6 weeks consumption of pure fresh coconut milk caused up-regulation of eNOS and CSE protein expression in middle-aged male rats

I only copied the abstract and discussion portion, much more at link. What the hell is your doctor and stroke hospital going to do to get this tested in humans? Or will they DO NOTHING like usual? It is not their problem to solve? YOU are on your own to solve it, get going. 

6 weeks consumption of pure fresh coconut milk caused up-regulation of eNOS and CSE protein expression in middle-aged male rats

Chaweewan Jansakul^{1  2} 

Jomkarn Naphatthalung^{1  2} 

Sakda Pradab^{1  2} 

Somruedee Yorsin⁵ 

Kanyanatt Kanokwiroon^{3  4} 

¹ Faculty of Traditional Thai Medicine, Prince of Songkla University, Hat-Yai, Thailand

² Natural Product Research Centre of Excellence, Prince of Songkla University, Hat-Yai, Thailand

³Department of Biomedical Sciences, Faculty of Medicine, Prince of Songkla University, Hat-Yai, Thailand

⁴The Excellent Research Laboratory of Cancer Molecular Biology, Prince of Songkla University, Hat-Yai, Thailand

⁵Faculty of Medicine, Princess of Naradhiwas University, Narathiwas, Thailand

ABSTRACT

Coconut milk (CCM) has been an important cooking ingredient in the Asia-Pacific region since ancient time. Due to its high content of saturated fatty acids, it has been considered atherogenic. We have tested if chronic consumption of fresh coconut milk by middle-aged male rat affects vascular function, plasma glucose and lipid profiles. Compared to control, CCM caused lower maximal contraction to phenylephrine of thoracic aortic rings and increased relaxation to acetylcholine that was abolished by N ^G-nitro-L-arginine (L-NA) or disruption of the endothelium. DL-propargylglycine caused slight increase in baseline tension of L-NA treated aortic rings of CCM-treated rats and produced higher contractile response of the aortic rings to low concentrations of phenylephrine. The aortic eNOS- and cystathionine-γ-lyase(CSE) proteins expression of the CCM-treated rats were also higher than in controls. Except for lower fasting plasma glucose there were no changes in blood chemistry for the CCM treated rats. CCM consumption caused up-regulation of eNOS and CSE protein expression which resulted in increased production of NO and H₂S from the blood vessels with attenuation of vasocontraction to phenylephrine and increased relaxation to acetylcholine. These novel benefits may be expected to reduce the development of cardiovascular risk factors in the aging rat.

