1. 1 2013; 128: A18068

2. ReserachGate.net-Antonio Delgado-Almeida Core 7. Vascular Disease: Biology and Clinical Science Session Title: Hypertension: Clinical---Heart, Oxygen and Blood Pressure Circulation. Nov 17, 2013;128:A18068 Core 7. Vascular Disease: Biology and Clinical Science Session Title: Hypertension: Clinical---Heart, Oxygen and Blood Pressure Circulation. Nov 17, 2013;128:A18068 2 AHA 2013 E-Poster

3. BACKGROUND For a century, the pathogenesis and management of hypertension (HT) have been focused on dietary salt consumption, low potassium (K) intake, or hormonal factors triggering the elevation of BP. In this context, our description of an inherited defect of low RBC-K content, despite normal cell Na (Intermediate Phenotype) recorded in hypertensive subjects and half of their siblings and their adolescent offspring (Figure 1) implies a genetic defect from erythropoietic cells in the bone marrow, and a biochemistry and vascular disorders in circulating RBC. In effect, several RBC-K functions may be affected by a low RBC-K content including pyruvate kinase, a K-dependent enzyme activity involved on ATP synthesis and release in presence of low pH or low PO2, or the allosteric reaction of K and Oxygen biding by oxyhemoglobin, documented in our laboratory (Delgado-Almeida A. From Bohr Effect to the Electrical Regeneration of the Heart. FASEB J. March 29, 2012; 26:1126.7. Experimental Biology2012, San Diego, CA). Therefore, we explore this inherited defect in RBC-K content and transport, a major genetic alteration in hypertensive families, compared with control ones. The results in normotensive families will be presented elsewhere. For a century, the pathogenesis and management of hypertension (HT) have been focused on dietary salt consumption, low potassium (K) intake, or hormonal factors triggering the elevation of BP. In this context, our description of an inherited defect of low RBC-K content, despite normal cell Na (Intermediate Phenotype) recorded in hypertensive subjects and half of their siblings and their adolescent offspring (Figure 1) implies a genetic defect from erythropoietic cells in the bone marrow, and a biochemistry and vascular disorders in circulating RBC. In effect, several RBC-K functions may be affected by a low RBC-K content including pyruvate kinase, a K-dependent enzyme activity involved on ATP synthesis and release in presence of low pH or low PO2, or the allosteric reaction of K and Oxygen biding by oxyhemoglobin, documented in our laboratory (Delgado-Almeida A. From Bohr Effect to the Electrical Regeneration of the Heart. FASEB J. March 29, 2012; 26:1126.7. Experimental Biology2012, San Diego, CA). Therefore, we explore this inherited defect in RBC-K content and transport, a major genetic alteration in hypertensive families, compared with control ones. The results in normotensive families will be presented elsewhere. 3

4. FIGURE 1 Delgado MC, Delgado-Almeida A. Abnormal Potassium. In: Mohler III ER, Townsend RR (Eds). Advanced Therapy in Hypertension and Vascular Disease. BC Decker Inc. Publisher, Ontario, Canada.2006, Chapter 35: 291-99 (Copyright Figure) 4

