1. Soy isoflavones and nicotinamide
down-regulate pro-inflammatory mediators in vitro
R Di Caprio1, S Lembo2, A Balato3, F Gasparri4, G Monfrecola1
1Department of Clinical Medicine and Surgery, University of Naples “Federico II”, Italy
2Department of Medicine, Surgery and Dentistry “Scuola Medica Salernitana”, University of Salerno, Italy
3Department of Advanced Biomedical Sciences, University of Naples Federico II, Italy
4Department of Pharmacy (DIFARMA), University of Salerno, Italy
INTRODUCTION:
Isoflavones, the major soybean flavonoids, exist predominantly as glucosides such as daidzin and genistin. They have recently attracted considerable interest because of their beneficial properties for coronary heart disease, cancer prevention, and osteoporosis. The metabolism and absorption of isoflavones ingested with food is well documented, but little is known about their use as topical photo-protective agents. Nicotinamide (pyridine-3-carboxamide) is an amide active form of Vitamin B3 or niacin (pyridine-3-carboxylic acid).
Both nicotinamide (NCT) and niacin are precursors for the synthesis of nicotinamide adenine dinucleotide (NAD+) and its phophorylated derivative (NADP+): therefore NCT plays an important role in maintaining energy for cellular functions. Several animal studies have shown that the use of topical NCT and/or systemic supplementation of niacin, can protect against UV-induced immunosuppression thus reducing the development of non-melanoma skin cancers.
AIM OF THE STUDY:
To evaluate, in vitro, the anti-inflammatory properties of soy isoflavones and nicotinamide,
singularly or in combination, in UVB-irradiated keratinocytes.
MATERIALS AND METHODS:
Soy isoflavones were used in two different forms: ISOSOY and Lo ISO-3s. The first, is a standardized glycine soy extract titrated to 90% in isoflavone aglycones (with genistein:daidzein=1:4), soluble in dimethylsulfoxide. The second, consists in microencapsulated soy extracts, enriched in daidzein and genistein, obtained by spray-drying technique using a sodium-carboxymethylcellulose based matrix, soluble in water. Immortalized human keratinocytes were pre-incubated for 2h with ISOSOY and NCT or with Lo ISO-3s and NCT, singularly or in combination, and irradiated with UVB 60 mJ/cm2. Substances toxicity was assessed through cell viability and their possible anti-inflammatory effect was explored analyzing IL-6, TNF-α and COX-2 gene expression (qRT-PCR) 24h after UVB irradiation.
STUDY DESIGN:
RESULTS:
Incubation of HaCaT cells with substances for 2h
RNA extraction and processing h24
UVB-irradiation (60 mJ/cm2)
HaCaT (DMEM)-5% CO2, 37°
IL-6, TNF-α and COX-2 gene expression
NCT= Nicotinamide 20µM; ISOSOY= Soy isoflavones 10µM (DMSO 0,01%);
Lo ISO-3s= Soy isoflavones 10µM
Effects of NCT, ISOSOY and Lo ISO-3s:
cell viability
C, 80% confluence, h-2
None of the tested substances, used alone or in combination, appreciably affected cell viability respect to the controls.
UVB-induced cytotoxicity rate and statistical significance were determined with respect to the 100% viability of untreated control cells; viability of HaCaT cells pretreated with NCT, ISOSOY and Lo ISO-3s, and irradiated with UVB 60mJ/cm2, was compared to cells irradiated only.
IL-6, TNF-α and COX-2 gene expression
Lo ISO-3s was more effective than ISOSOY in down-regulating IL-6 and COX-2 increases (p<0.01), while ISOSOY was more effective than Lo ISO-3s in down-regulating TNF-α increases (p<0.01). A part from TNF-α, Lo ISO-3s was more effective than NCT in reducing UVB-induced inflammation, while ISOSOY, generally, resulted less effective.
Data were compared to UVB-irradiated cells and normalized to the housekeeping gene 18S. Substances-induced cytokines decreases and statistical significance were determined with respect to the 100% fold increase of UVB-irrdiated cells (*) and between cells pretreated with ISOSOY and Lo ISO-3s (#).
Data are expressed as mean ± SD of three independent experiments, each performed in triplicate. Student’s t-test was used to calculate significant differences
(**P < 0.01; ***P < 0.001; ##P < 0.01).
CONCLUSION:
The results of our study indicate that soy isoflavones and NCT, used alone or in combination,
have a potential immuno-modulatory and cytoprotective effect
representing an interesting option to improve or prevent UV-induced/aggravated clinical conditions.