Jayadev Raju

Trigonella foenum graecum (fenugreek) seed powder improves glucose homeostasis in alloxan diabetic rat tissues by reversing the altered glycolytic, gluconeogenic and lipogenic enzymes

Trigonella foenum graecum (fenugreek) seed powder has been suggested to have potential antidiabetic effects. The effect of oral administration of Trigonella whole seed powder (5% in the diet) for 21 days on glycolytic, gluconeogenic and NADPlinked lipogenic enzymes were studied in liver and kidney tissues of alloxan-induced diabetic Wistar rats. Diabetic rats were characterised by a 4fold higher blood glucose level and a 0.7fold lower body weight compared to normal controls. The activities of the glycolytic enzymes were significantly lower in the diabetic liver and higher in the diabetic kidney. The activities of gluconeogenic enzymes were higher in both liver and kidney during diabetes, however the activities of the lipogenic enzymes were decreased in both tissues during diabetes. Trigonella seed powder treatment to diabetic rats for 21 days brought down the elevated fasting blood glucose levels to control levels. The altered enzyme activities were significantly restored to control values in both the liver and kidney after Trigonella seed powder treatment. The therapeutic role of Trigonella seed powder in type1 diabetes as exemplified in this study can be attributed to the change of glucose and lipid metabolising enzyme activities to normal values, thus stabilizing glucose homeostasis in the liver and kidney. These biochemical effects exerted by Trigonella seeds make it a possible new therapeutic in type1 diabetes.

β-Escin inhibits colonic aberrant crypt foci formation in rats and regulates the cell cycle growth by inducing p21waf1/cip1 in colon cancer cells

Extracts of Aesculus hippocastanum (horse chestnut) seed have been used in the treatment of chronic venous insufficiency, edema, and hemorrhoids. Most of the beneficial effects of horse chestnut are attributed to its principal component β-escin or aescin. Recent studies suggest that β-escin may possess anti-inflammatory, anti-hyaluronidase, and anti-histamine properties. We have evaluated the chemopreventive efficacy of dietary β-escin on azoxymethane-induced colonic aberrant crypt foci (ACF). In addition, we analyzed the cell growth inhibitory effects and the induction of apoptosis in HT-29 human colon cancer cell line. To evaluate the inhibitory properties of β-escin on colonic ACF, 7-week-old male F344 rats were fed experimental diets containing 0%, 0.025%, or 0.05% β-escin. After 1 week, the rats received s.c. injections of azoxymethane (15 mg/kg body weight, once weekly for 2 weeks) or an equal volume of normal saline (vehicle). Rats were continued on respective experimental diets and sacrificed 8 weeks after the azoxymethane treatment. Colons were evaluated histopathologically for ACF. Administration of dietary 0.025% and 0.05% β-escin significantly suppressed total colonic ACF formation up to ∼40% (P < 0.001) and ∼50% (P < 0.0001), respectively, when compared with control diet group. Importantly, rats fed β-escin showed dose-dependent inhibition (∼49% to 65%, P < 0.0001) of foci containing four or more aberrant crypts. To understand the growth inhibitory effects, HT-29 human colon carcinoma cell lines were treated with various concentrations of β-escin and analyzed by flow cytometry for apoptosis and cell cycle progression. β-Escin treatment in HT-29 cells induced growth arrest at the G1-S phase, which was associated with the induction of the cyclin-dependent kinase inhibitor p21WAF1/CIP1, and this correlated with reduced phosphorylation of retinoblastoma protein. Results also indicate that β-escin inhibited growth of colon cancer cells with either wild-type or mutant p53. This novel feature of β-escin, a triterpene saponin, may be a useful candidate agent for colon cancer chemoprevention and treatment. [Mol Cancer Ther 2006;5(6):1459–66]

Low doses of β-carotene and lutein inhibit AOM-induced rat colonic ACF formation but high doses augment ACF incidence

