Physiologic homeostasis after pig-to-human kidney xenotransplantation

Abstract

Demand for kidney grafts outpaces supply, limiting kidney transplantation as a treatment for kidney failure. Xenotransplantation has the potential to make kidney transplantation available to many more patients with kidney failure, but the ability of xenografts to support human physiologic homeostasis has not been established. A brain-dead adult decedent underwent bilateral native nephrectomies followed by 10 gene-edited (four gene knockouts, six human transgenes) pig-to-human xenotransplantation. Physiologic parameters and laboratory values were measured for seven days in a critical care setting. Data collection aimed to assess homeostasis by measuring components of the renin-angiotensin-aldosterone system, parathyroid hormone signaling, glomerular filtration rate, and markers of salt and water balance. Mean arterial blood pressure was maintained above 60 mmHg throughout. Pig kidneys secreted renin (post-operative day three to seven mean and standard deviation: 47.3 ± 9 pg/mL). Aldosterone and angiotensin II levels were present (post-operative day three to seven, 57.0 ± 8 pg/mL and 5.4 ± 4.3 pg/mL, respectively) despite plasma renin activity under 0.6 ng/mL/hr. Parathyroid hormone levels followed ionized calcium. Urine output down trended from 37 L to 6 L per day with 4.5 L of electrolyte free water loss on post-operative day six. Aquaporin 2 channels were detected in the apical surface of principal cells, supporting pig kidney response to human vasopressin. Serum creatinine down trended to 0.9 mg/dL by day seven. Glomerular filtration rate ranged 90-240 mL/min by creatinine clearance and single-dose inulin clearance. Thus, in a human decedent model, xenotransplantation of 10 gene-edited pig kidneys provided physiologic balance for seven days. Hence, our in-human study paves the way for future clinical study of pig-to-human kidney xenotransplantation in living persons.

Keywords: aldosterone; parathyroid hormone; renin-angiotensin system; transplantation; water channels.

Figure 1 ∣. Renin-angiotensin-aldosterone system.

Xenotransplant recipient hormone plasma concentrations over time; shaded areas represent normal human ranges for each hormone. (a) Renin (pg/ml): normal, <45.7 pg/ml. Plasma renin activity was <0.6 ng/ml per hour at all time points. (b) Angiotensinogen (μg/ml); 71 μg/ml is the upper limit of normal. (c) Angiotensin II (pg/ml): normal range, 3 to 30 pg/ml. (d) Aldosterone (pg/ml) normal range, 31 to 354 pg/ml.

Figure 2 ∣. Parathyroid hormone (PTH) levels and ionized calcium (Ca) levels in decedent following xenotransplantation.

Figure 3 ∣. Renal clearance physiology.

(a) Inulin decay curve, concentration at timed intervals after a 10-g bolus injection (see Supplementary Figure S1 for method). (b) Pig kidney clearance grouped by method of measurement. (c) Serum creatinine trend. (d) Tacrolimus pharmacokinetics. AUC_0-12, area under curve 0–12 hours; CKD-EPI, Chronic Kidney Disease Epidemiology Collaboration; CrCl, creatine clearance; eGFR, estimated glomerular filtration rate; GFR, glomerular filtration rate.

Figure 4 ∣. Water and sodium balance.

(a) Decedent’s daily urine output after xenotransplantation (L). Intraoperative furosemide, 100 mg, and mannitol, 25 g, were administered i.v. prior to reperfusion. (b) Serum sodium. (c) Water clearance after xenotransplantation (L). (d) Urine osmolarity (mOsm/kg H₂O).

Figure 5 ∣. Aquaporin (AQP) expression in the 10 gene-edited xenokidney.

(a) AQP1 in the apical side of the proximal tubule. (b) AQP4 in the basolateral membrane of the principal cells of the collecting duct. Arrows indicate principal cells positive for AQP4. (c) AQP2 in the apical membrane of the principal cells. (d) AQP2 phosphorylation S256, a known activated form of AQP2, is also expressed in the principal cells. (e) Immunofluorescent labeling of principal cells with AQP2–488– (green) and vacuolar-type H+–adenosine triphosphatase B1/2 (V-ATPase)–positive staining of intercalated cells (red); see Supplementary Figure S4 for negative control. (f) Cortex and (g) medulla: representative trichrome-stained sections with proximal tubules (PTs) and collecting ducts lined by pale staining principal cells and rare darkly stained intercalated cells. Asterisks (*) denote intercalated cells. Bar = 50 μm.