Generation of tryptophan hydroxylase 2 gene knockout pigs by CRISPR/Cas9-mediated gene targeting

Unbalanced brain serotonin(5-HT) levels have implications in various behavioral abnormalities and neuropsychiatric disorders. The biosynthesis of neuronal 5-HT is regulated by the rate-limiting enzyme, tryptophan hydroxylase-2(TPH2). In the present study, the clustered regularly interspaced short palindromic repeat(CRISPR)/CRISPR-associated(Cas) system was used to target the Tph2 gene in Bama mini pig fetal fibroblasts. It was found that CRISPR/Cas9 targeting efficiency could be as high as 61.5%, and the biallelic mutation efficiency reached at38.5%. The biallelic modified colonies were used as donors for somatic cell nuclear transfer(SCNT) and 10 Tph2 targeted piglets were successfully generated. These Tph2 KO piglets were viable and appeared normal at the birth.However, their central 5-HT levels were dramatically reduced, and their survival and growth rates were impaired before weaning. These Tph2 KO pigs are valuable large-animal models for studies of 5-HT deficiency induced behavior abnomality.

Predictors of heart rhythm disturbances in hypertensive obese patients with obstructive sleep apnea

Objective To assess the incidence and predictors of heart rhythm and conduction disturbances in hypertensive obese patients with andwithout obstructive sleep apnea （OSA）. Methods This is an open, cohort, prospective study. Out of 493 screened patients, we selected 279hypertensive, obese individuals without severe concomitant diseases： 75 patients without sleep-disordered breathing （non-SDB group）, and204 patients with OSA （OSA group）. At baseline, all patients underwent examination, including ECG, Holter ECG monitoring, and sleepstudy. During follow-up （on 3, 5, 7 and 10th years; phone calls once per 6 months）, information about new events, changes in therapy and lifestyle was collected, diagnostic procedures were performed. As the endpoints, we registered significant heart rhythm and conduction disordersas following： atrial fibrillation （AF）, ventricular tachycardia, atrioventricular block （AV） 2-3 degree, sinoatrial block, significant sinus pauses（〉 2000 ms）, and the required pacemaker implantation. Results The median follow-up was 108 （67.5-120） months. The frequency of heartrhythm disorders was higher in OSA patients （29 cases, X^2= 5.5; P = 0.019） compared to the non-SDB patients （three cases; OR： 3.92, 95%CI： 1.16-13.29）. AF was registered in 15 patients （n = 12 in OSA group; P = 0.77）. Heart conduction disturbance developed in 16 patients,without an association with the rate of coronary artery disease onset. Regression analysis showed that only hypertension duration was anindependent predictor olAF （OR： 1.10, 95% CI： 1.04-1.16; P = 0.001）. In case of heart conduction disturbances, apnea duration was thestrongest predictor （P = 0.002）. Conclusions Hypertensive obese patients with OSA demonstrate 4-fold higher incidence of heart rhythmand conduction disturbances than subjects without SDB. Hypertension duration is an independent predictor for AF development, while sleepapneaJhypopnea duration is the main factor for heart conduction disorders onset in hypertensive obese patients with OSA.