5. MATERIALS AND METHODS Family Subjects: This was obtained from randomized hypertensive patients (n=316, 47± 11 years), their parents, siblings and their offspring (100 families), at times comprising three consecutive generations (Pedigree analysis) a)Ion Transport Research study: Evaluating a defective RBC-K along with plasma, renal electrolytes aimed to assess body K and Na physiology and disorders (Figure 2). b)Body composition (Bio-electrical Impedance, Quantum X, RJL Systems, Clinton, MI) For total body water (TBW), intra/extracellular water (ICW, ECW), Fat-Mass, Fat-Free- Mass, and estimated total body potassium (TBK). c)Central aortic Analysis: Recording of aortic BP, pulse waveform (PW) and reflected wave (time and pressure) by a non-invasive device in Office (DP 200M, Pulse Metric, Vista, CA) along with cardiac, vascular systemic and brachial hemodynamics by online Analysis. The aortic PW reflection was grouped as normal (Type I, in diastole) and hypertensive (Type II-IV, in systole). Travel Time of PW reflection was also measured and will be reported elsewhere. We have previously reported that gender differences in aortic PW reflection and CV hemodynamics are related to TBK (lower in females) in healthy subjects from Family Blood Pressure Program, NIH, at University of Michigan, (Figure 3) Family Subjects: This was obtained from randomized hypertensive patients (n=316, 47± 11 years), their parents, siblings and their offspring (100 families), at times comprising three consecutive generations (Pedigree analysis) a)Ion Transport Research study: Evaluating a defective RBC-K along with plasma, renal electrolytes aimed to assess body K and Na physiology and disorders (Figure 2). b)Body composition (Bio-electrical Impedance, Quantum X, RJL Systems, Clinton, MI) For total body water (TBW), intra/extracellular water (ICW, ECW), Fat-Mass, Fat-Free- Mass, and estimated total body potassium (TBK). c)Central aortic Analysis: Recording of aortic BP, pulse waveform (PW) and reflected wave (time and pressure) by a non-invasive device in Office (DP 200M, Pulse Metric, Vista, CA) along with cardiac, vascular systemic and brachial hemodynamics by online Analysis. The aortic PW reflection was grouped as normal (Type I, in diastole) and hypertensive (Type II-IV, in systole). Travel Time of PW reflection was also measured and will be reported elsewhere. We have previously reported that gender differences in aortic PW reflection and CV hemodynamics are related to TBK (lower in females) in healthy subjects from Family Blood Pressure Program, NIH, at University of Michigan, (Figure 3) AIMS To explored an inherited defect of low RBC-K content and function, unrelated to RBC-Na, in hypertensive families 5

6. Hypertensive Male No drugs Abnormal Results In Blue Figure 2 Ion Transport Research Laboratory 6

7. Figure 3 Histograms of TTRW in Healthy Subjects Males Females 7 Delgado AJ, Delgado MC, Weder AB. Am J Hypertens. 2003; 16(5, Suppl. 1): A139

8. In 100 hypertensive families and 585 offspring, there were 316 hypertensives (54%) and 269 normotensives (46%): Half of the adult offspring in families with 1 hypertensive parent, and 76 ± 21% if both parents were hypertensives. There were 234 grandchildren from 58% of hypertensive families, but only 14 cases (6%) had hypertension at age ≥ 30 years, suggesting that the genetic defect evolved to clinical hypertension around the 3rd decade of live. RESULTS Pedigree Analysis Low RBC-K was recorded in all hypertensives subjects, in 52% of their siblings including normotensive subjects, half of their adolescent offspring, and few of the available grandchildren. In hypertensive subjects (67%) and half of their siblings the renal function was altered: large 12-hours overnight urine volume (>1.2 ml/min) and increased Trans Tubular K gradient (TTKG), regardless of low or high salt intake or sodium excretion (Figure 2) These RBC-K and renal K disorders, despite normal dietary K, normal or low salt intake, and unchanged RBC-Na, indicated an ion transport mechanism independently of Na-K ATPase. In effect, this ATPase functions as a Na-specific pump, rejecting K and Ca ions (Nature. 2013;502(7470):201-6). Low RBC-K was recorded in all hypertensives subjects, in 52% of their siblings including normotensive subjects, half of their adolescent offspring, and few of the available grandchildren. In hypertensive subjects (67%) and half of their siblings the renal function was altered: large 12-hours overnight urine volume (>1.2 ml/min) and increased Trans Tubular K gradient (TTKG), regardless of low or high salt intake or sodium excretion (Figure 2) These RBC-K and renal K disorders, despite normal dietary K, normal or low salt intake, and unchanged RBC-Na, indicated an ion transport mechanism independently of Na-K ATPase. In effect, this ATPase functions as a Na-specific pump, rejecting K and Ca ions (Nature. 2013;502(7470):201-6). Ion Transport Studies 8

