Study On Encapsulated ANP-producing Cell Used In Treatment Of Hypertension

OBJECTIVEA technique based on release of the human atrial natriuretic peptide (hANP) from plasmid hANP cDNA transfected Chinese hamster ovary (CHO) cells encapsulated in polycaprolactone (PCL)-capsules was investigated for a potential therapeutic approach to hypertension or congestive heart failure (CHF).METHODSThe plasmid hANP cDNA was transfected into CHO cells, then, the cells were encapsulated in PCL-capsules and the levels of hANP secreted by encapsulated hANP-producing CHO cells were detected by RIA specific for hANP. Circadian rhythm of hANP secreted by encapsulated hANP-producing CHO cells was studied and regulated with melatonin. Encapsulated plasmid hANP cDNA transfected CHO cells were implanted into two-kidney, one-clip (2K1C) hypertensive rats intraperitoneally. The physiological parameters and morphological changes, histological changes, the circadian rhythm of renal excretion were investigated post-implantation of PCL-capsulesin 2K1C hypertensive rats. In addition, several parameters on safety of gene therapy were analyzed post-implantation of PCL-capsules in 2K1C hypertensive rats. RESULTSDuring culturing, the encapsulated plasmid liANP cDNA transfected CHO cells remained viable and the mean levels of hANP reached 246.1pg/ml/24hr in 2ml medium containing one PCL-capsule that is 20mm in length and 3mm in diameter, whereas the control was negative. The secretion of hANP showed the circadian variation and the level of hANP at night was higher, but at daytime was lower. The acrophase of circadian rhythm was 4:18. After given melatonin, the acrophase of circadian rhythm of hANP secreted from encapsulated plasmid hANP cDNA transfected cells shifted to 7:56.The implantation of encapsulated hANP-producing cells caused a significant delay of blood pressure (BP) increase 2 days post-implantation and the effect lasted for more than 5 months in 2K1C hypertensive rats. The encapsulated hANP-producing cells also caused significant increases in renal blood flow (RBF), glomerular filtration rate (GFR), sodium output, urine excretion, urinary cGMP levels in 2K1C hypertensive rats comparing with the control rats. These effects were reflected morphologically by an attenuation of the glomerular sclerotic lesions, reduction in cardiomyocyte size, the ratio of left ventricular weight to heart weight, tubular damage and renal arterial thickening. Plasma levels of hANP in 2K1C rats implanted with the PCL-capsules containing hANP-producing cellswere higher than that of the control rats. During a 12:12 light-dark cycle, the implantation of the encapsulated hANP-producing cells increased the net excretion of water and caused an advance shift of the acrophases in 2K1C rats comparing with 2K1C rats implanted with encapsulated control CHO cells. Similarly, the implantation of encapsulated hANP-producing cells caused an increase in the net excretion of sodium, potassium and also reversed the delay shifts of the acrophases caused by renovascular hypertension in 2K1C rats. The several parameters on safety of gene therapy were normal 10 days post-implantation in 2K1C hypertensive rats. The encapsulated transfected CHO cells remained viable 5 months after implanted. CONCLUSIONThis study demonstrated that the usefulness of encapsulated hANP gene transfected cells as a new tool for hANP gene delivery in studying renovascular hypertension and cardiovascular diseases. Thus, our results may have important implications for clinical use of gene transfected cells as therapeutic agents in the treatment of cardiovascular diseases. Moreover, this approach avoided the side effects of gene therapy by virus vectors. Encapsulation technique may be used for gene transfected cells implanted into human body for treatment in the future.