Cross-linked hemoglobin increases fractional reabsorption and GFR in hypoxic isolated perfused rat kidneys

Abstract

We compared the ability of human red blood cells (RBC) and a cell-free oxygen carrier to maintain isolated perfused kidney function under moderately hypoxic conditions. Recirculating perfusate was gassed initially with 93% air-7% CO2, and, after 30 min, the gas was changed to 12 O2-7 CO2-81% N2. Oxygen content of the perfusate was increased with RBC (30 g/l Hbg) or highly purified human hemoglobin Ao (HbAo) polymerized with O-raffinose (o-R-poly-Hb, 30 g/l Hbg). For comparison, kidneys were perfused with 60 g/l of bovine serum albumin (BSA) alone. The effects of unmodified hemoglobin were examined by adding 5 g/l of nonpolymerized HbAo to the BSA perfusate after 20 min. The effect of increasing oxygen delivery without hemoglobin was examined by switching to 93% O2 after 20 min during some BSA perfusions (BSA-HiO2). Vascular resistance decreased progressively in o-R-poly-Hb- and BSA-HiO2-perfused kidneys but remained constant in other experiments. Nitro-L-arginine methyl ester (L-NAME) prevented vasodilation and increased the filtration fraction of o-R-poly-Hb-perfused kidneys with no change in other functions. L-NAME also prevented the formation of methemoglobin. After a 70-min perfusion with BSA, Na reabsorption was 82 +/- 3% (means +/- SD), and inulin clearance [glomerular filtration rate (GFR)] was 0.66 +/- 0.33 ml.min-1.g-1. RBC increased reabsorption to 95% (85-98%) (median, 25th-75th percentile) but did not alter GFR (0.52 +/- 0.26 ml.min-1.g-1). o-R-poly-Hb increased Na reabsorption proportionately more than GFR, so that, while GFR was doubled to 1.04 +/- 0.40 ml.min-1.g-1, Na reabsorption increased to 98% (92-99.5%). HbAo increased GFR to 1.07 +/- 0.44 ml.min-1.g-1 and increased reabsorption to 89 +/- 6%.(ABSTRACT TRUNCATED AT 250 WORDS)

Arterial Blood Pressure Responses to Cell-free Hemoglobin Solutions and the Reaction with Nitric Oxide

“…Human hemoglobin cross-linked with bis- (3,5-dibromosalicyl) fumarate between the ␣ chains at lysine 99 (␣␣-Hb, 7.9 g/dl, 4.9 mM heme) was supplied as a gift from the U.S. Army, Blood Research Detachment, Walter Reed Army Institute for Research (8). Purified native human hemoglobin (HbA 0 , 9.0 g/dl, 5.5 mM heme) was supplied as a gift from Hemosol, Inc. Human hemoglobin cross-linked with trimesoyl tris (methyl phosphate) to make either a two-point intramolecular crosslink between the ␤ chains at lysine 82 (␤82-Hb, 6.8 g/dl, 4.3 mM heme) or a three-point cross-link between the ␤ chains at lysine 82 and valine 1 of one ␤ chain (Tm-Hb, 7.0 g/dl, 4.4 mM heme) were prepared as described by Kluger et al (9) and supplied as a gift from Hemosol, Inc. Human hemoglobin polymerized with ring-opened raffinose (o-R-polyHb, 9.4 g/dl, 5.9 mM heme) was also supplied as a gift from Hemosol, Inc. (10). Human hemoglobin modified by covalent attachment of pyridoxal-5Ј-phosphate and surface conjugation to ␣-carboxymethyl, -carboxymethoxypolyoxyethylene (PHP, 7.7 g/dl, 4.8 mM heme) was supplied as a gift from Apex Biosciences, Inc. (11).…”

Artificial Oxygen Carriers as Red Blood Cell Substitutes: A Selected Review and Current Status

“…For example, while unmodified Hb increased renal vascular resistance two fold and glomerular filtration rate (GFR, an indicator of renal function), by 58%, Hb‐raffimer had no deleterious effect on GFR, renal blood blow, or renal vascular resistance (55). Fractional GFR and reabsorption increased (67). It has also been shown that perfusion with a cell‐free HBOC better preserves or restores renal blood flow and function following an acute ischemic renal event (67–69).…”

“…Fractional GFR and reabsorption increased (67). It has also been shown that perfusion with a cell‐free HBOC better preserves or restores renal blood flow and function following an acute ischemic renal event (67–69).…”

O-Raffinose Crosslinking Substantially Ameliorates the Vasoconstrictive and Nitric-Oxide-Inactivating Effects of Unmodified Human Hemoglobin in the Rat

“…Unreacted aldehydes are simultaneously reduced to alcohols to prevent further crosslinking. Excess reagents and byproducts are removed by diafiltration and the final solution formulated in the oxygenated state, in Ringer's lactate (USP) for injection (24).…”

“…This study was designed to evaluate systemic and renal hemodynamic effects of a highly purified hemoglobin solution that has been chemically crosslinked with o-raffinose to form oligomers of hemoglobin with a weight-average molecular weight of 230,000 (24).…”