9. Abnormal aortic PW reflection (II-IV) was recorded in treated or untreated hypertensives (100%), and 52% of their normotensive siblings as compared with control subjects from normotensive parents, despite identical aortic BP (Figure 4). Such changes in aortic PWr became an easy vascular marker for hypertension, in normal adolescents offspring and adults subjects across three successive generations of individuals (Figure 5). Interestingly, this acute disturbed aortic BP, PW reflection and cardiovascular hemodynamics are recorded in nearly 40% of controlled hypertensives, regardless of the medical drug treatment. Up to date, it is the only reliable method to evaluate hemodynamic variables in treated hypertensive, and to assure effective BP control (Delgado-Almeida, et al. Recent Pat on Cardiovasc Drug Discov.2012;7:221-35). Abnormal aortic PW reflection (II-IV) was recorded in treated or untreated hypertensives (100%), and 52% of their normotensive siblings as compared with control subjects from normotensive parents, despite identical aortic BP (Figure 4). Such changes in aortic PWr became an easy vascular marker for hypertension, in normal adolescents offspring and adults subjects across three successive generations of individuals (Figure 5). Interestingly, this acute disturbed aortic BP, PW reflection and cardiovascular hemodynamics are recorded in nearly 40% of controlled hypertensives, regardless of the medical drug treatment. Up to date, it is the only reliable method to evaluate hemodynamic variables in treated hypertensive, and to assure effective BP control (Delgado-Almeida, et al. Recent Pat on Cardiovasc Drug Discov.2012;7:221-35). Bio-Impedance Analysis In most of randomized hypertensives (67%) TBW, ICW and estimated TBK were decreased or in low-normal ranges for age-sex and weight control groups. This decreased TWB/ICW was significantly related to the impaired renal water concentration, as measured by electrolyte free-water clearance (data no shown), and reversed after enhanced RBC-K transport. Central Aortic Hemodynamics 9

10. 10 Figure 4 Central Aortic PW, BP and PWr in Healthy Females Identical aortic BPs and HR in 26/y old healthy females from normotensive (Top) and hypertensive families (Bottom) Top: Aortic PW (right arrow), BPs, HR, PWr (type I, left arrow) and TTRW (248 ms) were normal. Bottom: Despite identical BPs and HR, aortic PWr (type IV, left arrow) impairs LV ejection time and pressures due to speedy TTRW (122 ms). Absence of pressure waves in diastole as in Top tracing. Identical aortic BPs and HR in 26/y old healthy females from normotensive (Top) and hypertensive families (Bottom) Top: Aortic PW (right arrow), BPs, HR, PWr (type I, left arrow) and TTRW (248 ms) were normal. Bottom: Despite identical BPs and HR, aortic PWr (type IV, left arrow) impairs LV ejection time and pressures due to speedy TTRW (122 ms). Absence of pressure waves in diastole as in Top tracing. 248 ms 122 ms

11. 11 Figure 5 Aortic PW Hemodynamics During Isometric Hand-Grip Test Resting aortic hemodynamics in normotensive 57 year old white female with no drugs Acute hemodynamic alterations after 3-minutes Isometric H-G test (maximal tension). Abnormal PWr type IV Acute hemodynamic alterations after 3-minutes Isometric H-G test (maximal tension). Abnormal PWr type IV Hemodynamic changes reversed 10 minutes after the Test, but the PWr type IV, as above tracings, implies risks of hypertensive crisis. Arterial pulse tracing identified specific studied areas (arrow). Hemodynamic changes reversed 10 minutes after the Test, but the PWr type IV, as above tracings, implies risks of hypertensive crisis. Arterial pulse tracing identified specific studied areas (arrow).

12. 12 Conclusions These results confirm a major hereditary defect in hypertensive subjects, in half of their siblings and normotensive offspring, probably explaining the large number of hypertensive population (1.1 billion, 2012), despite life style modification, dietary advice and use of anti-hypertensive drugs References in: ResearchGate.net-Antonio Delgado-Almeida PubMed.com These results confirm a major hereditary defect in hypertensive subjects, in half of their siblings and normotensive offspring, probably explaining the large number of hypertensive population (1.1 billion, 2012), despite life style modification, dietary advice and use of anti-hypertensive drugs References in: ResearchGate.net-Antonio Delgado-Almeida PubMed.com