Discussion

The present study demonstrated that consumption of the CCM caused beneficial effects on the cardiovascular system if consumed in a sufficient amount. As shown in the results section, consumption of the CCM at a dosage of 1 g/kg for 6 weeks by the middle-aged rats did not result in any difference in the parameters studied. When the dosage was increased to 3 g/kg, CCM caused lowered food intake.The reduction in food intake was not associated with any change in body weight or in the weight of the organs measured. The reasons for this might be due to high lipid content (70%) of the CCM composition which consists dominantly of medium chain fatty acids (8: 0 to 12: 0; 45%). These fatty acids are absorbed directly to the portal vein except for lauric acid (C12:0) that is partly absorbed via the lymphatic system (^{Mu, Hoy, 2000}; 2001; 2002). More recently, ^{Valente et al. (2017}) reported that coconut oil consumption promoted less appetite in women with excess body fat. In addition, Jambor de Sousa et al. (2006) found that hepatic portal vein infusion of caprylic acid, a medium chain fatty acid, caused reduction of food intake by about 40% in 18-h food-deprived male rats. Another possibility is that CCM might contain an active substance that at high doses it causes satiety or reduced appetite resulting in reduced food intake. However, further specific studies would be needed to clarify these possibilities.
The alteration in vascular function, especially endothelial function, is the key event in the pathophysiology of atherosclerosis, as it was shown that endothelial dysfunction preceded and predicted atherosclerosis (^{Bonetti, Lerman, Lerman, 2003}; ^{Davignon,Ganz, 2004}).In the present study endothelial dysfunction of the middle-aged male rat(^{Chongsa et al., 2015}) was used to study the effects of chronic consumption of the CCM.Although there were no changes in animal blood pressure and heart rate after taking CCM compared to the distilled water control group, adosage of 3 g/kg but not 1g/kg of CCM, caused some beneficial changes in vascular function. These included reduction in maximal contractile response of thoracic aortic rings to phenylephrine and a higher maximal relaxation to acetylcholine. These effects were abolished by L-NA or removal of the endothelium, which indicates that the lowering of maximal contractile response to phenylephrine of the CCM-treated aortic rings might be due to an increase in nitric oxide production from the vascular endothelium. This was confirmed by the finding that eNOS protein expression of the thoracic aorta obtained from CCM-treated was higher than that of the control rats.^{Salil, Nevin, Rajamohan (2011}, 2012)reported that coconut kernel protein is rich in arginine which was able to rescue the pancreatic β-cells and cytoarchitecture in alloxan-induced diabetic Wistar Albino rat via the arginine-nitric oxide pathway. Thus, it is possible that the up-regulation of the vascular eNOS of the CCM treated middle-aged rat in the present study might be mediated by the coconut protein. However, further study of the isolated CCM protein consumption by the middle-aged rat would be needed to clarify this possibility.
It has been reported that high-fat diet consumption by young rats or mice for 12-14 weeks causes deficiency of aortic CSE and H₂S (^{Jenkins, Nguyen, Hart, 2016}; ^{Peh et al., 2014}). Thus, it is possible that chronic consumption by the middle-aged rat of the CCM which contained high content of coconut oil might affect blood vessel H₂S production. To test this possibility another set of the endothelium-intactaortic rings was used and preincubated with L-NA in order to inhibit eNOS activity in order to prevent any disturbances by nitric oxide before adding PAG, a cystathionine-γ-lyase inhibitor; the vasocontraction to phenylephrine was then determined. As shown in the results section, adding PAG to the incubation medium caused a greater increase in basal baseline tension of the aortic rings obtained from CCM-treated rats than that of the control group, suggesting that the activity of the CSE enzyme of the CCM-treated rats was higher than that of the control rats. These results sequentially caused an increase in contractile response to low concentrations of the phenylephrine on the aortic rings that was higher than that of the control group. The finding that the blood vessel CSE protein expression was higher in the CCM treated rats is consistent with the finding that consumption of CCM caused an increase in blood vessel H₂S production. This result is in contrast to those of the Jenkins, Nguyen, Hart(2016) and Peh et al.(2014) who found that high fat diet caused a decrease in vascular CSE and H₂S production.The reason for this might be the differences in the types of fatty acid used. In the present study we used CCM which contained mostly medium-chain fatty acid which was found to cause an increase in mitochondrial and peroxisomal β-oxidation of fatty acids (^{Arunima, Rajamohan, 2014}). In contrast, Jenkins, Nguyen, Hart (2016) and Peh et al.(2014) used western diet and high fat diet, respectively, which contained mostly long-chain fatty acid, which was found to induce vascular oxidative stress and reduce endothelial function. As another possibility, ^{Zhao et al. (2001}) found that endogenous H₂S production from different vascular tissues including thoracic aorta was enhanced by the NO. Thus, the increased CSE protein expression which resulted in an increased H₂S production in the present study might be facilitated by the increased NO production elicited by the CCM, as mentioned above. However, further study is needed to clarify these possibilities.
In the present study it was also found that chronic consumption of CCM caused a slight decrease in basal fasting plasma glucose compared to that of the control group. As mentioned above, the reason for this might be due to the increase in basal level of NO since it has been reported that low concentration of NO exerted positive regulation of insulin sensitivity and secretion (^{Carvalho et al., 2016}; ^{Kurohane Kaneko, Ishikawa, 2013}). However, to clarify this possibility, further studyto measure basal plasma insulin level and/or the glucose insulin sensitivity of the CCM-treated and the control middle-aged rats would be necessary.
Taken together, consumption of CCM at the dosage of 3 g/kg increased blood vessel eNOS and CSE protein expression resulting in increased NO and H₂S production to attenuate the contractile response of thoracic aortic rings to phenylephrine and potentiate vasodilatation to acetylcholine, all of which could be expected to prolong vascular health of the middle-aged rat. In addition, CCM consumption caused lowering of plasma glucose level with no harm to liver or kidney functions, or on fat metabolism. Thus, CCM could be a novel food to develop as a nutraceutical for vascular health of the aging human being. Nevertheless, further work is required to identify the mechanism that causes CCM to lower plasma glucose and the vascular effects described.