Epidemiological studies suggest that carotenoids such as β‐carotene and lutein play an important role in reducing the risk for several cancers. However, in colon cancer there is ambiguity with regard to the role of these compounds in that both preventive effects and tumor promotion have been observed. In the present study we observed that male F344 rats were able to tolerate up to 2,500 ppm of β‐carotene as well as of lutein. We have then assessed the chemopreventive efficacy of β‐carotene and lutein at dose levels of ∼4 and 8% of the 2,500 ppm tolerated dose (TD) and also ∼40 and 80% of the TD on azoxymethane (AOM)‐induced colon carcinogenesis, using aberrant crypt foci (ACF) as a surrogate biomarker for colon cancer. Throughout the experiments, 5‐week‐old male F344 rats were fed the control diet (modified AIN‐76A) or experimental diets containing 100 or 200 ppm (∼4 or 8% of the 2,500 ppm TD), or 1,000 or 2,000 ppm (∼40 or 80% of the 2,500 ppm TD) of β‐carotene and lutein (n=10 rats/group). After 2 weeks on the experimental or control diets, all animals were injected with AOM (15 mg/kg body wt., once weekly for 2 weeks). At 14 weeks of age, all rats were killed, and their colons were evaluated for ACF. Administration of 100 or 200 ppm of β‐carotene inhibited AOM‐induced total colonic ACF formation by 24% (p<0.01) and 36% (p<0.001), respectively, whereas lutein at 200 ppm produced a 27% inhibition (p<0.01) yet had no significant effect at the 100 ppm dose level. Surprisingly, administration of 1,000 or 2,000 ppm of β‐carotene and lutein increased colonic ACF formation in a dose‐dependent manner, i.e., to 124% and 144% for the former and 110% and 159% for the latter. These results clearly suggest that further studies are warranted to determine whether the increase in ACF incidence by high doses of β‐carotene and lutein will also lead to an increase in tumor outcome. Taken together these data indicate that the chemopreventive activity of β‐carotene and lutein against colon carcinogenesis depends on the dose level.

Diosgenin, a Steroid Saponin Constituent of Yams and Fenugreek: Emerging Evidence for Applications in Medicine

Phytochemicals found in foods and spices are progressively gaining popularity over conventional synthetic drugs mainly because they act via multiple molecular targets that synergize to efficiently prevent or treat chronic illnesses. Phytochemicals are also safe (with minimal or no toxic or side effects) with better bioavailability. Food saponins have been used in complimentary and traditional medicine against a variety of diseases including several cancers. Diosgenin, a naturally-occurring steroid saponin is found abundantly in legumes (Trigonella sp.) and yams (Dioscorea sp.). Diosgenin is a precursor of various synthetic steroidal drugs that are extensively used in the pharmaceutical industry. Over the past two decades, a series of pre-clinical and mechanistic studies have been independently conducted to understand the beneficial role of diosgenin against metabolic diseases (hypercholesterolemia, dyslipidemia, diabetes and obesity), inflammation and cancer. In experimental models of obesity, diosgenin decreases plasma and hepatic triglycerides and improves glucose homeostasis plausibly by promoting adipocyte differentiation and inhibiting inflammation in adipose tissues. A limited number of experiments have been conducted to understand the preclinical efficacy of diosgenin as a chemopreventive/therapeutic agent against cancers of several organ sites. Mechanistic studies using in vitro models suggest that diosgenin suppresses cancer cell growth through multiple cell signaling events associated with proliferation, differentiation, apoptosis, inflammation and oncogenesis. This chapter provides a comprehensive review of the biological activity of diosgenin that contributes to several diseases in its role as a health beneficial phytochemical by citing new studies. In addition, diosgenin’s safety with regards to its potential toxicity is also critically discussed. Altogether, the findings from pre-clinical and mechanistic studies strongly implicate the use of diosgenin as a novel multi-target based chemopreventive or therapeutic agent against several chronic diseases.

Steady state levels of transforming growth factor-β1 and -β2 mRNA and protein expression are elevated in colonic tumors in vivo irrespective of dietary lipids intervention

Colonic tumors of human origin produce abundant transforming growth factor (TGF)‐β suggesting that TGF‐β is critical to their growth. Dietary lipids regulate a number of growth factors including TGF‐β. Whether elevated TGF‐β levels are consistently expressed in colonic tumors irrespective of the environmental milieu in an in vivo model is not known and forms the main objective of the present study. Male F344 rats were injected with azoxymethane, 10 weeks later, rats bearing preneoplastic lesions were fed a low fat (5% corn oil) diet and 3 high fat (5% corn oil with 18% corn oil, fish oil or beef tallow) diets for 16 weeks. Colonic tumors and mucosae were processed and assessed for TGF‐β status. TGF‐β1 and ‐β2 mRNA levels were upregulated in colonic tumors more than in mucosae of all diet groups. Dietary lipids modulated TGF‐β mRNA in both tumors and mucosae, high corn and fish oil diets upregulated TGF‐β1 significantly more than the low fat corn oil or high fat beef tallow diets. Immunohistochemical assessments of tissues with different biological features revealed that TGF‐β1 and ‐β2 were elevated in tumors and in selected microscopic preneoplastic lesions compared to normal mucosae. This is the first in vivo study, documenting that developing colonic tumors acquire upregulated TGF‐β phenotype even in the presence of lipid environments capable of differentially regulating TGF‐β in normal mucosae. Elevated expression of TGF‐β in a selected subset of microscopic preneoplastic lesions suggests that TGF‐β plays an important role on both early and late stages of colon carcinogenesis.

