 3 activity, but not to p53 activation. Further studies will deepen our knowledge of specific apoptosis pathways and cellular stress processes such as oxidative damage.

Plant Lectins as Medical Tools against Digestive System Cancers.

Digestive system cancers-those of the esophagus, stomach, small intestine, colon-rectum, liver, and pancreas-are highly related to genetics and lifestyle. Most are considered highly mortal due to the frequency of late diagnosis, usually in advanced stages, caused by the absence of symptoms or masked by other pathologies. Different tools are being investigated in the search of a more precise diagnosis and treatment. Plant lectins have been studied because of their ability to recognize and bind to carbohydrates, exerting a variety of biological activities on animal cells, including anticancer activities. The present report integrates existing information on the activity of plant lectins on various types of digestive system cancers, and surveys the current state of research into their properties for diagnosis and selective treatment.