Protection of Clitoria ternatea flower petal extract against free radical-induced hemolysis and oxidative damage in canine erythrocytes

The present study assessed the antioxidant activity and protective ability of Clitoria ternatea flower petal extract (CTE) against in vitro 2,2′-azobis-2-methyl-propanimidamide dihydrochloride (AAPH)-induced hemolysis and oxidative damage of canine erythrocytes. From the phytochemical analysis, CTE contained phenolic compounds, flavonoids, and anthocyanins. In addition, CTE showed antioxidant activity as measured by oxygen radical absorbance capacity (ORAC) method and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assay. CTE (400 µg/ml) remarkably protected erythrocytes against AAPH-induced hemolysis at 4 h of incubation. Moreover, CTE (400 µg/ml) reduced membrane lipid peroxidation and protein carbonyl group formation and prevented the reduction of glutathione concentration in AAPH-induced oxidation of erythrocytes. The AAPH-induced morphological alteration of erythrocytes from a smooth discoid to an echinocytic form was effectively protected by CTE. The present results contribute important insights that CTE may have the potential to act as a natural antioxidant to prevent free radical-induced hemolysis, protein oxidation and lipid peroxidation in erythrocytes.     

Anti-inflammatory effects of topical supernatant from human amniotic membrane cell culture on canine deep corneal ulcer after human amniotic membrane transplantation

The objective of this study was to examine the effect of topically applied human amniotic epithelial cell (HAEC) culture supernatant on corneal inflammatory reaction in dogs. Twenty-five dogs were randomly assigned into five groups. The control group consisted of five dogs with normal cornea. Inductions of corneal ulcers were performed using 0.45 cm trephine and human amniotic membrane was transplanted in 20 dogs. These 20 dogs were assigned into four treatment groups: topical antibiotic, topical corticosteroid, topical mock media and topical culture supernatant from HAEC, respectively. Administrations of the testing agents started at 24 h (h) after transplantation four times daily for nine consecutive days. Tears were collected before an operation 24 h after transplantation, but before application of the testing agents on consecutive odd days following transplantation. The concentrations of interleukin-1β (IL-1β) and nitric oxide (NO) in tear fluid were measured using canine IL-1β ELISA kit and Griess assay, respectively. Our analysis indicates that elevations of IL-1β and NO concentrations are associated with inflammatory conditions in the eyes. Corticosteroid, a reference anti-inflammatory drug, and the culture supernatant from HAEC significantly decreased IL-1β and NO concentrations. In addition, the clinical signs such as conjunctivitis and neovascularization were decreased in both topical corticosteroid and supernatant from HAEC treated groups. Mock and antibiotic solutions failed to decrease NO and IL-1β concentrations. In conclusion, topical application of the culture supernatant from HAEC alleviated inflammation in induced-corneal ulcer of dogs, possibly via inhibition of IL-1β and NO production.     

Insulin-releasing properties of a series of cinnamic acid derivatives in vitro and in vivo

Cinnamic acid derivatives are naturally occurring substances found in fruits, vegetables, and flowers and are consumed as dietary phenolic compounds. In the present study, cinnamic acid and its derivatives were evaluated for insulin secreting activity in perfused rat pancreas and pancreatic beta-cells (INS-1) as well as an increase in [Ca(2+)]i in vitro. The presence of m-hydroxy or p-methoxy residues on cinnamic acid was a significantly important substituent as an effective insulin releasing agent. The introduction of p-hydroxy and m-methoxy-substituted groups in cinnamic acid structure (ferulic acid) displayed the most potent insulin secreting agent among those of cinnamic acid derivatives. In particular, the stimulatory insulin secreting activities of test compounds were associated with a rise of [Ca(2+)]i in INS-1. In perfused rat pancreas, m-hydroxycinnamic acid, p-methoxycinnamic acid, and ferulic acid (100 microM) significantly stimulated insulin secretion during 10 min of administration. The onset time of insulin secretion of those compounds was less than 1 min and reached its peak at 4 min that was about 2.8-, 3.3-, and 3.4-fold of the baseline level, respectively. Intravenous administration of p-methoxycinnamic acid and ferulic acid (5 mg/kg) significantly decreased plasma glucose and increased insulin concentration in normal rats and maintained its level for 15 min until the end of experiment. Meanwhile, m-hydroxycinnamic acid induced a significant lowering of plasma glucose after 6 min, but the effects were transient with plasma glucose concentration, rapidly returning to basal levels. Our findings suggested that p-methoxycinnamic acid and ferulic acid may be beneficial for the treatment of diabetes mellitus because they regulated blood glucose level by stimulating insulin secretion from pancreatic beta-cells.