Evidence for the involvement of dietary lipids on the modulation of transforming growth factor-β1 in the platelets of male rats

Transforming growth factor β1 (TGF-β1), a multifunctional cytokine participates in the proliferation and differentiation of various cell types. Platelets are an important source of TGF-β1 and are physiologically linked to a variety of chronic illnesses including cancer, heart disease and inflammation. It is well known that dietary lipids modulate platelet function. Whether dietary lipids affect growth factor status of platelets is not known. This study addresses the effect of dietary lipids on TGF-β1 status of the platelets. Male 8 month-old Sprague Dawley rats were allocated to different diet groups. The high fat diets (18% by weight) comprising of high fat beef tallow (HFB), high fat corn oil (HFC), high fat fish oil (HFF) and high fat olive oil (HFO) and one low fat diet containing low fat soybean oil (LFS) (5% by weight) were fed to the experimental animals for 6 weeks. The TGF-β1 status in the platelet lysate was assessed by using the CCL-64 mink lung cell bioassay and by Western blot analysis. Platelet lysates were evaluated for their ability to inhibit the growth of the CCL-64 mink lung cells, unexpectedly platelet lysates stimulated growth. The stimulatory effect of platelet lysate was in the order HFF > HFO > HFB > HFC > LFS. Acidification of the lysates to activate the latent form of TGF-β1 resulted in the loss of the growth stimulatory potential of the platelet lysates in all the groups. Western blot analysis of the platelet lysates to detect the level of TGF-β1 protein demonstrated that HFB diet group had the highest level of TGF-β1 and the HFC diet group had the lowest level of TGF-β1 and were significantly different (p < 0.05) as compared to the other three diet groups. These findings demonstrate that dietary lipids varying in their fatty acid composition, profoundly affect the level of growth modulating constituents of the platelets. Further studies are warranted to refine our understanding of the effect of dietary constituents on the physiology of the platelets.

Differential modulation of transforming growth factor-βs and cyclooxygenases in the platelet lysates of male F344 rats by dietary lipids and piroxicam

Platelets are implicated in the pathogenesis of various chronic diseases including cancer. The main objective of the present study was to determine if dietary fish oil and piroxicam, known modulators of colon tumorigenesis, effect transforming growth factor (TGF)-βs and cyclooxygenase (COX) isozymes in the platelets of colon tumor-bearing male F344 rats. TGF-βs and COXs are important in the development of chronic illnesses including colon cancer. Animals harboring preneoplastic colonic lesions were randomly allocated to a low fat diet (5% by weight – low corn oil, LFC) and three high fat diets (23% by weight – high corn oil, HFC; high corn oil containing 150-ppm piroxicam, HFC+P; and high fish oil, HFF) for 16 weeks. TGF-β1, TGF-β2, COX-1 and COX-2 protein levels were assessed in the platelets by Western blot analysis. Active TGF-β1 (12.5 kDa) level was significantly lower in the platelets of the HFC+P group (p < 0.001), whereas precursor TGF-β1 (39 kDa) level was significantly lower in the platelets of the HFF group (p < 0.001). The anti-rabbit TGF-β2 polyclonal antibody did not detect the 13-kDa active TGF-β2 protein in the platelets. However a 29-kDa protein, potentially a precursor of TGF-β2, was detected in the platelets of all the groups and was significantly lower in the HFC+P and HFF groups than in LFC and HFC (p < 0.001). COX-1 level was significantly lower in the HFF group than the other three groups (p < 0.001). COX-2 protein was detected in the platelets of all diet groups. Piroxicam in the presence of high corn oil (HFC+P) significantly lowered the level of COX-2 (p < 0.001), without having any effect on COX-1 level. These findings conclusively show that LFC and HFC differ from HFF and HFC+P, and piroxicam differs from fish oil, in regulating the levels of TGF-βs and COX in the platelets. This supports the conjecture that the levels of bioactive constituents of the platelets are profoundly modulated by dietary lipids, which in turn could influence the pathogenesis of chronic illnesses.